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A Simple Method for Estimating Evolutionary Rate of Base Substitutions Through Comparative Studies of Nucleotide Sequences
Abstract
Some simple formulae were obtained which enable us to estimate evolutionary distances in terms of the number of nucleotide substitutions (and, also, the evolutionary rates when the divergence times are known). In comparing a pair of nucleotide sequences, we distinguish two types of differences; if homologous sites are occupied by different nucleotide bases but both are purines or both pyrimidines, the difference is called type I (or "transition" type), while, if one of the two is a purine and the other is a pyrimidine, the difference is called type II (or "transversion" type). Letting P and Q be respectively the fractions of nucleotide sites showing type I and type II differences between two sequences compared, then the evolutionary distance per site is K = -(1/2) ln [(1-2P-Q) square root of 1-2Q]. The evolutionary rate per year is then given by k = K/(2T), where T is the time since the divergence of the two sequences. If only the third codon positions are compared, the synonymous component of the evolutionary base substitutions per site is estimated by K'S = -(1/2) ln (1-2P-Q). Also, formulae for standard errors were obtained. Some examples were worked out using reported globin sequences to show that synonymous substitutions occur at much higher rates than amino acid-altering substitutions in evolution.
... The sequences were BLAST searched (Table 2) and CLUSTAL W (Thompson et al., 1994) aligned for precise understanding of the phylogenetic status of the candidate specimen. The Maximum Likelihood (ML) phylogram (Kimura, 1980) and the genetic p-distance were constructed using the software MEGA 11. ...
... B. guerini and B. mccanni holds a robust phylogenetic relation, judged from a bootstrap (BS) value 84% (Fig. 1). Pertinently, previous species and the distant ones, the present study had employed the Kimura-two-parameter (K2P) model (Kimura, 1980). The results of comparison of the nucleotide sequences from the conserved regions (18S rRNA) of the candidate species (B. ...
River, Kanniyakumari using morphological and molecular tools.
Samples of B. cunicularis were collected from Kumba River flowing in Kilamalai,
Kanniyakumari district. The species identification was carried out through morphological keys constructed
out of a coding matrix and phylogenetic tree (Maximum Parsimony) using Mesquite and PAUP4 software. 18S
rRNA sequence was subjected to BLAST analysis, and the phylogenetic tree was constructed through
Maximum Likelihood method using MEGA 11 software. Pairwise genetic distance of the species (p-distance,p
= nd/n) was also assessed by comparing the K2P values involving phylogenetically close and distant
relatives.
Coding matrix prepared using the morphological keys, raised out of 28 distinctive characteristics of
B. cunicularis and comparison with its phylogenetic relatives, have brought in valuable information on status
of the species. These findings were further established by Maximum Likelihood Analysis, using the PCR
amplicons of 18S rRNA. The phylogram prepared out of the sequence clearly reveals the candidate species'
phylogenetic proximity to other members of the genus Barytelphusa. Further, the species' monophyletic
status (with a BS value of 81%) suggests its early divergence from its congeners. The K2P pairwise genetic 'p'
distance analysis (p = nd/n) of the 18S rRNA has helped us not only to further ascertain the extent of its genetic
identity with its congeners, but as well has clearly provided us with the valuable cues per precise identification
of the species.
Along with the morphological parameters, the present study, using molecular tools, provides
valuable information for precise identification of the commercially important freshwater crab.
Key words: Barytelphusa cunicularis, Crustacea, Freshwater crab, Gecarcinucidae, Phylogram, Taxonomy
... For phylogenetic reconstructions, two methodologies were followed, including neighbor joining and minimum evolution by using MEGA software version 7.0 18 (Kumar et al., 2016). To finalize the sequence divergences, we used Kimura two-parameter distances (Kimura, 1980), with 1000 bootstrap iterations (Felsenstein, 1985). ...
... The process began by configuring a bootstrap consensus tree derived from 1000 replicates, representing the evolutionary history of the taxa (Felsenstein 1985). Additionally, evolutionary distances were computed using the Kimura 2-parameter (K2P) method, measured in terms of base substitution numbers per site (Kimura 1980;Nishimaki and Sato 2019). The ME tree was searched using the Close-Neighbor-Interchange (CNI) algorithm at search level 2 (Nei and Kumar 2000), and the neighbor-joining algorithm was used to generate the initial tree (Saitou and Nei 1987;Mailund et al. 2006). ...
Ambarwati R, Rahayu DA. 2024. Revealing species identity of edible razor clam Cultellus sp. (Bivalvia: Pharidae) from Madura Island, Indonesia. Biodiversitas 25: 1505-1513. Razor clams have an essential role as commercial commodities. One of the poorly-known edible razor clams from Madura, Indonesia, has not been identified as the species. This research aimed to identify the edible razor clam (Bivalvia: Pharidae) species from Madura Island, Indonesia. The samples of razor clams (Cultellus sp.) were collected from Bangkalan, Madura Island, Indonesia. Identification was conducted based on morphological and molecular characterization. The foot tissues of three individuals were processed for DNA isolation, electrophoresis, amplification, and sequencing to obtain the sequences of COI gene. The morphological characters of razor clam shells were analyzed descriptively and quantitatively. Data were collected from the COI gene sequences of razor clam Cultellus sp. and descriptively analyzed using bioinformatics software. The morphological identification revealed razor clams (Cultellus sp) from Bangkalan, Madura, Indonesia, Cultellus subellipticus. The nucleotide base composition sequences of C. subellipticus A, B & C consisted of Thiamine (T), Cytosine (C), Adenine (A), and Guanine (G) with a mean of 18.25, 12.92, 23.42, and 46.39%. Maximum Likelihood (ML) and Minimum Evolution (ME) phylogenetic analysis was also conducted using the Kimura 2 Parameter model to establish one major clade on C. subellipticus from the gene bank and a different group with Cultellus attenuatus and one out-group significantly different from the Ensis macha. This finding was strengthened by molecular identification results, which indicated the similarity with C. subellipticus from GenBank, with very high BOLD System data between 98.10-98.86%.
... The maximum likelihood (ML) method was used for constructing phylogenetic tree with 100 bootstrap replicates in MEGA ver. 11 (Tamura et al., 2021) and genetic distances was determined using the Kimura 2-parameter (Kimura, 1980). The best-fit-model was a general time-reversible model (GTR) with a gamma distribution ( + G) using MEGA. ...
The first complete mitogenome sequence of the Red-tongue Pit Viper (Gloydius ussuriensis) from Korea was characterized using next-generation sequencing. The mitogenome is a circular molecule (17,209 bp) with a typical vertebrate mitogenome arrangement, which consists of 2 ribosomal RNA genes (rRNA), 22 transfer RNA genes (tRNA), two non-coding regions (D-loop), and 13 protein-coding genes (PCGs). The base composition of the mitogenome is 32.7% of A, 27.5% of C, 13.9% of G, and 25.9% of T, with a slight AT bias(58.6%). This phylogenetic analysis infers that G. ussuriensis is in the same group as the Chinese G.
ussuriensis (Accession No. KP262412) and is closely related to G. blomhoffi and other species of the genus Gloydius. In our study, the complete mitogenome sequence of Korean G. ussuriensis was characterized and we provided basic genetic information on this species.
... The transition/transversion ratio (R) was calculated, where R tends to be 0.5 when there is no bias towards either substitution. To evaluate the clustering further, the neighbour-joining (NJ) method was also employed using the K2P model substitution (Kimura 1980). ...
Saccharumspontaneum L. commonly known as wild sugarcane, is a close wild relative of sugarcane, representing the majority of terrestrial environments from the open ground to the shoreline. Due to its diverse applications as food, fodder, food ingredients, medicine, household products, pulp material, and even religious offerings, understanding the genetic relationships among populations is crucial. The present study attempted to understand the phylogenetic and evolutionary relationships of eight genotypes (six field samples and two commercial samples) from the chosen population of S. spontaneum in Tamil Nadu. The genomic material was extracted and amplified using nuclear ribosomal DNA internal transcribed spacer (nrDNA-ITS) and ribulose-1,5-bisphosphate carboxylase/oxygenase large subunit (rbcL) sequences and compared with those obtained from the NCBI database. The evolutionary history was inferred using MEGA11 software. The results showed that the presence of highly conserved sites in the rbcL region led to a close phylogenetic relationship falling under a common clade, which could be useful for molecular identification of this species. On the other hand, ITS markers were found to be useful for determining the phylogeny of S. spontaneum via both phenetic and cladistic approaches. The ITS region in this wild sugarcane has better discrimination ability despite the highly conserved sites with few rapidly evolving sites. Four of the collected genotypes showed a close relationship, along with two commercial genotypes, suggesting shared adaptations or a common geographic origin. Additionally, PTMTMC 002 and PTMTMC 008 were found to have diverged from the rest of the genotypes, exhibiting low genetic distance, indicating recent evolutionary divergence.
... The most simple one is the percentage of sites that differ between sequences, referred to as the "raw" difference. In evolutionary biology, the Kimura 2-parameter distance (referred to as "K80") [22] is more commonly used. It assumes that the probabilities for transition and transversion substitutions are different. ...
Image-based species identification could help scaling biodiversity monitoring to a global scale. Many challenges still need to be solved in order to implement these systems in real-world applications. A reliable image-based monitoring system must detect out-of-distribution (OOD) classes it has not been presented before. This is challenging especially with fine-grained classes. Emerging environmental monitoring techniques, DNA metabarcoding and eDNA, can help by providing information on OOD classes that are present in a sample. In this paper, we study if DNA barcodes can also support in finding the outlier images based on the outlier DNA sequence's similarity to the seen classes. We propose a re-ordering approach that can be easily applied on any pre-trained models and existing OOD detection methods. We experimentally show that the proposed approach improves taxonomic OOD detection compared to all common baselines. We also show that the method works thanks to a correlation between visual similarity and DNA barcode proximity. The code and data are available at https://github.com/mikkoim/dnaimg-ood.
