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Lander, E. S., Green, P., Abrahamson, J., Barlow, A., Daly, M. J., Lincoln, S. E. and Newburg, L.. MAPMAKER; An interactive Computer Package for Constructing Primary Genetic Linkage Maps of Experimental and Natural Populations. Genomics, 1: 174-181
Abstract
With the advent of RFLPs, genetic linkage maps are now being assembled for a number of organisms including both inbred experimental populations such as maize and outbred natural populations such as humans. Accurate construction of such genetic maps requires multipoint linkage analysis of particular types of pedigrees. We describe here a computer package, called MAPMAKER, designed specifically for this purpose. The program uses an efficient algorithm that allows simultaneous multipoint analysis of any number of loci. MAPMAKER also includes an interactive command language that makes it easy for a geneticist to explore linkage data. MAPMAKER has been applied to the construction of linkage maps in a number of organisms, including the human and several plants, and we outline the mapping strategies that have been used.
... Finally, a linkage map is constructed by coding data for each molecular marker on every individual in the population and utilizing computer algorithms. Frequently used software programs (Table2) for developing linkage maps consist of Mapmaker/EXP (Lander et al. 1987;Lincoln et al. 1993), JoinMap (Stam 1993) and MapManager QTX (Manly et al. 2001) which are easily accessible from the Internet. ...
... Higher degrees of connection can be distinguished using lower LOD values, and additional markers can be added to maps made using higher LOD values. MapManager QTX (Manly et al., 2001) and Mapmaker/EXP (Lander et al., 1987 andLincoln et al., 1993) are two frequently used software programmes that are both freely accessible online. Another popular programme for creating connectivity maps is called Join Map (Stam, 1993). ...
Human population is expanding in every part
of the planet earth and limited agricultural
produce will definitely raise the feeding
problems in future. Identification of varieties
at the genetic level for specific traits,
especially high yield, is very important for
fruitful breeding. Traditional methods of
germplasm screening are based upon its
performance in the field which is tedious and
time consuming and due to which many years
are required for the development of a new
variety. Thus, to fasten this process different
areas of technologies involving, molecular
biology, biotechnology and genetic
engineering are being utilized in the crop
improvement sector. This chapter focuses on
the different DNA fingerprinting techniques
which can be used in varietal identification of
crop plants. The unique DNA banding pattern
or the DNA fingerprints is the most suitable
tool for studying the relationship between the
closely related plant species and for assessing
the genetic diversity also. DNA fingerprinting
begins with the non-PCR based technique,
RFLP but with advent of the thermal cycler the
PCR based techniques (RAPD, SSR, ISSR and
SRAP) became the methods of choice. Further,
combination of PCR and hybridization-based
method, AFLP being more reliable. The new
generation sequencing techniques has
improved the DNA fingerprinting techniques
also to the next level and is the first choice of
the breeders.
... A QTL mapping study correlates the genotypic information from the parents and their progeny of a bi/multi-parental population to the phenotypic variations. QTLs are determined based on the availability of polymorphic molecular markers, through which a linkage map is generated, and the observable phenotypic spectrum from a mapping population, as well as the genetic constitution of the parents for the trait of interest, using statistical softwares like Mapmaker [6], QTLCartographer [7], Joinmap [8], and WinQTLCart [9]. Although QTL mapping is a time taking process as it involves generation and advancement of mapping population, the simplistic nature of the approach has led to discovery of QTLs for a varied number of traits in all the currently cultivated crops. ...
... Xu et al. [43] Triticum aestivum Fusarium head blight resistance 31 6 Cai et al. [44] Triticum Zheng et al. [48] Triticum to conduct a meta-analysis study. While other softwares can perform meta-analysis on data generated in multiple fields, MetaQTL and BioMercator softwares are solely dedicated to handling QTL data. ...
Meta-analysis consists of pooling results from multiple studies, and allows prediction of more precise and meaningful data. A plethora of QTL mapping studies for multiple traits in different crop plants have generated vast and often redundant information. A meta-QTL analysis helps in collating available trait specific QTL data from multiple studies and purging the redundant data, thereby leading to identification of more robust genomic regions known as “Meta-QTLs”. The identified meta-QTL regions have shorter confidence intervals from the yielding QTLs of individual mapping studies, thereby helping in gaining a deeper insight into the genetic framework of complex traits. This chapter provides an overview of the available tools for meta-analysis and a user-friendly protocol for using “BioMercator”, the most widely used software for performing a meta–QTL analysis. A good meta-QTL analysis is dependent on the availability of multiple independent trait specific QTL studies and therefore involves an extensive literature survey as an essential primary step. The compiled QTL information is processed to generate map files and a consensus map followed by projection of QTLs on the consensus map ultimately leading to identification of meta-QTL regions. Together, the reduction in the confidence interval and integration of meta-QTL regions with additional functional genomics datasets helps in shortlisting potential candidate genes and reduce the overall efforts for crop improvement through marker-assisted breeding or genetic engineering.
... Mapmaker/Exp3.0 was used to construct the genetic linkage map [37], while the composite interval mapping method included in the WinQTLCart software version 2.5 was used to perform QTL scanning on population phenotype data and genetic maps, as well as to screen QTLs with a logarithm of odds (LOD) threshold of 2.0 [38]. ...
