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Fluorescent in situ hybridization to P. edulis chromosomes. a LINE (long interspersed nuclear element) and bTy3-gypsy LTR retrotransposon, showing a dispersed and uniform distribution. cTy1-copia LTR retrotransposon showing a dispersed but not entirely uniform distribution (arrow heads indicate four unlabeled small chromosomes). d BAC Pe55J16 (red) showing a subtelomeric pattern on most chromosome ends. Bar = 5 μm
Source publication
Passiflora edulis, the yellow passion fruit, is the main crop from the Passiflora genus, which comprises 525 species with its diversity center in South America. Genetic maps and a BAC (bacterial artificial chromosome) genomic library are available, but the nine chromosome pairs of similar size and morphology (2n = 18) hamper chromosome identificati...
Citations
... Long terminal repeat retroelements predominated, consisting mainly of Gypsy elements, with overrepresentation of the RLG_peDel (or Tekay lineage according to Neumann et al. [2019]), and single elements of P. edulis were obtained for the first time . Additionally, Sader, Dias, et al. (2019), who used a Gypsy element sequence as a probe for in situ hybridization, found a dispersed and uniform distribution pattern along P. edulis chromosomes, implying that they were abundant and could have significantly influenced the genome size. Other information on the nuclear genome of P. edulis was derived from lowcoverage sequencing data, facilitating microsatellite marker development (Araya et al., 2017) and providing cytogenomic markers, especially satellite sequences that are possibly associated with 5S and 35S rDNA or subtelomeres (Pamponét et al., 2019). ...
The genus Passiflora comprises a large group of plants popularly known as passionfruit, much appreciated for their exotic flowers and edible fruits. The species (∼500) are morphologically variable (e.g., growth habit, size, and color of flowers) and are adapted to distinct tropical ecosystems. In this study, we generated the genome of the wild diploid species Passiflora organensis Gardner by adopting a hybrid assembly approach. Passiflora organensis has a small genome of 259 Mbp and a heterozygosity rate of 81%, consistent with its reproductive system. Most of the genome sequences could be integrated into its chromosomes with cytogenomic markers (satellite DNA) as references. The repeated sequences accounted for 58.55% of the total DNA analyzed, and the Tekay lineage was the prevalent retrotransposon. In total, 25,327 coding genes were predicted. Passiflora organensis retains 5,609 singletons and 15,671 gene families. We focused on the genes potentially involved in the locus determining self‐incompatibility and the MADS‐box gene family, allowing us to infer expansions and contractions within specific subfamilies. Finally, we recovered the organellar DNA. Structural rearrangements and two mitoviruses, besides relics of other mobile elements, were found in the chloroplast and mt‐DNA molecules, respectively. This study presents the first draft genome assembly of a wild Passiflora species, providing a valuable sequence resource for genomic and evolutionary studies on the genus, and support for breeding cropped passionfruit species.
... In our study, we have seen that some Tekay clusters were present in the last common ancestral among the three species. Apart from methodological differences (we are investigating clusters of TEs, not individual elements), a possible explanation for this discrepancy could be that the authors analysed TEs from gene-rich regions, which are located at chromosomal ends (Sader et al. 2019b). In our case, we used elements from the most abundant clusters that may have accumulated in the pericentromeric regions for longer time. ...
... Here we have observed that Ty1/ copia-Angela and Ty3/gypsy-Tekay retroelements were scattered in P. quadrangularis and P. cincinnata chromosomes, although Angela showed more intense labelling at proximal regions. An uneven scattered distribution was also observed in P. edulis, as well as in P. alata and P. watsoniana, all from the subgenus Passiflora, where terminal or terminal regions of the chromosome arms are gene-rich, and the proximal regions are gene-poor and consist of dispersed repetitive sequences (Pamponét et al. 2019;Sader et al. 2019b;Stack et al. 2009;Dias et al. 2020). This distribution pattern suggests that gene-rich regions, and probably recombination, is higher in chromosomal ends of larger genome species, such as P. quadrangularis, P. cincinnata, and P. edulis. ...
Main conclusions
While two lineages of retrotransposons were more abundant in larger Passiflora genomes, the satellitome was more diverse and abundant in the smallest genome analysed.