... The bootstrap method was used to test the stability of the phylogenetic tree topology, using 1000 replicates overall. Utilizing the Kimura two-parameter method, the evolutionary distance was determined (Kimura 1980;Tamura et al. 2013). ...
A Gram stain-positive, non-spore-forming, non-motile, short-rod actinomyces strain GXQ1321 T was isolated from maritime surface sediments in Beihai, Guangxi Zhuang Autonomous Region, and a number of categorization studies were performed. Following a period of 72 hours of incubation at a temperature of 30°C within an actinomycetes culture medium, the colony was yellow, circular, smooth, central bulge, convex, opaque, with a 1.8-3.0 mm diameter. Chemotaxonomic studies revealed that the major menaquinone in strain GXQ1321 T is MK-8. The most prevalent cellular fatty acids were anteiso -C 19:0 (27.28%), anteiso -C 15:0 (18.97%), anteiso -C 17:0 (15.95%), and iso -C 16:0 (12.21%). The whole-cell sugars of the strain GXQ1321 T identified were rhamnose, xylose and glucose. Strain GXQ1321 T exhibited the presence of meso-diaminopimelic acid (m-DAP) as a distinctive cell-wall diamino acid, and the polar lipids were identified as diphosphatidylglycerol (DPG), three phosphoglycolipidsone, phosphatidylethanolamine (PE), one unknown phospholipid (UP) and one unknown glycolipid (UG). This strain had 69.6% DNA G + C content. Strain GXQ1321 T is classified as Brevibacterium based on its 16S rRNA gene sequence. It is closely related to Brevibacterium samyangense SST-8 T (96.77%) and Brevibacterium rongguiense 5221 T (96.32%). The results showed that the average nucleotide identity (ANI) values of GXQ1321 T and the above two strain tyoes were 73.91–77.14%, and the digital DNA-DNA hybridisation (dDDH) values were 15.3–21.1%. Based on the phylogenetic, chemotaxonomi and physiologicalc data, strain GXQ1321 T was considered to be a new species of the genus Brevibacterium , named Brevibacterium litoralis sp. nov, with the type strain GXQ1321 T (= MCCC 1K08964 T = KCTC 59167 T ).
... Out of the 46 specimens, 12 were sequenced in this study and the rest were obtained from GenBank (see Supp. file 1: Table S1). The substitution models selected according to ModelFinder were TVM+F+I+G4 (Posada 2003;Le et al. 2012) for the three codon positions of COI and 16S, and K2P+I (Kimura 1980) for H3, accounting for the three codon positions. ML and BI analyses yielded slightly different results for the concatenated alignment ( Regarding our results, we believe that the specimen Doris sp. ...
In the present study, a revision of the phylogeny and taxonomy of the family Dorididae is carried out focusing on the genus Doris Linnaeus, 1758. The type species D. verrucosa Linnaeus, 1758 and a blueish and yellow morphotype of D. ocelligera collected in different localities in the Mediterranean Sea and the NorthEast Atlantic were sequenced, as well as D. bertheloti and the elusive D. marmorata for the first time. The genetic markers include the cytochrome c oxidase subunit I, 16S rRNA, and histone 3. The phylogenetic results suggest that the genus Doris is paraphyletic, and D. ocelligera morphotypes separate into two species, as confirmed with species delimitation tests. To complement the phylogenetic evidence with morphoanatomical data, the dissection of two specimens of each morphotype is conducted. Significant differences in morphological traits such as body shape, colouration patterns, and mantle tubercles come to light, together with anatomical differences in the relative shape and size of the radular teeth and reproductive structures. Considering the modern and old descriptions of D. ocelligera, it is finally concluded that the blueish morphotype belongs to D. ocelligera. In contrast, the yellow morphotype responds to the actual synonym Aldisa berghi (Vayssière, 1901), which is resurrected here as Doris berghi comb. rest. Considering the broad phylogeny of the family, some systematic notes at the genus level are here provided.
... As shown in Figure 2 by using the maximum likelihood approach and the Kimura two-parameter model, the evolutionary history was determined [31]. A total of 1000 replicates were used to get a bootstrap consensus tree [32] which is taken to represent the evolutionary history of the taxa analyzed [32]. ...
Bacterial isolate which can produce acetic acid has been isolated from a coffee pulping process. Primary screening of acetic acid-producing bacteria was performed by using YPG medium. Then, positive isolates on YPG were further screened on GYC medium. It has been identified by using a molecular method by sequencing 16srRNA gene and was very closely related to Pseudomonas aeruginosa. Its optimum growth conditions have been optimized by design expert software by using the RSM method. Temperature, ethanol content, pH, and time of incubation of the isolate have been optimized. This Pseudomonas species was used to produce vinegar from barley malt wine. In this process, the commercial yeast strain, Saccharomyces cerevisiae, was used to produce ethanol from barley malt extract with 7.3 Brix degree. Malt wine with 3.49% ethanol content was obtained in 3 days of fermentation. For acetous fermentation by the bacterial isolate, barley malt wine with 5% ethanol content and 4.7 Brix was used as a substrate. Malt vinegar with a pH of 3.76 and 3.5% titratable acidity was obtained by using the flask method, and malt vinegar with a pH of 3.79 and 3.4% titratable acidity was obtained by using a bioreactor.
... Genetic distance between the species was calculated with Kimura-2 parameter model (Jin & Nei, 1990;Kimura, 1980), and the genetic tree was constructed for genera consisting of at least four species by maximum likelihood (ML) method with 803-976-bp region in the amplified 991-bp fragment using MEGA v. 7.0.26 (Kumar et al., 2016). ...
Genetic relationships were examined for a total of 90 individuals of 90 species or species var. from 13 genera of Malawian cichlids based on the sequences of an amplified 991‐bp fragment of the mtDNA control region (mtDNA‐CR). In the network analysis, no exclusive clades were made by all the members of any genera in this study. However, congeneric clades were observed by genera Buccochromis, Copadichromis, Protomelas, and Sciaenochromis, whereas no congeneric clades were observed by genera Mylochromis, Nimbochromis, and Otopharynx. In non‐mbuna, an Aulonocara–Lethrinops group was divided into two groups, and the mean genetic distance of the larger group was much lower from mbuna than from other non‐mbuna. Overall mean genetic distance within a genus was generally low in mbuna, whereas it was relatively high in non‐mbuna. In the genetic tree of each genus, two or more large clades were observed, and some clades, such as those of Aulonocara hansbaenschi and Aulonocara nyassae in genus Aulonocara, Lethrinops micrentodon and Lethrinops sp. “gold harbor” in Lethrinops, and Otopharynx ovatus and Otopharynx brooksi in Otopharynx, were very deeply differentiated. Besides, a mbuna species, Pseudotropheus crabro, was extremely deeply differentiated from other members of this genus. These results suggest a widespread morphological convergence across the taxa in parallel with deep genetic differentiation in the long evolutionary story and some possibility of generation of the species of Aulonocara–Lethrinops group by hybridization of small non‐mbuna and mbuna species. Furthermore, taxonomical reexamination is necessary based on a strong support by genetic connection.
... The percentage of replicate trees in which the associated taxa clustered together in the bootstrap test (500 replicates) are shown next to the branches 68 . The evolutionary distances were computed using the Kimura 2-parameter method 69 and are in the units of the number of base substitutions per site. All positions containing gaps and missing data were eliminated (complete deletion option). ...
Metabolites exploration of the ethyl acetate extract of Fusarium solani culture broth that was isolated from Euphorbia tirucalli root afforded five compounds; 4-hydroxybenzaldehyde (1), 4-hydroxybenzoic acid (2), tyrosol (3), azelaic acid (4), malic acid (5), and fusaric acid (6). Fungal extract as well as its metabolites were evaluated for their anti-inflammatory and anti-hyperpigmentation potential via in vitro cyclooxygenases and tyrosinase inhibition assays, respectively. Azelaic acid (4) exhibited powerful and selective COX-2 inhibition followed by fusaric acid (6) with IC50 values (2.21 ± 0.06 and 4.81 ± 0.14 μM, respectively). As well, azelaic acid (4) had the most impressive tyrosinase inhibitory effect with IC50 value of 8.75 ± 0.18 μM compared to kojic acid (IC50 = 9.27 ± 0.19 μM). Exclusive computational studies of azelaic acid and fusaric acid with COX-2 were in good accord with the in vitro results. Interestingly, this is the first time to investigate and report the potential of compounds 3–6 to inhibit cyclooxygenase enzymes. One of the most invasive forms of skin cancer is melanoma, a molecular docking study using a set of enzymes related to melanoma suggested pirin to be therapeutic target for azelaic acid and fusaric acid as a plausible mechanism for their anti-melanoma activity.
... The final dataset comprised 568 nucleotide-length sequences, which were aligned using the ClustalW algorithm. The evolutionary history was determined using the maximum likelihood method with the Kimura-2 parameter model [45], which is a widely accepted approach in the field. In order to account for potential variations in the speed of evolution among the sites, a discrete gamma distribution was also utilized. ...