The quality of rice, evaluated using multiple quality-related traits, is the main determinant of its market competitiveness. In this study, two japonica rice varieties with significant differences in quality-related traits were used as parents to construct two populations, BC3F2 and BC3F2:3, with Kongyu131 (KY131) as the recurrent parent. A genetic linkage map was constructed using the BC3F2 population based on 151 pairs of SSR/InDel polymorphic markers selected between the parents. Grain-shape-related traits (grain length GL, grain width GW, and length-to-width ratio LWR), chalkiness-related traits (white-core rate WCR, white-belly rate WBR, white-back rate BR, and chalkiness rate BR), and amylose content (AC) were investigated in the two populations in 2017 and 2018. Except for BR and CR, the traits showed similar characteristics with a normal distribution in both populations. Genetic linkage analysis was conducted for these quality-related traits, and a total of 37 QTLs were detected in the two populations. Further validation was performed on the newly identified QTLs with larger effects, and three grain shape QTLs and four chalkiness QTLs were successfully validated in different environments. One repeatedly validated QTL, qWCR3, was selected for fine mapping and was successfully narrowed down to a 100 kb region in which only two genes, LOC_0s03g45210 and LOC_0s03g45320, exhibited sequence variations between the parents. Furthermore, the variation of LOC_Os03g45210 leads to a frameshift mutation and premature protein termination. The results of this study provide a theoretical basis for positional cloning of the qWCR3 gene, thus offering new genetic resources for rice quality improvement.
... The function mstmap in the R/ASMap package (Taylor and Butler 2017) was used to adjust the marker positions and reorder the markers based on maximum likelihood (ML). The function est.map in the R/qtl package was used to estimate the genetic distance using Kosambi mapping function (Lander et al. 1987). ...
Key message
Two major-effect QTL GlcA07.1 and GlcA09.1 for green leaf color were fine mapped into 170.25 kb and 191.41 kb intervals on chromosomes A07 and A09, respectively, and were validated by transcriptome analysis.
Abstract
Non-heading Chinese cabbage (NHCC) is a leafy vegetable with a wide range of green colors. Understanding the genetic mechanism behind broad spectrum of green may facilitate the breeding of high-quality NHCC. Here, we used F2 and F7:8 recombination inbred line (RIL) population from a cross between Wutacai (dark-green) and Erqing (lime-green) to undertake the genetic analysis and quantitative trait locus (QTL) mapping in NHCC. The genetic investigation of the F2 population revealed that the variation of green leaf color was controlled by two recessive genes. Six pigments associated with green leaf color, including total chlorophyll, chlorophyll a, chlorophyll b, total carotenoids, lutein, and carotene were quantified and applied for QTL mapping in the RIL population. A total of 7 QTL were detected across the whole genome. Among them, two major-effect QTL were mapped on chromosomes A07 (GlcA07.1) and A09 (GlcA09.1) corresponding to two QTL identified in the F2 population. The QTL GlcA07.1 and GlcA09.1 were further fine mapped into 170.25 kb and 191.41 kb genomic regions, respectively. By comparing gene expression level and gene annotation, BraC07g023810 and BraC07g023970 were proposed as the best candidates for GlcA07.1, while BraC09g052220 and BraC09g052270 were suggested for GlcA09.1. Two InDel molecular markers (GlcA07.1-BcGUN4 and GlcA09.1-BcSG1) associated with BraC07gA023810 and BraC09g052220 were developed and could effectively identify leaf color in natural NHCC accessions, suggesting their potential for marker-assisted leaf color selection in NHCC breeding.
... MAPMAKER/EXP 3 developed by Lander et al., (1987), was used for the linkage mapping. Composite interval mapping for QTL analysis was conducted with 1000 permutation time at 0.05 probability level using WinQTL Cartographer version 2.5 (Wang, 2007). ...
The study was undertaken to construct a linkage map and to identify the QTLs for heading date, plant height, and culm strength using a F2 population derived from Ayeyarmin/IR 64, two varieties distinctly different in heading date and plant height. A total of 233 F2 individuals were genotyped using 70 SSR primers, and phenotypic data were collected for heading date, plant height and culm strength traits. Fifty-four out of 70 marker loci were mapped across twelve rice chromosomes spanning a total map length of 1592.30 cM with an average distance of 29.49 cM between adjacent marker loci using MAPMAKER/EXP 3. Composite interval mapping for QTL analysis was conducted with 1000 permutation time at 0.05 probability level using WinQTL Cartographer version 2.5. One major QTL for heading date was mapped between marker RM204 and RM50 on chromosome 6. One major QTL for culm strength was mapped between marker RM1003-RM5931, and two major QTLs for plant height were mapped between RM84-RM1003 and RM1003-RM5931 on chromosome 1. The molecular markers linked to the major QTLs could be potentially used in marker-assisted selection for the rainfed rice improvement breeding program.
... Based on the genotyping of the 190 plants of the F 2 population with 133 SSR and InDel markers, twelve linkage groups corresponding to 12 chromosomes were established by using the computer program MAP-MAKER/EXP 3.0 [59]. The recombination frequencies among the markers in the same linkage group were converted into genetic distances using the kosambi function. ...
Grain yield in rice is a complex trait and it is controlled by a number of quantitative trait loci (QTL). To dissect the genetic basis of rice yield, QTL analysis for nine yield traits was performed using an F2 population containing 190 plants, which was developed from a cross between Youyidao (YYD) and Sanfenhe (SFH), and each plant in the population evaluated with respect to nine yield traits. In this study, the correlations among the nine yield traits were analyzed. The grain yield per plant positively correlated with six yield traits, except for grain length and grain width, and showed the highest correlation coefficient of 0.98 with the number of filled grains per plant. A genetic map containing 133 DNA markers was constructed and it spanned 1831.7 cM throughout 12 chromosomes. A total of 36 QTLs for the yield traits were detected on nine chromosomes, except for the remaining chromosomes 5, 8, and 9. The phenotypic variation was explained by a single QTL that ranged from 6.19% to 36.01%. Furthermore, a major QTL for grain width and weight, qGW2-1, was confirmed to be newly identified and was narrowed down to a relatively smaller interval of about ~2.94-Mb. Collectively, we detected a total of 36 QTLs for yield traits and a major QTL, qGW2-1, was confirmed to control grain weight and width, which laid the foundation for further map-based cloning and molecular design breeding in rice.
... Of the various types of mapping populations, F 2 can be quickly developed and possesses all potential parental allelic combinations [81]. A recombinant inbred line (RIL), also known as an immortal or permanent mapping population can be produced from an F 2 generation using the single-seed descent (SSD) approach [82]. ...