AbstractRepetitive sequences are ubiquitous and fast-evolving elements responsible for size variation and large-scale organization of plant genomes. Within Passiflora genus, a tenfold variation in genome size, not attributed to polyploidy, is known. Here, we applied a combined in silico and cytological approach to study the organization and diversification of repetitive elements in three species of this genus representing its known range in genome size variation. Sequences were classified in terms of type and repetitiveness and the most abundant were mapped to chromosomes. We identified long terminal repeat (LTR) retrotransposons as the most abundant elements in the three genomes, showing a considerable variation among species. Satellite DNAs (satDNAs) were less representative, but highly diverse between subgenera. Our results clearly confirm that the largest genome species (Passiflora quadrangularis) presents a higher accumulation of repetitive DNA sequences, specially Angela and Tekay elements, making up most of its genome. Passiflora cincinnata, with intermediate genome and from the same subgenus, showed similarity with P. quadrangularis regarding the families of repetitive DNA sequences, but in different proportions. On the other hand, Passiflora organensis, the smallest genome, from a different subgenus, presented greater diversity and the highest proportion of satDNA. Altogether, our data indicates that while large genomes evolved by an accumulation of retrotransposons, the smallest genome known for the genus has evolved by diversification of different repeat types, particularly satDNAs.
... Seventy-eight BAC probes were assigned to specific chromosomes. These data are similar to those obtained in other works where BAC probes were used to establish a cytogenetic map, as in the case of Passiflora edulis, where 36 BAC clones were used [41], or Saccharum spontaneum, with 114 BAC clones [42]. It was possible to obtain chromosomal markers for all the chromosomes of Senegalese sole. ...
Solea senegalensis aquaculture production has experienced a great increase in the last decade and, consequently, the genome knowledge of the species is gaining attention. In this sense, obtaining a high-density genome mapping of the species could offer clues to the aquaculture improvement in those aspects not resolved so far. In the present article, a review and new processed data have allowed to obtain a high-density BAC-based cytogenetic map of S. senegalensis beside the analysis of the sequences of such BAC clones to achieve integrative data. A total of 93 BAC clones were used to localize the chromosome complement of the species and 588 genes were annotated, thus almost reaching the 2.5% of the S. senegalensis genome sequences. As a result, important data about its genome organization and evolution were obtained, such as the lesser gene density of the large metacentric pair compared with the other metacentric chromosomes, which supports the theory of a sex proto-chromosome pair. In addition, chromosomes with a high number of linked genes that are conserved, even in distant species, were detected. This kind of result widens the knowledge of this species’ chromosome dynamics and evolution.
... In our study, we have seen that some Tekay clusters were present in the last common ancestral among the three species. Apart from methodological differences (we are investigating clusters of TEs, not individual elements), a possible explanation for this discrepancy could be that the authors analysed TEs from gene-rich regions, which are located at chromosomal ends (Sader et al. 2019b). In our case, we used elements from the most abundant clusters that may have accumulated in the pericentromeric regions for longer time. ...
... Here we have observed that Ty1/ copia-Angela and Ty3/gypsy-Tekay retroelements were scattered in P. quadrangularis and P. cincinnata chromosomes, although Angela showed more intense labelling at proximal regions. An uneven scattered distribution was also observed in P. edulis, as well as in P. alata and P. watsoniana, all from the subgenus Passiflora, where terminal or terminal regions of the chromosome arms are gene-rich, and the proximal regions are gene-poor and consist of dispersed repetitive sequences (Pamponét et al. 2019;Sader et al. 2019b;Stack et al. 2009;Dias et al. 2020). This distribution pattern suggests that gene-rich regions, and probably recombination, is higher in chromosomal ends of larger genome species, such as P. quadrangularis, P. cincinnata, and P. edulis. ...
Repetitive sequences are ubiquitous and fast-evolving elements responsible for size variation and large-scale organization of plant genomes. Within Passiflora genus, a ten-fold variation in genome size, not attributed to polyploidy, is known. Here, we applied a combined in silico and cytological approach to study the organization and diversification of repetitive elements in three species of these genera representing its known range in genome size variation. Sequences were classified in terms of type and repetitiveness and the most abundant were mapped to chromosomes. We identified Long Terminal Repeat (LTR) retrotransposons as the most abundant elements in the three genomes, showing a considerable variation among species. Satellite DNAs (satDNAs) were less representative, but highly diverse between subgenera. Our results clearly confirm that the largest genome species ( Passiflora quadrangularis ) presents a higher accumulation of repetitive DNA sequences, specially Angela and Tekay elements, making up most of its genome. Passiflora cincinnata , with intermediate genome and from the same subgenus, showed similarity with P. quadrangularis regarding the families of repetitive DNA sequences, but in different proportions. On the other hand, Passiflora organensis , the smallest genome, from a different subgenus, presented greater diversity and the highest proportion of satDNA. Altogether, our data indicate that while large genome evolve by an accumulation of retrotransponsons, small genomes most evolved by diversification of different repeat types, particularly satDNAs.