In this study, yeasts from the gut of O. barnabita larvae were isolated and molecularly identified. It is worth noting that this research provides the first analysis of the gut yeast community in O. barnabita larvae in Lithuania, which is a significant contribution to the field. Two hermit-like L3-praepupa instars were collected from a decaying oak log in Lithuania. The isolation, morphology, biochemistry, and physiology of the yeast isolates were characterized using standards commonly employed in yeast taxonomy studies. The isolates were identified by sequencing the large subunit (26S) rDNA (D1/D2 domain of the LSU). All gut compartments were colonized by the yeast. A total of 45 yeast strains were obtained from the gut of both O. barnabita larvae, with 23 strains originating from Larva 1, 16 strains from Larva 2, and 6 strains from the galleries. According to our identification results of the 45 yeast strains, most of the species were related to Ascomycota, with most of them belonging to the Saccharomycetales order. Yeasts of the genera Candida, Debaryomyces, Meyerozyma, Priceomyces, Schwanniomyces, Spencermartinsiella, Trichomonascus, and Blastobotrys were present in gut of O. barnabita larvae. Species of the Trichosporonales order represented the Basidiomycota phylum.
... Phylogenetic relationships were inferred using the neighbor-joining (NJ) and maximum parsimony (MP) methods. NJ trees were constructed using MEGA X (Kumar et al., 2018) and the Kimura two-parameter model of base substitution (Kimura, 1980b) with 1,000 bootstrap repetitions (Felsenstein, 1985). MP tree topology was estimated using the tree bisection-reconnection algorithm. ...
Blue sprat (Spratelloides gracilis) is an economically valuable species that inhabits the central Taiwan Strait. However, despite the implementation of a local management scheme involving a closed season between 1 May and 1 June, the annual catch has rapidly decreased over the decades. Thus, the efficacy of the implemented regulations must be investigated. This study sought to clarify the reproductive biology, spawning ground, and behavior of this species in the waters of Penghu. In total, 6,549 specimens were collected between March 2021 and September 2022. Considering gonadosomatic index, group maturity, and oocyte diameter, we found that the spawning season (March–September) peaked in April; new cohorts were distinctly recruited after the spawning peak. The distribution of the spawning ground (latitude, 23°41′N to 23°43′N; longitude, 119°32′E to 119°39′E) was determined by considering the fishing area of mature females and DNA analysis of adhered eggs. We further noted shoals of fish darting in the bottom water without clear schooling movement, females’ abdomen frequently contacting substrata for eggs able to attach on during oviposition, and males releasing sperms making water milky white; these indicate that S. gracilis exhibits promiscuous spawning behavior. Our findings may facilitate the management and conservation of S. gracilis in this region.
... In addition, MEGA 11 were also used for constructing the Maximum-Likelihood phylogenetic tree (ML, 1000 bootstrap replications), and estimating the genetic distances within and among the populations. (Kimura, 1980;Kumar et al., 2016). The basic nucleotide configurations, nucleotide diversity (p), haplotype diversity (h), haplotype number, fixation index (FST, 100 permutations for significance, and 1000 permutations for mantel test), gene flow [Nm, calculated by the formula Nm = (1-FST)/2FST], Tajima's D and Fu's Fs test values were analyzed by Arlequin 3.5 (Tajima, 1989;Fu, 1997;Excoffier and Lischer, 2010). ...
The Pseudaspius leptocephalus is a unique fish in the Heilongjiang River Basin and has important economic and ecological value. In the present study, the complete mitochondrial genome of P. leptocephalus were determined, and COI partial sequences of 85 individuals from Erguna river (EH), Mohe (MH), Fuyuan (FY), Hulan (HL) were used to evaluated the genetic diversity of four populations of P. leptocephalus in Heilongjiang River Basin. The mitogenome is 16,607 bp in length and contained one D-loop, 2 rRNA, 13 PCG, and 22 tRNA. 4 variable sites and 5 haplotypes were detected in 705 bp COI, and 705 bp COI exhibited a lower content of C + G (45.95%) than A + T (54.05%). The nucleotide diversity (π) and haplotype diversity (h) indices ranged from 0.00027 (HL) to 0.00065 (EH and FY) and from 0.192 (HL) to 0.462 (EH), respectively. The genetic distance within the population and between populations ranged from 0.0006554 to 0.0002728 and from 0.0003541 to 0.0006974, respectively. Pairwise values of FST and Nm showed that there was moderate genetic differentiation between EH population and other populations and individuals between EH population and other populations can mate randomly (0.15 > FST > 0.05, Nm > 4). Significant negative values of neutrality tests (P < 0.05) indicated that MH and FY populations may had experienced population expansion, but mismatch distribution analysis suggested that all populations have remained basically stable. These results provide strong basis for the protection and utilization of P. leptocephalus germplasm resources, and provide valuable information for the population structure and genetic diversity of P. leptocephalus.
... DNA sequence data was matched to databases available at the National Center for Biotechnological Information (NCBI) online (www.ncbi.nml.nih.gov) and the Barcode of Life Data System (BOLD System). Phylogenetic analysis was done based on the Neighbor Joining (NJ) method (Saitou and Masatoshi, 1987) and the Kimura 2parameter model (K2P) (Kimura, 1980) using Mega XI software (Tamura et al., 2021). ...
Environmental and genetic variables can exert an influence on alterations in morphological traits. Within fish species inhabiting diverse aquatic settings, there can be observed variations in morphological traits. Genetically, variations in fish morphological characteristics can occur through mating and gene flow. To date, there has been a lack of research conducted on the variability in morphological traits and genetic relationships between Lutjanus timoriensis, L. malabaricus, and L. erythropterus. Thus, the current research aimed to identify variations in the morphological characteristics as well as in the intra- and inter-specific relationships between three red snapper species from the genus Lutjanus. Cytochrome oxidase I (COI) gene was used to study the molecular relationship among species of red snapper. The results showed that L. timoriensis had high intraspecific morphological variation in young individuals. Morphologically, L. timoriensis, L. malabaricus, and L. erythropterus are very similar. Unlike L. malabaricus and L. erythropterus, young and adult L. timoriensis have black patches in the pectoral fin axils. The adult L. erythropterus has a comparatively small mouth, no hump on its head, and no black saddle at the base of its tail. Meanwhile, L. malabaricus has a comparatively large mouth, a head with a hump, and a black saddle at the base of its tail. In terms of body size, L. erythropterus is larger than L. timoriensis and L. malabaricus. Based on NCBI and Bold System data, molecular analyses determined that the observed fish were L. timoriensis, L. malabaricus, L. erythropterus with a similarity of between 99.85 and 100%. The phylogenetic tree construction demonstrated that L. malabaricus, L. timoriensis and L. erythropterus were closely related.
... Branch support was assessed using the ultrafast bootstrap analysis (Hoang et al., 2018) based on 1000 bootstrap alignments (Felsenstein, 1985), while default settings were used for all other parameters. The resulting tree was visualized in MEGA 11 (Tamura et al., 2021), and this program was also used to calculate Kimura 2-parameter distances (Kimura, 1980) between selected sequences. A minimum-spanning haplotype network (Bandelt et al., 1999) was constructed between the species comprising a monophyletic cluster in the maximum likelihood tree, using popArt (Leigh & Bryant, 2015). ...
... Taxonomic annotation was done using NCBI-nt database. Homologous sequences were obtained from the GenBank database of NCBI and the phylogenetic tree was inferred using the Maximum Likelihood method based on the Kimura 2-parameter model (Kimura, 1980) with the aid of MEGA XI (Tamura et al., 2021). The bootstrap consensus tree was inferred from 1000 replicates (Felsenstein, 1985) to represent the evolutionary history of the taxa analyzed. ...
The Paper Nautilus, Argonauta hians Lightfoot 1786, belongs to the superfamily Argonautoidea and is widely distributed along the tropical and subtropical oceans. The present study describes the record of A. hians recorded from the gut of a long-snouted lancetfish, Alepisaurus ferox (133cm TL and 2 kg weight). A. ferox was hooked during the regular multifilament tuna longlining onboard MFV Yellow Fin on 20 th October 2021 off Goa (Lat. 15º 19.44 N; Long. 70º 50.20 E) at 2652 m depth along the Central west coast of India. A detailed note on the distribution of A. hians in Indian waters is provided along with morphological characterization of the specimen and validation with molecular marker.
... Additionally, the variations among species were assessed by averaging pairwise comparisons, and the count of haplotypes was evaluated using DnaSp 5.10 [20]. Utilizing the Kimura 2-parameter (K2P) model, and Neighbors Joining (NJ) trees were constructed with the assistance of MEGA 11 [21,22]. ...
Objective
Channidae family, are major freshwater fish species amongst the local aquatic fauna of Pakistan, while, there is limited availability of local data on their molecular identification and phylogenetic analysis.
Methods
Channa species were collected from four different geographical sites in the tertiary of Punjab province on the Indus and Chenab rivers of Pakistan. Morphometric records and molecular techniques were used to determine the intraspecific variations among populations of Channa marulius. Mitochondrial DNA was extracted from the flesh of C. marulius, while, COI gene was used for molecular identification and variation levels were estimated by using Principal Component Analysis.
Results
Data recorded on the basis of morphometric parameters clearly divided the C. marulius of different locations into two distinct categories, which accounted for a cumulative variability of 97.6%. Non-significance (P < 0.05) among the C. marulius showed that it contains a unique control haplotype localized within the sub-population. The intra-species distance ranged from 0.000 to 0.001 for four different populations, in contrast, the sequences retrieved from the NCBI database exhibited a range span of 0.000–0.003, while, sequence diversity ranged from 0.000 to 0.006 for this intra-specific comparison. The cladogram was also constructed for C. marulius of different geographical locations for observation of phylogenetic relationship. The conclusion drawn from the phylogenetic analysis of C. marulius populations used in this study, contributes significantly to the understanding of genetic variations within populations of this species. The findings provide valuable insight to devise conservation strategies in fisheries management programs in Pakistan.