Castor (Ricinus communis L.) is an industrially important oil producing crop belongs to Euphorbiaceae family. Castor oil has unique chemical properties make it industrially important crop. It is a member of monotypic genus even though it has ample amount of variability. Using this variability, conventionally many varieties and hybrids have been developed. But, like other crops, the modern and unconventional methods of crop improvement has not fully explored in castor. This article discusses the use of polyploidy induction, distant/wide hybridization and mutation breeding as tools for generating variety. Modern approaches accelerate the speed of crop breeding as an alternative tool. To achieve this goal, molecular markers are employed in breeding to capture the genetic variability through molecular analysis and population structuring. Allele mining is used to trace the evolution of alleles, identify new haplotypes and produce allele specific markers for use in marker aided selection using Genome wide association studies (GWAS) and quantitative trait loci (QTL) mapping. Plant genetic transformation is a rapid and effective mode of castor improvement is also discussed here. The efforts towards developing stable regeneration protocol provide a wide range of utility like embryo rescue in distant crosses, development of somaclonal variation, haploid development using anther culture and callus development for stable genetic transformation has reviewed in this article. Omics has provided intuitions to the molecular mechanisms of (a)biotic stress management in castor along with dissected out the possible genes for improving the yield. Relating genes to traits offers additional scientific inevitability leading to enhancement and sympathetic mechanisms of yield improvement and several stress tolerance.
... After analyzing the polymorphism of SSR markers in Cheongcheong and Nagdong, very similar markers were excluded. The selected SSR marker was used to construct a genetic map by Mapmaker version 3.0 (Lander et al., 1987). For linkage mapping, the MAP function of lciMapping software was used (Meng et al., 2015). ...
Introduction
Rice is an important food source that can provide a stable supply of calories for most people around the world. However, owing to the recent rapid temperature rise, we are facing social issues related to the increase in the Winkler scale. In this study, a strategy for screening potential candidate genes related to the yield according to the Winkler scale is presented, and the possibility of using a candidate gene identified through sequence haplotype and homology analysis as a breeding source is suggested.
Methods
QTL for the Winkler scale was identified using a population of 120 double haploids derived from a cross between Cheongchoneg, Indica, and Nagdong, Japonica.
Results and discussion
A total of 79 candidate genes were detected in the identified QTL region, and OsHAq8 was finally screened. Through haplotype analysis, OsHAq8 was derived from the Indica group and orthologous to Graminae’s activator of Hsp90 ATPase, suggesting that it is a candidate gene involved in yield according to temperature during the growing period. The expression level of OsHAq8 increased as the Winkler scale increased. The findings of this study can serve as a crucial indicator for predicting harvest time and grain quality while achieving a stable yield through marker selection and adaptation to climate change. Climate change occurs more frequently. In these situations, it is very important to predict harvest time and apply relevant candidate genes to breeding. The candidate genes presented in this study can be effectively applied to rice breeding in preparation for climate change.
... A standard paddy field was used for growing parental, F 1 , and F 2 plants. Employing Mapmaker/ Exp 3.0, a linkage analysis for molecular markers and the OsAL50 locus was carried out based on genotyping data for 1,211 recessive F 2 individuals displaying the mutant phenotype (Lander et al. 1987). After converting recombination rates into genetic distances using the Kosambi operating function, a genetic linkage map was produced using the MapDraw program (Liu and Meng 2003). ...
Leaf color is a highly important agronomic trait, and mutants with altered leaf coloration can serve as excellent models for studies on chloroplast development and chlorophyll biosynthesis, enabling the cloning of genes involved in these processes in rice (Oryza sativa L.). In this study, we isolated a stable genetic rice mutant, oryza sativa albino leaf 50 (osal50), from a breeding population of the japonica cultivar GP50. This mutant exhibited a distinctive albino phenotype, with white-striped leaves in seedlings and white panicles at the heading stage. Compared with wild-type GP50, the osal50 mutant showed lower chlorophyll and carotenoid accumulation, together with abnormal chloroplast ultrastructure. Genetic analysis demonstrated that a recessive nuclear gene was responsible for the albino phenotype of osal50, and a map-based cloning strategy delimited OsAL50 to a 160-kb physical interval on chromosome 1, flanked by two single nucleotide polymorphism (SNP) markers, CAPS-08 and CAPS-37, that included 26 putative open reading frames. Sequence and expression analyses revealed LOC_Os01g20110 as the candidate OsAL50 gene, which was confirmed by knockout using CRISPR/Cas9. Subcellular localization and protein sequence analyses suggested that OsAL50 likely encodes an endoribonuclease E-like protein localized to the chloroplasts. Further investigation indicated that OsAL50 plays a vital role in the regulation of photosynthetic pigment metabolism, photosynthesis, and chloroplast biogenesis. In summary, we identified a novel albino mutant that will serve as useful genetic material for studies of chlorophyll biosynthesis and chloroplast development in rice.
... Currently, numerous software are available for linkage map construction and mapping QTLs in bi-parental or multi-parental populations and most are open access/free (Table 5.1). MAPMAKER was the first landmark software for the construction of linkage maps and QTL mapping i.e. interval mapping [61]. Although it is still a preferred software by many research groups but it cannot compute CIM (effect of other QTLs on locating target QTL) and is limited to mapping of F 2 populations. ...
With the onset of the genomic era, mapping of Quantitative Traits Loci (QTLs) appears to be a feasible solution for dissecting the complex architecture of numerous multigenic traits of significance. Great progress has been made in the recent past in QTL mapping, advances in fine mapping and expression studies conjugated with cheaper prices present a new arena for QTL mapping of under-studied species that lack reference genome sequences. Here we reviewed, basic to advances in mapping of QTLs, QTG (Quantitative Traits Gene) sequence, Meta-QTL analysis, statistical software in mapping and validating QTL. This review also emphasizes precision phenotyping by an amalgamation of AI in the phenotyping of traits. Despite the identification of numerous major and minor QTLs governing various traits, only a few have been utilized in crop improvement programs. Furthermore, validations of identified QTL and their introgression in the elite lines are strongly recommended.