MAIN CONCLUSIONS
While two lineages of retrotransposons were more abundant in larger Passiflora genomes, the satellitome was more diverse and abundant in the smallest genome.
... In Corchorus L., for example, fragments of the retroelement Ty3-Gypsy were used for the construction of two species-specific cytogenetic maps (Begum et al. 2013). Recently, single-copy BAC clones, as well as repetitive elements, have been used to establish a cytogenetic map of P. edulis showing that the 5S rDNA and 35S rDNA sites are located on chromosome 5 and chromosomes 7 and 8, respectively (Sader et al. 2019b). ...
... In the present work, BACs corresponding to a gene-rich fraction of the P. edulis genome (Santos et al. 2014) and previously cytogenetically mapped as single-copy markers in this species (Sader et al. 2019b) were used as probes for the construction of comparative maps for P. alata and P. watsoniana using BAC-FISH. The repetitive elements Ty1-Copia and Ty3-Gypsy of P. edulis were also used to search for a distribution pattern that could be useful for chromosome identification in P. alata and P. watsoniana. ...
... Both repetitive and single-copy probes were used for FISH. Partial sequences of Ty1-Copia and Ty3-Gypsy retrotransposons from P. edulis (Santos et al. 2014) were amplified by PCR using specific primers (Sader et al. 2019b). In addition, 5S and 35S ribosomal DNA were located with the D2 clone from Lotus japonicus (Regel) K.Larsen (Pedrosa et al. 2002) and pTa71 from Triticum aestivum L. (Gerlach and Bedbrook 1979), respectively. ...
The genus Passiflora shows a large variation in chromosome numbers (2n = 12, 14, 18, 20, 22, 24, 36, 72) and in the distribution of ribosomal DNA (rDNA) sites. Most species of the subgenus Passiflora, however, have n = 9 and two pairs of 35S rDNA sites, such as species of economic importance Passiflora edulis and Passiflora alata. Species of another clade from the same subgenus, which includes Passiflora watsoniana, have three pairs of 35S rDNA sites. This karyotypic variability suggests a possible break in synteny within the subgenus Passiflora, but with maintenance of n = 9 in most species. To investigate synteny in three species of this subgenus, comparative cytogenetic mapping was performed in P. alata and P. watsoniana using BAC markers previously developed for P. edulis. In addition, the distribution of 35S and 5S rDNA sites and LTR retrotransposons Ty1-Copia and Ty3-Gypsy were investigated. The used single-copy BAC clones resulted in distinct dot-like signals exclusively in terminal or subterminal regions of the chromosomes. No synteny break with regard to the chromosomal distribution of these BACs or the rDNA sites could be detected, except for an additional 35S rDNA site on chromosome 3 of P. watsoniana. LTR elements showed proximal or uniformly dispersed patterns. The results revealed a similar distribution of BAC markers between species, preferably in the terminal regions, which is compatible with an increased accumulation of repetitive sequences preferentially in proximal regions of the chromosomes associated with an increase in genome size.
... Gypsy was predominant in the gene-rich fraction, and represented a large proportion of the draft genome. The data recently released by [76,99] corroborate our hypothesis, implying that this element is highly frequent and could have significantly influenced genome size. ...
A significant proportion of plant genomes is consists of transposable elements (TEs), especially LTR retrotransposons (LTR-RTs) which are known to drive genome evolution. However, not much information is available on the structure and evolutionary role of TEs in the Passifloraceae family (Malpighiales order). Against this backdrop, we identified, characterized, and inferred the potential genomic impact of the TE repertoire found in the available genomic resources for Passiflora edulis, a tropical fruit species. A total of 250 different TE sequences were identified (96% Class I, and 4% Class II), corresponding to ~ 19% of the P. edulis draft genome. TEs were found preferentially in intergenic spaces (70.4%), but also overlapping genes (30.6%). LTR-RTs accounted for 181 single elements corresponding to ~ 13% of the draft genome. A phylogenetic inference of the reverse transcriptase domain of the LTR-RT revealed association of 37 elements with the Copia superfamily (Angela, Ale, Tork, and Sire) and 128 with the Gypsy (Del, Athila, Reina, CRM, and Galadriel) superfamily, and Del elements were the most frequent. Interestingly, according to insertion time analysis, the majority (95.9%) of the LTR-RTs were recently inserted into the P. edulis genome (< 2.0 Mya), and with the exception of the Athila lineage, all LTR-RTs are transcription-ally active. Moreover, functional analyses disclosed that the Angela, Del, CRM and Tork lineages are conserved in wild Passiflora species, supporting the idea of a common expansion of Copia and Gypsy superfamilies. Overall, this is the first study describing the P. edulis TE repertoire, and it also lends weight to the suggestion that LTR-RTs had a recent expansion into the analyzed gene-rich region of the P. edulis genome, possibly along WGD (Whole genome duplication) events, but are under negative selection due to their potential deleterious impact on gene regions.