... The first and second codon positions of the RAG genes were assigned the general time reversible (GTR) model (Tavar e, 1986), as were the first and second codon positions for the mitochondrial genes. The third codon position of the RAG and mitochondrial genes were assigned a Kimura two-parameter (K2P) model, which indicates varying transition or transversion rates and equal base frequencies (Kimura, 1980). All partitions included a mixture model of rate heterogeneity across sites (gamma model). ...
... using the default values to compute distance calculation for all three available substitution models: Jukes-Cantor (JC69), Kimura (K80) ts/tv and simple p-distances. Nucleotide divergence, representing genetic distances between and within the lineages identified in the different species delimitation analyses, was calculated using Kimura's two-parameter (K2P) model (Kimura 1980), including transitions and transversions, with uniform rates among sites, a pairwise deletion of gaps and 500 bootstrap replicates to estimate the variance on the COI and 16S sequences using MEGA X (ver. 10.2.6, see https://www.megasoftware.net/ ...
Syllis prolifera (Syllidae, Syllinae) is an abundant species of marine annelids commonly found in warm to temperate waters worldwide. Although morphological variability occurs among populations , S. prolifera has long been considered a cosmopolitan species, widely distributed in coastal environments, including acidified and polluted areas. However, the increasing number of cases of cryptic and pseudocryptic speciation in several polychaete families in recent years has led us to question whether S. prolifera represents a single globally distributed taxon or is a species complex. To address this question, we conducted an integrative study, combining morphological, ecological and molecular data of 52 S. prolifera specimens collected in different localities across the western Mediterranean Sea and the Gulf of Cadiz. Our phylogenetic and species delimitation analyses that included two mitochondrial DNA markers (COI and 16S rRNA) were congruent in not considering S. prolifera a unique entity. Five distinct lineages that can also be recognised by certain morphological and ecological traits were identified from these analyses instead. Overall, our study does not support the homogeneity of S. prolifera across the Mediterranean Sea, providing a new example of pseudocrypticism in marine invertebrates.
... and then the pairwise comparisons were performed using the NCBI (www.ncbi.nlm.nih.gov) and EzBioCloud databases (www.ezbiocloud.net). Phylogenetic analyses of the two strains and other type species were performed by mega X [12] with three tree-making sets of computer instructions, including the neighbourjoining algorithm [13] with Kimura's two-parameter model [14], the maximum-likelihood algorithm [15] with the Tamura-Nei model and the maximum-parsimony algorithm [16] based on 1000 bootstrap replications. ...
Two Gram-stain-positive, aerobic, oxidase-and catalase-negative, non-motile, and short rod-shaped actinomycetes, named SYSU T00b441 T and SYSU T00b490, were isolated from tidal flat sediment located in Guangdong province, PR China. The 16S rRNA gene sequence similarity, average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values between SYSU T00b441 T and SYSU T00b490 were 99.3, 99.5 and 97.1 %, respectively. Strains SYSU T00b441 T and SYSU T00b490 exhibited the highest 16S rRNA gene sequence similarities to Actinotalea ferrariae CF 5-4 T (97.1 %/98.2 %), with ANI values of 74.01/73.88 % and dDDH values of 20.5/20.4 %. In the phylogenomic tree, the two isolates were affiliated with the genus Acti-notalea. The genomes of strains SYSU T00b441 T and SYSU T00b490 were 3.31 and 3.34 Mb, and both had DNA G+C contents of 72.8 mol%, coding 3077 and 3085 CDSs, three and three rRNA genes, and 53 and 51 tRNAs, respectively. Growth occurred at 15-40 °C (optimum, 28-30 °C), pH 4.0-10.0 (optimum, 7.0) and in the presence of 0-7 % (w/v) NaCl (optimum, 3 %). The major fatty acids (>10 %) of strains SYSU T00b441 T and SYSU T00b490 were anteiso-C 15 : 0 and C 16 : 0. The major respiratory quinone was identified as MK-10(H 4). The polar lipids of strains SYSU T00b441 T and SYSU T00b490 were diphosphatidyl glycerol, phos-phatidylglycerol, phosphoglycolipid, phosphatidyl ethanolamine, two phosphatidylinositol mannosides, two glycolipids and two phospholipids. Based on these data, the two strains (SYSU T00b441 T and SYSU T00b490) represent a novel species of the genus Actinotalea, for which the name Actinotalea lenta sp. nov is proposed. The type strain is SYSU T00b441 T (=GDMCC 1.3827 T =KCTC 49943 T).
... A majority consensus tree was generated to obtain the final topology after discarding 25% of the initial trees. The Kimura-2-parameter evolutionary model (K2P; Kimura, 1980) was utilised, with complete gap deletion, to calculate the genetic distances both within and between species using the MEGA X software (Kumar et al., 2018). ...
Aspects of the biology and natural history of the endemic anuran Crossodactylus dantei remain unknown even almost 30 years since its description. In this study, we describe its larval stage, along with the advertisement and territorial calls, and assess its phylogenetic positioning based on a fragment of the 16S rRNA mitochondrial gene. The larval stage of C. dantei follows the morphological pattern of Hylodidae, corroborating the relatively uniform larval morphology of the family. However, it is still possible to observe characters that distinguish C. dantei from other species in the genus, such as the elliptical body in dorsal view, nostrils located closer to the snout than to the eyes, an elongated pre-nasal arena, and the absence of taenia tecti medialis and taenia tecti transversalis in the chondrocranium. Its advertisement call is characterised by a sequence of about 23 pulsed notes, with ascending amplitude modulation, and has a mean duration of 2.5 seconds, with a dominant frequency of 3356 Hz. The territorial call has a mean duration of 23 seconds, a dominant frequency of 3160 Hz, and two distinct types of notes. Phylogenetic analysis indicated the existence of two main lineages in the genus. Crossodactylus dantei was recovered with high statistical support as a sister species to the clade composed of C. gaudichaudii and C. timbuhy. Furthermore, the group formed by these three species was recovered as the sister lineage of C. trachystomus. In summary, the information presented here represents a significant advance in the knowledge of this enigmatic and rare species of the northern Atlantic Forest, providing baseline data for future ecological and evolutionary studies of this unique frog.
... The percentage of replicate trees, in which the associated taxa clustered together in the bootstrap test (1000 replicates), was shown next to the branches [15]. The evolutionary distances were computed using the Kimura 2-parameter method [16], and measured as the number of base substitutions per site. All positions containing gaps and missing data were eliminated (complete deletion option). ...
Berberine (BBR) is widely used as a botanical pesticide due to its broad-spectrum antibacterial and antifungal activities. However, BBR degradation pathway in soil microorganisms, which determines its impact on soil environment, remains poorly understood. Herein, a novel BBR-degrading bacterium Agrobacterium sp. V1 was isolated and characterized. Agrobacterium sp. V1 was able to utilize BBR as the sole carbon source for cell growth, and 50 μg/mL of BBR was completely degraded within 48 h. To reveal the possible BBR degradation pathway, whole genome sequencing of Agrobacterium sp. V1 was conducted, and proteins in Agrobacterium sp. V1 were aligned with enzymes involved in BBR biosynthesis in Rhizoma Coptidis. The results indicated that more than 60% of enzymes in BBR biosynthesis pathway had orthologs in Agrobacterium sp. V1. Combined with the primary mass spectra of BBR metabolites, a novel BBR degradation pathway in this bacterium was proposed. In summary, the proposed BBR degradation pathway offered new insights into the impact of BBR to the environment and also provided a reference for studying BBR metabolism in microorganisms.
... The alignment of 13 protein-coding genes (PCGs) was based on amino acid sequences translated with the invertebrate mitochondrial genetic codon (Folmer et al. 1994;Hebert et al. 2003aHebert et al. , 2003b were calculated using MEGA (ver. 7.0; Kumar et al. 2016) under the Kimura 2 Parameter (K2P) model (Kimura 1980). ...
The antlion genera Gatzara and Nepsalus (Myrmeleontidae: Dendroleontinae) inhabit mountain forests and are characterised by camouflaging larvae. Both genera remain poorly known despite recent findings on systematics and distribution. We report the discovery of new specimens and the previously unknown larvae of the rare species Gatzara jubilaea Navás, 1915, Nepsalus insolitus (Walker, 1860) and N. decorosus (Yang, 1988). These provide new evidence regarding the affinities of these species, and updated knowledge of the distribution, larval morphology and biology. Moreover, a new species of Nepsalus, N. maclachlani Badano, Zheng & Liu, sp. nov. is described from Sri Lanka based on historical museum collections. The discovery of the immature stages of Gatzara shows that the larvae of this genus share the same specialised ecological characteristics and habits as those of Nepsalus but are less morphologically derived. We also reconstruct a molecular phylogeny of this lineage, estimating the divergence time and biogeographical history by adding the new samples. The evolution of the Gatzara + Nepsalus lineage is associated with two major mountain ranges on the southern Tibetan Plateau, i.e. the Himalayas and the Hengduan Mountains. ZooBank: urn:lsid:zoobank.org:pub:68E68211-DFC1-4D98-997B-8A23BA8F9B69
... The chain length was set to 5,000,000 generations, with sampling every 2,000 generations. Pairwise genetic distances of the cytb gene sequences were estimated with the help of the Kimura-2 parameter model (Kimura 1980) implemented in MEGA7 (Kumar et al. 2016). ...