... A Chi-squared (χ 2 ) test was used to evaluate the goodness of fit of observed and expected segregation ratios. The MAP-MAKER software version 3.0 (Lander et al. 1987) was used to map the R-BPMV resistance gene on the integrated linkage map of common bean using the set of 142 markers as described in Geffroy et al. (2008). Linkage groups (LG) were established with a LOD threshold of 3.0 and a maximum recombination fraction of 0.3. ...
Key message
R-BPMV is located within a recently expanded TNL cluster in the Phaseolus genus with suppressed recombination and known for resistance to multiple pathogens including potyviruses controlled by the I gene.
Abstract
Bean pod mottle virus (BPMV) is a comovirus that infects common bean and legumes in general. BPMV is distributed throughout the world and is a major threat on soybean, a closely related species of common bean. In common bean, BAT93 was reported to carry the R-BPMV resistance gene conferring resistance to BPMV and linked with the I resistance gene. To fine map R-BPMV, 182 recombinant inbred lines (RILs) derived from the cross BAT93 × JaloEEP558 were genotyped with polymerase chain reaction (PCR)-based markers developed using genome assemblies from G19833 and BAT93, as well as BAT93 BAC clone sequences. Analysis of RILs carrying key recombination events positioned R-BPMV to a target region containing at least 16 TIR-NB-LRR (TNL) sequences in BAT93. Because the I cluster presents a suppression of recombination and a large number of repeated sequences, none of the 16 TNLs could be excluded as R-BPMV candidate gene. The evolutionary history of the TNLs for the I cluster were reconstructed using microsynteny and phylogenetic analyses within the legume family. A single I TNL was present in Medicago truncatula and lost in soybean, mirroring the absence of complete BPMV resistance in soybean. Amplification of TNLs in the I cluster predates the divergence of the Phaseolus species, in agreement with the emergence of R-BPMV before the separation of the common bean wild centers of diversity. This analysis provides PCR-based markers useful in marker-assisted selection (MAS) and laid the foundation for cloning of R-BPMV resistance gene in order to transfer the resistance into soybean.
... Markers that followed the expected Mendelian segregation ratio were used to genotype the BC 4 F 2 populations in the 16WJ, 16JH, and 16CZ environments. Third, a local genetic linkage map was constructed using MAPMAKER/EXP 3.0 (Lander et al. 1987), and the genetic map distances were calculated using the Kosambi mapping function (Kosambi 1994). By combining genetic linkage information and phenotyping data, the QTL analysis for TBN was conducted with QTL IciMapping 4.1 using inclusive composite interval mapping of additive (ICIM-ADD) and two-locus epistatic QTL (ICIM-EPI) modules. ...
The tassel competes with the ear for nutrients and shields the upper leaves, thereby reducing the yield of grain. The tassel branch number (TBN) is a pivotal determinant of tassel size, wherein the reduced TBN has the potential to enhance the transmission of light and reduce the consumption of nutrients, which should ultimately result in increased yield. Consequently, the TBN has emerged as a vital target trait in contemporary breeding programs that focus on compact maize varieties. In this study, QTL-seq technology and advanced population mapping were used to rapidly identify and dissect the major effects of the TBN on QTL. Advanced mapping populations (BC4F2 and BC4F3) were derived from the inbred lines 18–599 (8–11 TBN) and 3237 (0–1 TBN) through phenotypic recurrent selection. First, 13 genomic regions associated with the TBN were detected using quantitative trait locus (QTL)-seq and were located on chromosomes 2 and 5. Subsequently, validated loci within these regions were identified by QTL-seq. Three QTLs for TBN were identified in the BC4F2 populations by traditional QTL mapping, with each QTL explaining the phenotypic variation of 6.13–18.17%. In addition, for the major QTL (qTBN2-2 and qTBN5-1), residual heterozygous lines (RHLs) were developed from the BC4F2 population. These two major QTLs were verified in the RHLs by QTL mapping, with the phenotypic variation explained (PVE) of 21.57% and 30.75%, respectively. Near-isogenic lines (NILs) of qTBN2-2 and qTBN5-1 were constructed. There were significant differences between the NILs in TBN. These results will enhance our understanding of the genetic basis of TBN and provide a solid foundation for the fine-mapping of TBN.
... Chi-squared (v 2 ) analyzes were performed to check goodness of fit of observed segregations for stripe rust response with the expected ratios. Linkage between DNA markers and the resistance gene was established with MAPMARKER/EXP 3.0b (Lander et al. 1987). Markers were placed with a LOD threshold of 3.0 and a maximum distance of 30 cM. ...
Stripe rust, caused by Puccinia striiformis f. sp. tritici, is one of the most devastating foliar diseases of common wheat (Triticum aestivum L.) worldwide. Growing resistant cultivars is the most effective approach to control the disease. To determine inheritance of stripe rust resistance and map the resistance gene in a common wheat line D31, developed from Triticum sphaerococcum Perc. (accession number AS348), F 1 , F 2 , and BC 1 progenies derived from the Taichung 29 9 D31 cross were firstly inoculated with Chinese PST race CYR32 during whole growth stages under the field conditions. Genetic analysis indicated that the resistance to CYR32 in the line D31 was conferred by one recessive gene, temporarily designated as YrSph. A total of 400 simple sequence repeat (SSR), 315 pairs of sequence-related amplified polymorphism and 42 pairs of target region amplified polymorphism markers were screened, and four SSR markers and three TRAP markers were found to be polymorphic between the resistant and susceptible DNA bulks as well as their parents. Genetic linkage was tested on segregating F 2 population and indicated that all of the ten markers were linked to the resistance gene, two of which flanked the locus at 8.5 and 6.9 cM, respectively. The SSR markers mapped the resistance gene on chromosome arm 2AS. The results of chromosome location and pedigree analysis indicate that YrSph was probably a novel stripe rust resistance gene.