DNA in situ hybridization (DNA-ISH) is a widely used method in molecular cytogenetics that allows the localization of specific DNA sequences in particular regions of chromosomes. Implementation of DNA-ISH requires the use of DNA probes, which can be commercial or developed for specific research purposes as non-commercial (homemade) DNA probes. One of the significant drawbacks of non-commercial probes is the difficulty in obtaining a high signal intensity with a small DNA probe size. Therefore, developing approaches to enhance non-commercial DNA probes is an important task in modern molecular cytogenetics. To directly visualize small DNA sequences on a chromosome, the tyramide signal amplification (TSA) method is used. The TSA system is based on the formation of a covalent bond between electron-rich protein fragments in the sample and tyramide molecules linked to a hapten (in chromogenic in situ hybridization) or a fluorophore (in fluorescent in situ hybridization). This is achieved by converting tyramide molecules into free-radical intermediate compounds under the action of horseradish peroxidase (HRP), followed by deposition of precipitated molecules nearby. As a result, a low-intensity signal is amplified. Thus, TSA is a good complement to the DNA-ISH method, thanks to its high sensitivity and ability to detect small genomic imbalances, and can therefore become a valuable tool for diagnosing chromosomal rearrangements in clinical practice.
Passion fruit (Passiflora edulis) possesses a complex aroma and is widely grown in tropical and subtropical areas. Here, we conducted the de novo assembly, annotation and comparison of purple (P. edulis Sims) and yellow passion fruit (P. edulis f. flavicarpa) reference genomes using PacBio, Illumina and Hi-C technologies. Notably, we discovered evidence of recent whole-genome duplication events in P. edulis genomes. Comparative analysis revealed 7.6∼8.1 million single nucleotide polymorphisms, one million insertions/deletions, and over 142 Mb presence/absence variations among different P. edulis genomes. During the ripening of yellow passion fruit, metabolites related to flavor, aroma, and color were substantially accumulated or changed. Through joint analysis of genomic variations, differentially expressed genes, and accumulated metabolites, we explored candidate genes associated with flavor, aroma and color distinctions. Flavonoid biosynthesis pathways, anthocyanin biosynthesis pathways and related metabolites are pivotal factors affecting the coloration of passion fruit, and terpenoids metabolites accumulated more in purple passion fruit. Finally, by heterologous expression in yeast (Saccharomyces cerevisiae), we functionally characterized 12 terpene synthases (TPSs). Our findings revealed that certain TPS homologues in both yellow and purple passion fruit varieties produce identical terpene products, while others yield distinct compounds or even lose their functionality. These discoveries revealed the genetic and metabolic basis of unique characteristics in aroma and flavor between the two passion fruit varieties. This study provides resources for better understanding the genome architecture and accelerating genetic improvement of passion fruits.
Two forms of the genus Passiflora, belonging to the Passifloraceae family, are commonly called yellow and purple passion. These perennial woody climbers are found in the cooler regions at higher altitudes and in lowlands of tropical areas. The presence of alkaloids, terpenes, stilbenes, flavonoids, glycosides, carotenoids, etc. in different parts of the plant provides several pharmacological properties. Because of the various uses in foods and pharmaceuticals, in vitro propagation of this genus has been performed hugely and is of great interest to researchers. From different explants via direct organogenesis under controlled aseptic conditions, callus, root, shoot, and somatic embryos are induced successfully. Different PGRs are augmented in the media for the rapid multiplication or organogenesis, especially, the high ratio of cytokinin and auxin in the basal media efficiently regenerates the shoot and root respectively. The in vitro regenerated plantlets are then acclimatized and hardened properly before transferring to the field conditions. Thus, the present first of its kind review on P. edulis exclusively encompasses the wide applications of biotechnology for this species alongside its organogenesis, embryogenesis, cytology, and endophytic microbes with special emphasis on the role of genetic transformation studies mediated by Agrobacterium sp.
Key points
• Critical assessment on in vitro biotechnology in P. edulis.
• Agrobacterium-mediated transformation in P. edulis.
• Role of endophytic microbes in P. edulis.