The Ethiopian highlands represent a wide spectrum of ecological gradients that provide suitable conditions for gradient speciation. Previous studies support the gradient model of speciation for two Ethiopian shrew species: Crocidura thalia and C. glassi. Here, we aimed to elucidate for the first time the phylogenetic position of C. afeworkbekelei and to test the gradient model of speciation for these three species. On the basis of a dataset collected from the whole south slope of the Bale Mountains, we reconstructed phylogenetic relationships among these species using mitochondrial and nuclear markers. Additionally, we examined shape and size differentiation of the skull and mandible. The molecular data revealed a similarity of the three species with lack of reciprocal monophyly among them. We demonstrated differences both in size and shape of the skull and mandible between low-and high-elevation forms albeit without a significant morphological hiatus. We identified the most changeable parts of the skull and mandible, which imply adaptive shifts in diet. We revealed the distribution, phylogenetic and morphological patterns that match predictions of the gradient model of speciation for three mammalian forms. Our data suggest intense processes of adaptation to the markedly different habitats along the considerable altitudinal gradient that fit the first stage of the gradient model of speciation. We believe that C. afeworkbekelei and C. thalia should be regarded as different ecotypes, and these species names must be reduced to junior synonyms of C. glassi.
... ML trees were constructed in Mega7 (Kumar et al., 2018). For the beta subfamily, the best model was Kimura two-parameter + G + I, and for the gamma subfamily the best model was Hasegawa-Kishino-Yano + G + I (Hasegawa et al., 1985;Kimura, 1980). Node support was assessed using 500 bootstraps. ...
Public health concerns about recent viral epidemics have motivated researchers to seek novel ways to understand pathogen infection in native, wildlife hosts. With its deep history of tools and perspectives for understanding the abundance and distribution of organisms, ecology can shed new light on viral infection dynamics. However, datasets allowing deep explorations of viral communities from an ecological perspective are lacking. We sampled 1086 bats from two, adjacent Puerto Rican caves and tested them for infection by herpesviruses, resulting in 3131 short, viral sequences. Using percent identity of nucleotides and a machine learning algorithm (affinity propagation), we categorized herpesviruses into 43 operational taxonomic units (OTUs) to be used in place of species in subsequent ecological analyses. Herpesvirus metacommunities demonstrated long‐tailed rank frequency distributions at all analyzed levels of host organization (i.e., individual, population, and community). Although 13 herpesvirus OTUs were detected in more than one host species, OTUs generally exhibited host specificity by infecting a single core host species at a significantly higher prevalence than in all satellite species combined. We describe the natural history of herpesvirus metacommunities in Puerto Rican bats and suggest that viruses follow the general law that communities comprise few common and many rare species. To guide future efforts in the field of viral ecology, hypotheses are presented regarding mechanisms that contribute to these patterns.
... Sequence readings were compared with sequences of reported isolates using BLAST in GenBank. A maximum likelihood phylogenetic tree was constructed using MEGA X software [38], with bootstrap values estimated using 1,000 replicates based on Kimura's two-parameter substitution model [39]. ...
Background
Tick-borne diseases cause economically significant losses to animal production globally, and anaplasmosis and theileriosis are associated with the greatest losses. However, the spread of the relevant pathogens in flocks of domesticated animals in southern Egypt is little understood. Accordingly, in this study, we aimed to determine the prevalences of Anaplasma ovis, Theileria ovis, and Theileria lestoquardi in southern Egyptian sheep and goats through blood tests, and to make a molecular characterization of the A. ovis detected in sheep targeting a specific gene.
Results
We collected blood samples collected from 300 sheep and goats (n=150 /species) in Luxor Province in southern Egypt, and analyzed them for the presence of A. ovis, T. ovis and T. lestoquardi with screening by conventional and nested PCR targeting the msp4 and msp5, 18S rRNA, and merozoite surface protein genes. For A. ovis 140/300 samples (46.66%) were positive overall, with 90/150 (60%) and 50/150 (33.33%) positive samples in sheep and goats, respectively. Two major surface protein genes of A. ovis, msp4 and msp5, were sequenced using DNA extracted from sheep and goat blood samples, for phylogenetic analysis and genotyping. The msp4 gene sequence revealed no significant genetic diversity, to contrast to data on A. ovis strains from other countries. For T. lestoquardi, 8/150 (5.33%) samples were positive in sheep, but no samples were positive in goats (0%). For T. ovis, 32/150 (21.33%) samples were positive in sheep, but no samples were positive in goats (0%). Sequencing targeting the merozoite surface protein gene for T. lestoquardi and the small subunit ribosomal RNA gene for T. ovis revealed no significant genetic diversity in the study, another contrast to data on A. ovis strains from other countries.
Conclusion
This study provides valuable data on phylogenetic and molecular classifications of A. ovis, T. ovis and T. lestoquardi found in southern Egyptian sheep and goats. It also represents the first report on detection and molecular characterization of T. lestoquardi in southern Egyptian sheep based on the specific merozoite surface protein gene, thus providing valuable data for molecular characterization of this pathogen in southern Egypt.
... The new sequences were aligned with MAFFT 7.490 (Katoh & Standley, 2013) to ITS and Cox1 sequences from type or authentic strains of ingroup and outgroup species retrieved from Gen-Bank. For each DNA barcode sequence, a neighbour-joining (NJ) tree was constructed using the Kimura-2-parameter model (Kimura, 1980) with 1000 bootstrap replicates in PAUP 4.0b10 (Swofford, 2003). The resulting trees were used to determine relationships among barcode sequences based on sequence similarities, rather than for phylogenetic reconstruction. ...
Pythium sensu stricto (s.s.) and Globisporangium species are important components of the soil microbial community and exhibit diverse lifestyles, including mycoparasitism. However, a comprehensive understanding of the species diversity of these mycoparasites in the West Azarbaijan province of Iran is lacking. In this study, a total of 114 mycoparasitic Pythium s.s. and Globisporangium isolates were obtained from agricultural soils collected from six regions in the province. Through DNA barcoding, all Globisporangium isolates were identified as G. nunn, while the barcode markers were insufficient to accurately resolve species boundaries in Pythium s.s. By combining morphological and multilocus sequence data, five species within the genus Pythium s.s. were identified: P. salmasense sp. nov., a potentially new species, and three known species, P. acanthicum, P. ornamentatum, and P. periplocum. Pythium ornamentatum was the most common species and found in all regions studied, followed by G. nunn and P. acanthicum, which were both isolated from four regions. While the isolates of G. nunn showed no mycoparasitic activity against Sclerotinia sclerotiorum, all Pythium s.s. species were capable of infecting the hyphae of this pathogen. The existence of mycoparasitic species is promising for biological control of soil‐borne fungal pathogens in the province. The widespread occurrence of P. ornamentatum, G. nunn, and P. acanthicum may suggest their adaptation to local soil and environmental conditions, indicating their potentially superior effectiveness in controlling plant diseases across different regions if used as biocontrol agents.
... Acanthomyrmex careoscrobis Mofett, 1986, was used as the outgroup [26]. Te distance-based neighbor joining [27] method was used to infer the relationship between our query sequences and those downloaded from online repositories using the Kimura 2-parameter model [28]and 1000 bootstrap replicates. Analyses were performed with MEGA v7 [29]. ...
The genus Myrmecina was described basing on males of M. graminicola. Though representing the caste type, males were sufficiently described in only two out of 105 species known in the genus. However, the morphology of the male external genitalia remained undescribed. Re-examining by SEM the male of M. graminicola, we describe and illustrate in detail for the first time the external genitalia, redefining and updating the morphological male diagnosis of the genus. We also analyze the overall morphology, illustrating additional peculiar characters of this species as follows: (i) a very distinctive stipital groove in the dorsolateral stipes; (ii) a developed uncinate-shaped mesoscutellar arm; (iii) the antennal cleaning; and (iv) the absence of meso- and metatibial spurs. This morphological study will be useful as a base for further morphological descriptions of the males in other species of the same genus to support correct taxonomic identifications.
... Two molecular species delimitation analyses were conducted to test the congruence between molecular and morphological datasets (external and internal anatomical characters). The Bayesian Poisson tree processes (bPTP; Zhang et al., 2013) were implemented via a non-ultrametric best maximum likelihood (ML) tree estimated in IQ-TREE web server (http:// iqtree.cibiv.univie.ac.at/) (Trifinopoulos et al., 2016) under an Ultrafast Bootstrap analysis (Hoang et al., 2018) fr/abi/public/asap/). Additionally, we estimated pair-wise genetic distances for COI matrix in Mega 11 (Tamura et al., 2021) using Kimura 2-parameter model, K2P (Kimura, 1980). ...
A new species of Eigenmannia is described from the Rio Branco basin, Roraima, Brazil, based on morphological and molecular datasets. It is distinguished from all congeners by the following combination of characters: lateral line stripe extending from first perforated lateral line scale to distal portion of caudal filament, presence of superior midlateral stripe with origin posterior to end of body cavity anal‐fin hyaline, caudal filament corresponding to 15.2%–43.1% LEA, subterminal mouth, ii,14–16 pectoral‐fin rays, 166–219 anal‐fin rays, 10–13 scale rows above lateral line at vertical through posterior tip of pectoral fin, 100–128 scales on lateral line, 22–28 premaxillary teeth, 19–23 dentary teeth, 7–10 endopterygoid teeth, depth of posterodorsal expansion on infraorbitals 1 + 2 half as long as infraorbitals 1 + 2 length, basibranchial 1 unossified, 13 precaudal vertebrae, and length of coronomeckelian bone corresponding to 20% of Meckel's cartilage length. The new species has significant genetic divergence from species with accessible DNA sequences in public repositories, ranging from 10.8% to 17.7%. An osteological description of the new species, a review of Eigenmannia cytochrome c oxidase subunit I (COI) sequences available in public repositories based on voucher examination, and a hypothesis of phylogenetic relationships for the new species based on COI are provided. The critical importance of including voucher examination as one of the steps in the pipeline for using DNA sequences present in public repositories in taxonomic and phylogenetic studies is discussed.