... The two parental maps established by Saintagne et al. [12] using Mapmaker 2.0 [29] were first reconstructed ( . The total map lengths were however quite different (929 vs. 728.8 ...
... The nine markers (Gm03_3391237_A_G, Gm03_3828735_G_A, Gm03_4487138_A_C, Gm03_5451606_A_C, Gm03_5808835_C_T, Gm03_6844115_A_C, Gm_8205334_A_G, Gm03_14228358_T_C, and Gm03_22189671_C_T) were then used to genotype all the F2:3 families to map the resistance gene. Linkage maps were constructed using MAPMAKER/EXP Version 3.0 (Lander et al. 1987), and linkage groups were determined using a logarithm of the odds (LOD) score of 3.0b, with ...
Phytophthora root rot (PRR) is one of the most important diseases in soybean (Glycine max). PRR is mainly caused by Phytophthora sojae, but recent studies showed that P. sansomeana is another causal agent of PRR with more aggressive symptoms, especially in seeds and seedlings. Unlike P. sojae which can be effectively managed by Rps genes, no known resistance genes have yet been reported for P. sansomeana. Our previous study screened 470 soybean germplasm for resistance to P. sansomeana and found that soybean ‘Colfax’ (PI 573008) carries major resistance to the pathogen. In this study, we crossed ‘Colfax’ with a susceptible parent ‘Senaki’ and developed three mapping populations with a total of 234 F2:3 families. Inheritance pattern analysis indicated a 1:2:1 ratio for resistant: segregating: susceptible lines among all the three populations, indicating a single dominant gene conferring the resistance in ‘Colfax’ (designated Rpsan1). Linkage analysis using extreme phenotypes anchored Rpsan1 to a 30Mb region on chromosome 3. By selecting 9 polymorphic SNP markers within the region, Rpsan1 was genetically delimited into a 21.3 cM region between Gm03_4487138_A_C and Gm03_5451606_A_C, which corresponds to a 1.06 Mb genomic region containing 9 NBS-LRR genes based on Gmax2.0 assembly. The mapping results were then validated using two breeding populations derived from ‘E12076T-03’ × ‘Colfax’ and ‘E16099’ × ‘Colfax’. Marker-assisted resistance spectrum analyses with 9 more isolates of P. sansomeana indicated that Rpsan1 carries a broad resistance to P. sansomeana isolates and has strong merit in protecting soybean from PRR in the future.
... with the default settings. Mapmarker/ exp3.0 software was used to reconstruct a linkage map (Lander et al. 1987). ...
... with the default settings. Mapmarker/ exp3.0 software was used to reconstruct a linkage map (Lander et al. 1987). ...
Genomic deletions are pervasive in the maize (Zea mays L.) genome, and play important roles in phenotypic variation and adaptive evolution. However, little is known about the biological functions of these genomic deletions. Here, we report the biological function of a megabase-scale deletion, which we identified by position-based cloning of the multi-trait weakened (muw) mutant, which is inherited as a single recessive locus. MUW was mapped to a 5.16-Mb region on chromosome 2. The 5.16-Mb deletion in the muw mutant led to the loss of 48 genes and was responsible for a set of phenotypic abnormities, including wilting leaves, poor yield performance, reduced plant height, increased stomatal density, and rapid water loss. While muw appears to have resulted from double-stranded break repair that was not dependent on intragenomic DNA homology, extensive duplication of maize genes may have mitigated its effects and facilitated its survival.
... For each segregating marker, a Chi-square analysis 2 -Expected)/ Expected} was performed to test for deviation from the expected segregation ratio (1:2:1). Linkage analysis of SSR markers was conducted using the Kosambi (1944) mapping function with a minimum log10 odds ratio (LOD) of 2.0 and maximum recombination frequency of 0.4 performed by Map-Maker/EXP 3.0 (Lander et al. 1987). Quantitative trait loci (QTL) analysis for each individual environment and a combined one, across all environments were performed by composite interval mapping using Windows QTL Cartographer 2.5 (Basten et al. 2002). ...
Turcicum Leaf Blight (TLB) is an important maize disease which causes heavy loss every year. A study was conducted to identify quantitative trait loci (QTLs) for NCLB resistance in maize. A mapping population from F2:3 families were developed involving the two inbred lines viz., CM 212 (susceptible) and CM 338 (resistant) and evaluated in two Indian environments (Varanasi, Uttar Pradesh and Nagenahalli, Karnataka) to locate and map QTLs. Data on four disease severity traits viz., Percent Disease Index (PDI), Area under Disease Progress Curve (AUDPC-PDI) based on PDI, Lesion Area (LA), AUDPC based on lesion area (AUDPC-LA) were generated for QTL mapping. Four QTLs were identified on 4 th chromosome of maize.
... Too little genetic difference REVIEW between parents will reduce parental DNA polymorphism, and too much of a difference will prevent recombination of the chain loci, and even serious bias segregation [10] . The populations often used for mapping in tea plants are mainly hybrid F1 and backcross populations, etc [8,11] . For F1 populations with extreme phenotypic differences, the bulk segregant analysis (BSA) method can be used to quickly identify genomic loci that regulate the quality traits of tea plants. ...
Tea is one of the most significant non-alcoholic beverages globally due to its unique secondary metabolites. Therefore, it is essential to apply molecular technologies in conjunction with various phenotypes for candidate gene mining and identification, regulating the synthesis and degradation of secondary metabolites contributing to tea quality, in order to enhance effective tea breeding. To date, there are various tea genetic resources and numerous high-density genetic maps owing to the progress and development of the tea plant genome. In this review, we comprehensively reflect the mining and identification of quality-related candidate genes using quantitative trait loci (QTL) mapping and genome-wide association study (GWAS) in tea plants in recent years. Functional verification and promotion of these candidate genes were also discussed.
... Chi-square (χ 2 ) tests were used to determine the goodness of fit for the observed segregation and expected ratios of the F 2 and F 2:3 populations. Linkage analysis was performed by using MAPMAKER/EXP v3.0b [75]. The Kosambi function was used to convert recombination values into genetic distances [76]. ...
Lodging is one of the most important factors affecting the high and stable yield of wheat worldwide. Solid-stemmed wheat has higher stem strength and lodging resistance than hollow-stemmed wheat does. There are many solid-stemmed varieties, landraces, and old varieties of durum wheat. However, the transfer of solid stem genes from durum wheat is suppressed by a suppressor gene located on chromosome 3D in common wheat, and only hollow-stemmed lines have been created. However, synthetic hexaploid wheat can serve as a bridge for transferring solid stem genes from tetraploid wheat to common wheat. In this study, the F1, F2, and F2:3 generations of a cross between solid-stemmed Syn-SAU-119 and semisolid-stemmed Syn-SAU-117 were developed. A single dominant gene, which was tentatively designated Su-TdDof and suppresses stem solidity, was identified in synthetic hexaploid wheat Syn-SAU-117 by using genetic analysis. By using bulked segregant RNA-seq (BSR-seq) analysis, Su-TdDof was mapped to chromosome 7DS and flanked by markers KASP-669 and KASP-1055 within a 4.53 cM genetic interval corresponding to 3.86 Mb and 2.29 Mb physical regions in the Chinese Spring (IWGSC RefSeq v1.1) and Ae. tauschii (AL8/78 v4.0) genomes, respectively, in which three genes related to solid stem development were annotated. Su-TdDof differed from a previously reported solid stem suppressor gene based on its origin and position. Su-TdDof would provide a valuable example for research on the suppression phenomenon. The flanking markers developed in this study might be useful for screening Ae. tauschii accessions with no suppressor gene (Su-TdDof) to develop more synthetic hexaploid wheat lines for the breeding of lodging resistance in wheat and further cloning the suppressor gene Su-TdDof.
... Chi-square (χ 2 ) tests were used to determine the goodness of fit for the observed segregation and expected ratios of the F2 and F2:3 populations. Linkage analysis was performed using MAPMAKER/EXP v3.0b [40]. The Kosambi function was used to convert recombination values to genetic distances [41]. ...
Lodging is one of the important factors affecting the high and stable yield of wheat worldwide. Solid-stemmed wheat has higher stem strength and lodging resistance than hollow-stemmed wheat. There are many solid stemmed varieties, landraces and old varieties of durum wheat. However, the transfer of solid stem genes from durum wheat is suppressed by the suppressor gene located on chromosome 3D in common wheat, and only hollow-stemmed lines have been created. However, synthetic hexaploid wheat can serve as a bridge to transfer solid stem genes from tetraploid wheat to common wheat. In this study, the F1, F2, and F2:3 generations of the cross between solid-stemmed Syn-SAU-119 and semisolid-stemmed Syn-SAU-117 were developed. A single dominant gene, tentatively designated Su-TdDof, was identified in synthetic hexaploid wheat Syn-SAU-117 by genetic analysis, which suppresses stem solidity. Using bulked segregant RNA-seq (BSR-seq) analysis, Su-TdDof was mapped to chromosome 7DS and flanked by markers KASP-669 and KASP-1055 within a 4.53 cM genetic interval corresponding to 3.86 Mb and 2.29Mb physical region in the Chinese Spring (IWGSC RefSeq v1.1) and Ae. tauschii (AL8/78 v4.0) genome, respectively, in which three genes related to solid stem development were annotated. Su-TdDof differed from a previously reported solid stem suppressor gene based on its origin and position. Su-TdDof would provide a valuable example for research on the suppression phenomenon. The flanking markers developed in this study would be useful for screening Ae. tauschii accessions with no suppressor gene (Su-TdDof) to develop more synthetic hexaploid wheat lines for wheat lodging resistance breeding and further cloning the suppressor gene Su-TdDof.
... These new SNPs and SSR markers were used to screen the BC 4 F 2:3 population. Three-Point Analysis Mechanism was applied by using the MAPMAKER/ EXP Version 3.0b (Lander et al., 1987). Kosambi's mapping function was used to determine the order of the markers (Kosambi, 1943). ...
Spike length (SL) and spike compactness (SC) are crucial traits related to wheat (Triticum aestivum L.) yield potential. In this study, a backcrossed inbred lines (BILs) population segregating for SL/SC was developed by using a commercial variety chuanyu25 as recurrent parent and a backbone parent Chuanyu12D7. Bulked segregant analysis (BSA) combined with the Wheat 660K SNP array was performed to conduct quantitative trait locus (QTL) mapping. A major and stable SL/SC QTL (designated as QSl/Sc.cib-2D.1) was identified on chromosome 2DS, explaining 45.63-59.72% of the phenotypic variation. QSl/Sc.cib-2D.1 was mapped to a 102.29-Kb interval by flanking SNPs AX-110276364 and AX-111593853 using a BC4F2:3 population. Since QSl/Sc.cib-2D.1 is linked to the Rht8 gene, their additive effects on plant type and spike type were analysed. Remarkably, the superior allele of QSl/Sc.cib-2D.1 combined with Rht8 can increase SL and TGW, and decrese SC without any apparent trade-offs in other yield-related traits. In addition, the closely linked kompetitive allele-specific PCR (KASP) markers of this locus were developed for marker-assisted selection (MAS) breeding. Four genes within the physical interval were considered as potential candidates based on expression patterns as well as orthologous gene functions. These results laid the foundation for map-based cloning of the gene(s) underlying QSl/Sc.cib-2D.1 and its potential application in wheat ideotype breeding.
... MAPMAKER/EXP 3.0b [31] was used to determine the order of the SSR marker loci in the mapping populations. The Kosambi mapping function was used to convert the recombination values into map distances (cM). ...