Nepal holds a high ichthyofaunal diversity, nevertheless, the molecular study of Nepalese fish is still in its early stages. The first record of a cyprinid fish species the Garra kempi Hora, 1921 for Nepal was reported from eastern Nepal’s Lohandra River. Both morphology and molecular data affirmed the existence of G. kempi in Nepal’s aquatic system. This species was previously reported from China (Tibet) and northern India. The maximum likelihood phylogenetic analysis and pairwise genetic distance based on Kimura 2 parameters using cytochrome oxidase subunit 1 (COI) segment sequences (665 bp) also confirmed the identity of the species as G. kempi. The COI sequences of specimens from Nepal formed a monophyletic clade with the sequence of G. kempi from northeastern India and the two contained a pairwise genetic distance of 1.8% only. The new record of G. kempi from Nepal warrants a detailed ichthyofaunal survey for documenting the fish diversity in Nepal.
Okra (Abelmoschus esculentus) is a nutritious domesticated “fruit” that belongs to the Malvaceae family. India esteems to be the largest global producer of lady’s finger and its net annual output accounts for 60,00,000 metric tons. In this investigation, a peculiar kind of leaf spot disease was observed on a diseased leaf sample collected from Howrah, Kamalchowk, West Bengal, India (Latitude: N 22° 59.5770′, Longitude: E 88° 26.3641′). Large brownish spots mottled the leaves, with spots also lining the edges of the leaf. With disease progression, these lesions expanded to form patches, leading to necrosis and defoliation. Fungus isolated from infected plants when cultured showed golden to deep brown mycelial growth. Microscopic characterization revealed ovoid, muriform, or club-shaped multiseptated conidia, often existing in chains, with the majority tapering towards a swollen short beak. ITS sequencing rendered the identity of the infectious agent to be Alternaria aungustiovoidea. Phylogenetic results indicated that this pathogen was closely related to Alternaria arborescens, A. tenuissima, and Fusarium equiseti. Koch’s postulates were performed and ratified, where distinct symptoms were seen in okra. Alternaria species, including Alternaria solani cause early blight in tomatoes and potatoes, and Alternaria alternata are often associated with citrus fruit rot, induce plant diseases. Their intrusions are not restricted to plants as exposure to A. alternata and A. tenuissima have been associated with allergies, cutaneous alternariosis, and respiratory conditions in humans. Therefore, the significance of tracing such pathogens and thinking of preventive measures can never be undermined. No previous accounts were found, hence one can claim this is the first time such a peculiar host–pathogen combination scenario has been encased and presented in India.
Among the Palearctic species, butterflies of the genus Melanargia are known for their black and white wing patterns. The morphological character polarization of this genus is full of varying combinations of subgenus, species complex, and subspecies status. Its taxonomy is open to debate, especially in species and subspecies categories, with definitions mostly based on wing color. In recent years, cryptic species, phenotypically masked species, and species with intense intraspecific variation have been identified through the determination of lineages under the leadership of molecular systematics. The mtCOI gene, which is especially described as a species signature, is an important DNA barcode used for Lepidoptera. In the presented study, the mtCOI gene sequence of the populations of Melanargia larissa, M.grumi, M.hylata, M.syriaca, and M.russiae species in the southeastern Anatolia region was determined for the first time. To determine the boundaries of these species, gene characterization and genetic distances were carried out according to the Kimura-2 Parameter, and putative species analyses were carried out by the ABGD method. Trees were constructed with Maximum likelihood and Bayesian inference algorithms to determine the phylogenetic relationships between species of the genus. In light of these analyses, it has been shown that the genetic distance of morphological species M. larissa, M. grumi, M.hylata, and M. syriaca is not at the species level and that M.larissa maintains its species status according to the principle of priority. In addition, the M.russiae population presented in this study forms a monophyletic clade with other populations of the same species in the phylogenetic tree, proving that this taxon is a stable species.
A novel chemoheterotrophic iron-reducing micro-organism, designated as strain LSZ-M11000 T , was isolated from sediment of the Marianas Trench. Phylogenetic analysis based on the 16S rRNA gene revealed that strain LSZ-M11000 T belonged to genus Tepidibacillus , with 97 % identity to that of Tepidibacillus fermentans STGH T , a mesophilic bacterium isolated from the Severo-Stavropolskoye underground gas storage facility in Russia. The polar lipid profile of strain LSZ-M11000 T consisted of diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, as well as other unidentified phospholipids and lipids. The major fatty acids were C 16 : 0 (28.4 %), C 18 : 0 (15.8 %), iso-C 15 : 0 (12.9 %), and anteiso-C 15 : 0 (12.0 %). Strain LSZ-M11000 T had no menaquinone. Genome sequencing revealed that the genome size of strain LSZ-M11000 T was 2.97 Mb and the DNA G+C content was 37.9 mol%. The average nucleotide identity values between strain LSZ-M11000 T and its close phylogenetic relatives, Tepidibacillus fermentans STGH T and Tepidibacillus decaturensis Z9 T , were 76.4 and 72.6 %, respectively. The corresponding DNA–DNA hybridization estimates were 20.9 and 23.4 %, respectively. Cells of strain LSZ-M11000 T were rod-shaped (1.0–1.5×0.3–0.5 µm). Using pyruvate as an electron donor, it was capable of reducing KMnO 4 , MnO 2 , As(V), NaNO 3 , NaNO 2 , Na 2 SO 4 , Na 2 S 2 O 3 , and K 2 Cr 2 O 7 . Based on phenotypic, genotypic, and phylogenetic evidence, strain LSZ-M11000 T is proposed to be a novel strain of the genus Tepidibacillus , for which the name Tepdibacillus marianensis is proposed. The type strain is LSZ-M11000 T (=CCAM 1008 T =JCM 39431 T ).
One Gram-negative, rod-shaped bacterial strain, isolated from an undescribed Heterorhabditis entomopathogenic nematode species was characterized to determine its taxonomic position. The 16S rRNA gene sequences indicate that it belongs to the class Gammaproteobacteria, to the family Morganellaceae, to the genus Photorhabdus, and likely represents a novel bacterial species. This strain, designated here as CRI-LCT, was therefore molecularly, biochemically, and morphologically characterized to describe the novel bacterial species. Phylogenetic reconstructions using 16S rRNA gene sequences show that CRI-LCT is closely related to P. laumondii subsp. laumondii TT01T and to P. laumondii subsp. clarkei BOJ-47T. The 16rRNA gene sequences between CRI-LCT and P. laumondii subsp. laumondii TT01T are 99.1% identical, and between CRI-LCT and P. laumondii subsp. clarkei BOJ-47T are 99.2% identical. Phylogenetic reconstructions using whole genome sequences show that CRI-LCT is closely related to P. laumondii subsp. laumondii TT01T and to P. laumondii subsp. clarkei BOJ-47T. Moreover, digital DNA-DNA hybridization (dDDH) values between CRI-LCT and its two relative species P. laumondii subsp. laumondii TT01T and P. laumondii subsp. clarkei BOJ-47T are 65% and 63%, respectively. In addition, we observed that average nucleotide identity (ANI) values between CRI-LCT and its two relative species P. laumondii subsp. laumondii TT01T and P. laumondii subsp. clarkei BOJ-47T are 95.8% and 95.5%, respectively. These values are below the 70% dDDH and the 95–96% ANI divergence thresholds that delimits prokaryotic species. Based on these genomic divergence values, and the phylogenomic separation, we conclude that CRI-LCT represents a novel bacterial species, for which we propose the name Photorhabdus africana sp. nov. with CRI-LCT (= CCM 9390T = CCOS 2112T) as the type strain. The following biochemical tests allow to differentiate P. africana sp. nov. CRI-LCT from other species of the genus, including its more closely related taxa: β-Galactosidase, citrate utilization, urease and tryptophan deaminase activities, indole and acetoin production, and glucose and inositol oxidation. Our study contributes to a better understanding of the taxonomy and biodiversity of this important bacterial group with great biotechnological and agricultural potential.
Freshwater crabs (Potamiscus manipuriensis), commonly consumed as local delicacies by the native people in the state of Manipur, were found to harbour metacercariae of Microphallus sp. (Family Microphyllidae), which were morphologically different from metacercariae of Microphallus spp reported earlier from different regions. So, PCR-based molecular characterization of this metacercaria was done utilizing rDNA marker regions: larger subunit (LSU) or 28S (D1-D3 region) and inter-transcribed spacer 2 (ITS2). Sequence and phylogenetic analyses confirmed that the taxon under study belonged to family Microphyllidae of genus Microphallus.
Two myxobacterial strains (KH5-1T and NO1) were isolated from the activated sludge tanks treating municipal sewage wastewater in Japan. These strains were recognised as myxobacteria based on their phenotypic characteristics of swarming colonies and fruiting bodies. Phylogenetic analyses using the 16S rRNA gene revealed that strains KH5-1T and NO1 were affiliated with the genus Corallococcus, with the closest neighbours being Corallococcus exercitus AB043AT (99.77% and 99.84%, respectively). Genome comparisons using orthologous average nucleotide identity (orthoANI) and digital DNA-DNA hybridisation similarity (dDDH) with strains KH5-1T and NO1 and their phylogenetically close relatives in Corallococcus spp. were below the thresholds. The major cellular fatty acids of strains KH5-1T and NO1 were iso-C15:0 (31.9%, 30.0%), summed feature 3 (comprising C16:1ω7c and/or C16:1ω6c) (20.2%, 17.7%), and iso-C17:0 (12.1%, 14.8%), and the major respiratory quinone was found to be menaquinone (MK)-8. Based on the phenotypic, chemotaxonomic, and phylogenetic evidence, strains KH5-1T and NO1 represent a new species in the genus Corallococcus, for which the proposed name is Corallococcus caeni sp. nov. The type strain is KH5-1T (= NCIMB 15510T = JCM 36609T).