Asian soybean rust (ASR), caused by Phakopsora pachyrhizi, is one of the most serious soybean (Glycine max) diseases in tropical and subtropical regions. To facilitate the development of resistant varieties using gene pyramiding, DNA markers closely linked to seven resistance genes, namely, Rpp1, Rpp1-b, Rpp2, Rpp3, Rpp4, Rpp5, and Rpp6, were identified. Linkage analysis of resistance-related traits and marker genotypes using 13 segregating populations of ASR resistance, including eight previously published by our group and five newly developed populations, identified the resistance loci with markers at intervals of less than 2.0 cM for all seven resistance genes. Inoculation was conducted of the same population with two P. pachyrhizi isolates of different virulence, and two resistant varieties, ‘Kinoshita’ and ‘Shiranui,’ previously thought to only harbor Rpp5, was found to also harbor Rpp3. Markers closely linked to the resistance loci identified in this study will be used for ASR-resistance breeding and the identification of the genes responsible for resistance.
The development of Simple Sequence Repeat markers (SSRs) for lentil has played a pivotal role in enhancing the comprehension of the lentil genome through genetic mapping. The study aimed to determine the relative positions of newly developed microsatellites to the lentil genome using an F7-derived Recombinant Inbred Lines population (RIL) of 71 individuals developed from a cross between Eston and PI320937. Molecular analysis was performed with 100 newly developed lentil SSR markers and a linkage map was constructed using MapMaker/EXP 3.0b and MapChart 2.2 software. Among the 100 SSR markers, 12 markers exhibited polymorphism, 54 markers were identified as monomorphic, and 34 markers remained unamplified. While 10 out of the 12 polymorphic markers successfully integrated into two linkage groups, covering a cumulative length of 19.2cM, two markers remained unlinked. Linkage group-1, comprised of 8 markers, spanned 4.8cM, and linkage group-2 extended over a length of 14.4cM with two markers. Despite only partially representing 2 out of the 7 chromosomes in the lentil genome, this map holds promise for future mapping studies. Through the addition of markers, it could facilitate marker-assisted selection and the identification of QTLs associated with specific agronomic traits.
Wheat blast, a devastating disease having spread recently from South America to Asia and Africa, is caused by Pyricularia oryzae (synonym of Magnaporthe oryzae) pathotype Triticum, which first emerged in Brazil in 1985. Rmg8 and Rmg7, genes for resistance to wheat blast found in common wheat and tetraploid wheat, respectively, recognize the same avirulence gene, AVR-Rmg8. Here we show that an ancestral resistance gene, which had obtained an ability to recognize AVR-Rmg8 before the differentiation of Triticum and Aegilops, has expanded its target pathogens. Molecular cloning revealed that Rmg7 was an allele of Pm4, a gene for resistance to wheat powdery mildew on 2AL, whereas Rmg8 was its homoeologue on 2BL ineffective against wheat powdery mildew. Rmg8 variants with the ability to recognize AVR-Rmg8 were distributed not only in Triticum spp. but also in Aegilops speltoides, Aegilopsumbellulata and Aegilopscomosa. This result suggests that the origin of resistance gene(s) recognizing AVR-Rmg8 dates back to the time before differentiation of A, B, S, U and M genomes, that is, ~5 Myr before the emergence of its current target, the wheat blast fungus. Phylogenetic analyses suggested that, in the evolutionary process thereafter, some of their variants gained the ability to recognize the wheat powdery mildew fungus and evolved into genes controlling dual resistance to wheat powdery mildew and wheat blast.
Bioinformatics and computational biology integrate computer science, molecular biology, statistics, mathematics, and engineering to collect, manage, and analyze biological data. This interdisciplinary synergy has led to innovative advances in our understanding of plant diseases and the development of effective strategies for disease management. Bioinformatics plays a diverse and influential role in addressing the fundamental challenges faced by plant pathologists. Bioinformatics in plant pathology is vital for analyzing pathogen genomes, achieved through DNA sequencing and annotation. Comparative genomics involves comparing different pathogen genomes to identify commonalities and differences, shedding light on the genetic basis of pathogenicity, virulence, drug resistance, and evolutionary relationships. Bioinformatics also aids in studying pathogen evolution, reconstructing their history, and tracking new variants, essential for predicting disease outbreaks and developing mitigation strategies. Additionally, bioinformatics contributes to advanced diagnostic tools, detecting specific genetic markers for rapid and accurate disease identification, crucial for minimizing crop losses and ensuring food security. This chapter introduces and uses different bioinformatics tools and resources used to study plant pathogens. Moreover, applications and methods of the tools involved in BLAST, multiple sequence alignment, phylogenetics, gene structure analysis, protein domain and motif, gene localization on the chromosome, protein expression analysis, and molecular docking and interaction networks are also discussed.
Greenbug [Schizaphis graminum (Rondani)] is a serious insect pest that not only damages cereal crops, but also transmits several destructive viruses. The emergence of new greenbug biotypes in the field makes it urgent to identify novel greenbug resistance genes in wheat. CWI 76364 (PI 703397), a synthetic hexaploid wheat (SHW) line, exhibits greenbug resistance. Evaluation of an F2:3 population from cross OK 14319 × CWI 76364 indicated that a dominant gene, designated Gb9, conditions greenbug resistance in CWI 76364. Selective genotyping of a subset of F2 plants with contrasting phenotypes by genotyping-by-sequencing identified 25 SNPs closely linked to Gb9 on chromosome arm 7DL. Ten of these SNPs were converted to Kompetitive allele-specific polymerase chain reaction (KASP) markers for genotyping the entire F2 population. Genetic analysis delimited Gb9 to a 0.6-Mb interval flanked by KASP markers located at 599,835,668 bp (Stars-KASP872) and 600,471,081 bp (Stars-KASP881) on 7DL. Gb9 was 0.5 cM distal to Stars-KASP872 and 0.5 cM proximal to Stars-KASP881. Allelism tests indicated that Gb9 is a new greenbug resistance gene which confers resistance to greenbug biotypes C, E, H, I, and TX1. TX1 is one of the most widely virulent biotypes and has overcome most known wheat greenbug resistance genes. The introgression of Gb9 into locally adapted wheat cultivars is of economic importance, and the KASP markers developed in this study can be used to tag Gb9 in cultivar development.