Two Gram-stain-negative, rod-shaped, non-motile, strictly aerobic strains, forming yellow colonies and designated F6058 T and S2608 T , were isolated from marine sediment collected in Weihai, PR China. Both strains grow at 4–40 °C (optimum, 30–33 °C), pH 6.0–7.5 (optimum, pH 6.5) and in the presence of 0–7.0 % (w/v) NaCl. The optimum NaCl concentrations for strains F6058 T and S2608 T were 2.0 % and 2.5 %, respectively. Phylogenetic analysis of the 16S rRNA gene sequences indicated that strains F6058 T and S2608 T share an evolutionary lineage with members of the genus Aequorivita . The isolates exhibited a 16S rRNA gene sequence similarity of 96.7 % to each other. Strains F6058 T exhibited the highest 16S rRNA gene sequence similarity to Aequorivita xiaoshiensis F64183 T (98.8 %), and S2608 T was most similar to Aequorivita capsosiphonis A71 T (96.9 %). Iso-C 15:0 , anteiso-C 15:0 and iso-C 17:0 3-OH were the major fatty acids of strains F6058 T and S2608 T . The sole respiratory quinone of both isolates was menaquinone 6 (MK-6). The polar lipid profiles of the isolates both consisted of phosphatidylethanolamine and phosphoglycolipids; however, strain F6058 T exhibited one glycolipid, one aminolipid and two unidentified polar lipids, and strain S2608 T also had two glycolipids and one unidentified polar lipid. The DNA G+C contents of strains F6058 T and S2608 T were 34.6 % and 37.7 mol%, respectively. Based on their phenotypic, chemotaxonomic and genomic characteristics, strains F6058 T and S2608 T were considered to represent novel species of the genus Aequorivita , for which the names Aequorivita sediminis sp. nov. and Aequorivita marina sp. nov. were proposed. The type strains are F6058 T (=KCTC 92653 T =MCCC 1H01358 T ) and S2608 T (KCTC 92652 T =MCCC 1H01361 T ).
A new species of Delorhachis Karsch, 1896, Delorhachis nouabaleensis sp. n. is described from the Nouabalé-Ndoki National Park in northern Republic of Congo based on morphological and genetic evidence. This area is likely to have high levels of endemism due to the surrounding Sangha River Interval.
This study presents a comprehensive investigation into the evolutionary trajectories of Rhipicephalus ticks (Ixodidae) through the interpretation of molecular phylogenetics, elucidating their chromatographic spectrum. The use of advanced chromatographic tools in this study explored the dynamics chemical profiling, providing valuable insights into the evolutionary history and ecological adaptations. Prevalence of Rhipicephalus ticks was 4.5% in sheep and 3.9% in goats. The ITS2 sequence of the Rhipicephalus sanguineus (OK642408) and Rhipicephalus microplus (OK642409) form a distinct clade with sequences from other countries. The 16S rRNA sequences of R. sanguineus (OK560870) clustered with sequences form three lineages, tropical, temperate, and south‐eastern. The Cox I gene‐identified Rhipicephalus turanicus (OK623472) and R. microplus (OK623463) form separate clades with sequences. The HPLC chromatogram of tick samples reveals a diverse array of identified hydrocarbons, explained the complex chemical composition of their exoskeletons. This analytical approach provides valuable insights into the specific hydrocarbon profiles, allowing for potential applications in species differentiation, ecological studies, and a deeper understanding of the functional roles played by hydrocarbon compounds in tick physiology. The findings revealed the potential of applying molecular phylogenetics tools with chromatography not only to enhance our understanding of tick evolution but also to inform strategies for disease control and management in regions where Rhipicephalus ticks (Ixodidae) are endemic.
Research Highlights
Chemical mapping utilizing advanced chromatographic techniques.
Scanning microscopic insights high‐resolution scanning tool to observe structural and morphological features of ticks at a molecular level.
Molecular phylogeny data elucidate the evolutionary relationships among tick species.
Oryza is remarkable genus - with two domesticated (i.e. Asian and African rice) and 25 diploid and tetraploid wild species, 11 extant genome types, and a ∼3.4-fold genome size variation - that possesses a virtually untapped reservoir of genes that can be used for crop improvement. Here we unveil and interrogate 11 new chromosome-level assemblies of nine tetraploid and two diploid wild Oryza species in the context of ∼15 million years of evolution of the genus. We show that the core Oryza (sub)genome across all genome types is only ∼200 Mb and largely syntenic, while the remaining nuclear fractions, spanning ∼80-600 Mb, are intermingled, extremely plastic and rapidly evolving. For the halophyte O. coarctata, we show that - despite the detection of gene fractionation in the subgenomes - homoeologous genes are expressed at higher levels in one subgenome over the other in a mosaic form, thereby showing subgenome equivalence. The integration of these 11 new ultra-high quality reference genomes with our previously published genome data sets provide a nearly complete picture of the consequences of natural and artificial selection across the evolutionary history of Oryza. This in turn opens the door to unlock their genetic potential for future crop improvement and neodomestication.
The Pama Croaker, Otolithoides pama, is an economically important fish species in Bangladesh. Intra-family similarities in morphology and typical barcode sequences of cox1 create ambiguities in its identification. Therefore, morphology and the complete mitochondrial genome of O. pama, and comparative mitogenomics within the family Sciaenidae have been studied. Extracted genomic DNA was subjected to Illumina-based short read sequencing for De-Novo mitogenome assembly. The complete mitogenome of O. pama (Accession: OQ784575.1) was 16,513 bp, with strong AC biasness and strand asymmetry. Relative synonymous codon usage (RSCU) among 13 protein-coding genes (PCGs) of O. pama was also analyzed. The studied mitogenomes including O. pama exhibited consistent sizes and gene orders, except for the genus Johnius which possessed notably longer mitogenomes with unique gene rearrangements. Different genetic distance metrics across 30 species of Sciaenidae family demonstrated 12S rRNA and the control region (CR) as the most conserved and variable regions, respectively, while most of the PCGs undergone a purifying selection. Different phylogenetic trees were congruent with one another, where O. pama was distinctly placed. This study would contribute to distinguishing closely related fish species of Sciaenidae family and can be instrumental in conserving the genetic diversity of O. pama.
Ribosomal DNA (rDNA) repeat units are organized into tandem clusters in eukaryotic cells. In mice, these clusters are located on at least eight chromosomes and show extensive variation in the number of repeats between mouse genomes. To analyze intra- and inter-genomic variation of mouse rDNA repeats, we selectively isolated 25 individual rDNA units using Transformation-Associated Recombination (TAR) cloning. Long-read sequencing and subsequent comparative sequence analysis revealed that each full-length unit comprises an intergenic spacer (IGS) and a ∼13.4 kb long transcribed region encoding the three rRNAs, but with substantial variability in rDNA unit size, ranging from ∼35 to ∼46 kb. Within the transcribed regions of rDNA units, we found 209 variants, 70 of which are in external transcribed spacers (ETSs); but the rDNA size differences are driven primarily by IGS size heterogeneity, due to indels containing repetitive elements and some functional signals such as enhancers. Further evolutionary analysis categorized rDNA units into distinct clusters with characteristic IGS lengths; numbers of enhancers; and presence/absence of two common SNPs in promoter regions, one of which is located within promoter (p)RNA and may influence pRNA folding stability. These characteristic features of IGSs also correlated significantly with 5′ETS variant patterns described previously and associated with differential expression of rDNA units. Our results suggest that variant rDNA units are differentially regulated and open a route to investigate the role of rDNA variation on nucleolar formation and possible associations with pathology.
Sequences of human beta-globin mRNA were determined by analysis of complementary DNA. beta-mRNA was transcribed into double-stranded cDNA by RNA-dependent DNA polymerase. cDNA was cut by restriction endonucleases and the fragments were terminally labeled by means of polynucleotide kinase and [gamma-32P]ATP. After purification, fragments were degraded by snake venom phosphodiesterase. Alternatively single-stranded [32P]cDNA was prepared by transcription in the presence of [alpha-32P]dCTP and actinomycin D; the product was digested by endonuclease IV and degraded by snake venom phosphodiesterase. cDNA tracts obtained by both labeling methods enabled us to construct a sequence for the translated and 3'-terminal untranslated regions of human beta-mRNA.
The molecular cloning and nucleotide sequence analysis of adult chicken β globin mRNA is reported. DNA sequences derived from
in vitro transcription of globin mRNA were purified and amplified as recombinant DNA using the plasmid pBR322. Sequence analysis of
several clones coding for β globin strongly suggests that transcription errors may be generated near the 5′ end of transcripts
in vitro by reverse transcription. The complete sequence of the longest β globin insert containing 51 bases of the 5′ untranslated
region as well as the complete coding and 3′ untranslated regions has been determined.
The frequency of amino acid substitutions, relative to the frequency expected by chance, decreases linearly with the increase in physico-chemical differences between amino acid pairs involved in a substitution. This correlation does not apply to abnormal human hemoglobins. Since abnormal hemoglobins mostly reflect the process of mutation rather than selection, the correlation manifest during protein evolution between substitution frequency and physico-chemical difference in amino acids can be attributed to natural selection. Outside of 'abnormal' proteins, the correlation also does not apply to certain regions of proteins characterized by rapid rates of substitution. In these cases again, except for the largest physico-chemical differences between amino acid pairs, the substitution frequencies seem to be independent of the physico-chemical parameters. The limination of the substituents involving the largest physico-chemical differences can once more be attributed to natural selection. For smaller physico-chemical differences, natural selection, if it is operating in the polypeptide regions, must be based on parameters other than those examined.
Most evolutionary change in proteins may be due to neutral mutations and genetic drift.
The following five principles were deduced from the accumulated evidence
on molecular evolution and theoretical considerations of the population
dynamics of mutant substitutions: (i) for each protein, the rate of
evolution in terms of amino acid substitutions is approximately
constant/site per year for various lines, as long as the function and
tertiary structure of the molecule remain essentially unaltered. (ii)
Functionally less important molecules or parts of a molecule evolve (in
terms of mutant substitutions) faster than more important ones. (iii)
Those mutant substitutions that disrupt less the existing structure and
function of a molecule (conservative substitutions) occur more
frequently in evolution than more disruptive ones. (iv) Gene duplication
must always precede the emergence of a gene having a new function. (v)
Selective elimination of definitely deleterious mutants and random
fixation of selectively neutral or very slightly deleterious mutants
occur far more frequently in evolution than positive Darwinian selection
of definitely advantageous mutants.
Quantitative consideration of the operation and evolution of the information-processing system that constitutes the terrestrial biosphere indicates that non-Darwinian evolutionary changes cannot be expected to account for the increase in the amount of non-random biospheric structure that constitutes the information content of the biosphere. Changes that lead to such an increase must also be selectively advantageous and lead to preferential survival through natural selection.
A set of simple equations is derived which gives the relationship between the observed amino acid differences per 100 codons and the evolutionary distance per 100 codons using Holmquist's stochastic model of molecular evolution.
Sequences of human beta-globin mRNA were determined by analysis of complementary DNA. beta-mRNA was transcribed into double-stranded cDNA by RNA-dependent DNA polymerase. cDNA was cut by restriction endonucleases and the fragments were terminally labeled by means of polynucleotide kinase and [gamma-32P]ATP. After purification, fragments were degraded by snake venom phosphodiesterase. Alternatively single-stranded [32P]cDNA was prepared by transcription in the presence of [alpha-32P]dCTP and actinomycin D; the product was digested by endonuclease IV and degraded by snake venom phosphodiesterase. cDNA tracts obtained by both labeling methods enabled us to construct a sequence for the translated and 3'-terminal untranslated regions of human beta-mRNA.
The nucleotide sequence of a cloned rabbit chromosomal DNA segment of 1620 nucleotides length which contains a beta-globin gene is presented. The coding regions are separated into three blocks by two intervening sequences of 126 and 573 base pairs, respectively. The rabbit sequence was compared with a homologous mouse sequence. The segments flanking the rabbit gene, as well as the coding regions, the 5' noncoding and part of the 3' noncoding messenger RNA sequences are similar to those of the mouse gene; the homologous introns, despite identical location, are distinctly dissimilar except for the junction regions. Homologous introns may be derived from common ancestral introns by large insertions and deletions rather than be multiple point mutations.
The rabbit beta-globin DNA insertion of the hybrid plasmid PbetaG1 (Maniatis et al., 1976) was sequenced by the method of Maxam and Gilbert (1977). A sequence of 576 nucleotides was determined and verified by pyrimidine tract analysis of double-stranded DNA, synthesized in vitro starting from beta-globin mRNA. The derived sequence is in complete agreement with previously reported partial mRNA sequencing data and with the predictions from the primary structure of the protein. Moreover, the globin DNA insertion is missing only 13 nucleotides corresponding to the 5' terminal sequence of the mRNA. The rabbit beta-globin mRNA consists of a coding region of 438 nucleotides, flanked by a 5' noncoding region of 56 nucleotides (including the initiation codon AUG but not the 7-methyl-guanine of the "cap structure") and by a 3' noncoding region of 95 nucleotides (including a UGA termination codon). The features of the mRNA sequence are discussed with specific attention to the selective use of particular codons, the probable existence extensively base-paired segments at the 5' terminal region and the ribosome binding site. The faithful representation of beta-globin mRNA in the PbetaG1 DNA insertion establishes the validity of used cloned DNA, initially derived from double-stranded DNA transcripts of mRNA, for studying the structure of eucaryotic genes.
We have determined the entire nucleotide sequence of a cloned β-globinmaj gene derived from the BALB/c mouse. This sequence is 1567 bases long and includes the 5′ cap region as well as the presumptive poly(A) addition site of β-globin mRNA. The sequence establishes the fact that the gene is encoded in three discontinuous segments of DNA interrupted by two intervening sequences and precisely locates each. The smaller intervening sequence, 116 bases long, occurs between Arg and Leu codons at codon positions 30 and 31. The larger intervening sequence of 646 bases also occurs between Arg and Leu codons, but at codon positions 104 and 105. There is striking homology between the borders of the two intervening sequences, but no extensive dyad symmetry. Furthermore, the DNA region that just precedes and overlaps the 5′ cap structure of the mRNA shows homology to corresponding regions in other eucaryotic genes including the late adenovirus promoter. The 3′ untranslated sequence is closely homologous to that of the rabbit β-globin mRNA. The sequence thus allows us to identify several noncoding regions of potential importance for the expression and processing of genetic information. It also provides a basis for future comparison with other sequenced genes and a defined substrate for the development of direct tests of gene function.
The rabbit alpha-globin DNA insertion in the chimeric plasmid pHb 72 (Liu et al., 1977) has been sequenced by the method of Maxam and Gilbert (1977). This has enabled us to determine the messenger RNA(mRNA) sequence beginning in the 5' untranslated region 9 nucleotides before the initiation codon and extending through the first 361 nucleotides of the translated region. The data reported here overlap and are in complete agreement with sequences determined by Baralle (1977) for the 5' end of the mRNA and by Proudfoot et al. (1977) for the 3' end. Our sequence is also in agreement with the partial complementary RNA (cRNA) sequencing data which we reported previously (Paddock et al., 1977), this work marks the completion of the primary sequence of the rabbit alpha-globin mRNA. These observations reaffirm the high fidelity with which gene copies can be synthesized in vitro, cloned in a bacterial plasmid and maintained in the host. The general features of the mRNA nucleotide sequence are duscussed with particular attention given to the base composition and codon preferences observed and to comparison of this sequence with other completed mRNA gene sequences. A new computer program has been used to search for the most stable base-pairing arrangement of the completed mRNA.
ACCORDING to the neutral mutation-random drift hypothesis of molecular evolution and polymorphism1,2, most mutant substitutions detected through comparative studies of homologous proteins (and the nucleotide sequences) are the results of random fixation of selectively neutral or nearly neutral mutations. This is in sharp contrast to the orthodox neo-Darwinian view that practically all mutant substitutions occurring within species in the course of evolution are caused by positive Darwinian selection3-5. This paper shows that by comparative studies of messenger RNA (mRNA) sequences reliable estimates can be obtained of the evolutionary rates (in terms of mutant substitutions) at the third positions of the codon, and that the estimates conform remarkably well with the framework of the neutral theory.
3',5'-Linked hexa-adenylic acid with a 2',3'-cyclic phosphate terminus [(A5A less than p] couples on a polyuridylic acid template in the presence of ethylenediamine to form the dodecamer (24 percent) and octadecamer (5 percent). The bond produced is largely that of the 2',5' isomer.
Calculating the rate of evolution in terms of nucleotide substitutions seems to give a value so high that many of the mutations involved must be neutral ones.
A mouse alpha-globin-related pseudogene (psi alpha 30.5) completely lacks intervening sequences, and could not code for a functional globin poypeptide because of frameshifts. The widespread occurrence of globin pseudogenes in other species suggests that they are not 'dead' genes but may be important in controlling globin expression.
A method for estimating the evolutionary rates of synonymous and amino acid substitutions from homologous nucleotide sequences is presented. This method is applied to genes of phi X174 and G4 genomes, histone genes and beta-globin genes, for which homologous nucleotide sequences are available for comparison to be made. It is shown that the rates of synonymous substitutions are quite uniform among the non-overlapping genes of phi X174 and G4 and among histone genes H4, H2B, H3 and H2A. A comparison between phi X174 and G4 reveals that, in the overlapping segments of the A-gene, the rate of synonymous substitution is reduced more significantly than the rate of amino acid substitution relative to the corresponding rate in the non-overlapping segment. It is also suggested that, in the coding region surrounding the splicing points of intervening sequences of beta-globin genes, there exist rigid secondary structures. It is in only these regions that the beta-globin genes show the slowing down of evolutionary rates of both synonymous and amino acid substitutions in the primate line.
We have cloned and determined the nucleotide sequence of a mouse alpha-globin-like gene that entirely lacks the two intervening sequences that interrupt all globin genes thus far examined. The fact that this gene, alpha-3, is closely homologous to the normal adult alpha gene sequence suggests that it arose after the alpha/beta divergence and that it therefore must have lost its intervening sequences. The further fact that these intervening sequences have been lost cleanly--that is, in according with the G-T/A-G splicing rule of RNA--suggests, among other possibilities, that their loss may have been brought about by a gene conversion event involving the mediation of mature globin mRNA or its cDNA cognate. We propose such a mechanism that would permit the loss of either or both intervening sequences independently. Only the loss of both, however, should result in the inactivation of the globin gene, as seems to be the case with alpha-3.
The condensation of mononucleotides has been carried out in aqueous solution at neutral pH in the presence of cyanamide. Oligodeoxyribonucleotides
up to five units have been formed when montmorillonite was present.
The procession of life
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