Rice straw presents challenges as livestock feed due to its low digestibility and the presence of chemical residues. One potential solution is to focus on breeding brittle varieties that possess disease-resistance traits. In this study, AZ1803, a brittle mutant line isolated from the IR64 mutant pool, was chosen for gene identification and breeding. The AZ1803 mutant was crossed to the TNG67 variety to generate a mapping population and to the CS11 variety for fine mapping and breeding. The gene was mapped on chr. 10 between RM467 and RM171 SSR markers and was narrowed down to RM271 and RM5392 with 600 kb proximately interval. The AZ1803 and IR64 sequencing results revealed a substitution mutant in the Exon 9th of the OsCesA7 gene, resulting in an amino acid mutation at the end of the transmembrane domain 5th of the CESA7, responsible for cellulose synthesis for the secondary cell wall. The cellulose content of AZ1803 was reduced by 25% compared with the IR64. A new brittle and disease-resistant variety was bred by using developed markers in marker-assisted selection. In addition, bending tests and bacterial blight inoculation were applied. The bacterial lesion length of the bred variety is 64% lower than that of AZ1803. The rice straw of the new variety can be used for livestock feeding, which increases farmer income and reduces pesticide residues and air pollution from straw burning.
Plant defense responses to the soil-borne fungus Verticillium longisporum causing stem stripe disease on oilseed rape (Brassica napus) are poorly understood. In this study, a population of recombinant inbred lines (RILs) using the Arabidopsis thaliana accessions Sei-0 and Can-0 was established. Composite interval mapping, transcriptome data and T-DNA mutant screening identified the NITRATE/PEPTIDE TRANSPORTER FAMILY 5.12 gene (AtNPF5.12) as being associated with disease susceptibility in Can-0. Coimmunoprecipitation revealed interaction between AtNPF5.12 and the MAJOR LATEX PROTEIN family member AtMLP6, and fluorescence microscopy confirmed interaction in the plasma membrane and endoplasmic reticulum. CRISPR/Cas9 technology was applied to mutate the NPF5.12 and MLP6 genes in B. napus. Elevated fungal growth in the npf5.12 mlp6 double mutant of both oilseed rape and Arabidopsis demonstrated their importance in defense against V. longisporum. Colonization of this fungus depends also on available nitrates in the host root. Accordingly, the negative effect of nitrate depletion on fungal growth was less pronounced in Atnpf5.12 plants with impaired nitrate transport. In addition, suberin staining revealed involvement of the NPF5.12 and MLP6 genes in suberin barrier formation. Together, these results demonstrate a dependency of multiple plant factors that lead to successful V. longisporum root infection.
Powdery mildew, caused by Blumeria graminis f. sp. tritici (Bgt), is a severe disease that affects the yield and quality of wheat. Popularization of resistant cultivars in production is the preferred strategy to control this disease. In the present study, the Chinese wheat breeding line Jimai 809 showed excellent agronomic performance and high resistance to powdery mildew at the whole growth stage. To dissect the genetic basis for this resistance, Jimai 809 was crossed with the susceptible wheat cultivar Junda 159 to produce segregation populations. Genetic analysis showed that a single dominant gene, temporarily designated PmJM809, conferred the resistance to different Bgt isolates. PmJM809 was then mapped on the chromosome arm 2BL and flanked by the markers CISSR02g-1 and CIT02g-13 with genetic distances 0.4 and 0.8 cM, respectively, corresponding to a physical interval of 704.12–708.24 Mb. PmJM809 differed from the reported Pm genes on chromosome arm 2BL in origin, resistance spectrum, physical position and/or genetic diversity of the mapping interval, also suggesting PmJM809 was located on a complex interval with multiple resistance genes. To analyze and screen the candidate gene(s) of PmJM809, six genes related to disease resistance in the candidate interval were evaluated their expression patterns using an additional set of wheat samples and time-course analysis post-inoculation of the Bgt isolate E09. As a result, four genes were speculated as the key candidate or regulatory genes. Considering its comprehensive agronomic traits and resistance findings, PmJM809 was expected to be a valuable gene resource in wheat disease resistance breeding. To efficiently transfer PmJM809 into different genetic backgrounds, 13 of 19 closely linked markers were confirmed to be suitable for marker-assisted selection. Using these markers, a series of wheat breeding lines with harmonious disease resistance and agronomic performance were selected from the crosses of Jimai 809 and several susceptible cultivars.
In mechanically harvested soybean, green stem disorder (GSD) is an undesirable trait that causes green-stained seeds, which are graded lower in Japan. To obtain DNA markers for reduced GSD, we conducted a quantitative trait locus (QTL) analysis for 2 years using F4 and F5 lines from a cross between ‘Suzuotome’ (less GSD) and ‘Fukuyutaka’ (more GSD). We validated the effect of a detected QTL for GSD by first identifying F4 or F5 plants in which one or more markers in the QTL region were heterozygous. The F5 or F6 progeny of each plant was used to form a pair consisting of two groups in which the QTL region was homozygous for either the ‘Suzuotome’ or ‘Fukuyutaka’ allele in a similar genetic background, and the two groups within each pair were compared for GSD. Over 3 years of testing, the ‘Suzuotome’ allele of a QTL on chromosome 6 was found to reduce the level of GSD. This novel QTL was mapped to the region around DNA marker W06_0130, and was not closely linked to QTLs for important agronomic traits including yield components. Using this marker, the low level of GSD from ‘Suzuotome’ could be conferred to ‘Fukuyutaka’ or other high-GSD cultivars.
Aroma or fragrance in rice is a genetically controlled trait; Its high appreciation by consumers increases the rice market price. Previous studies have revealed that the rice aroma is controlled by a specific gene called BETAINE ALDEHYDE DEHYDROGENASE ( OsBADH2 ), and mutation of this gene leads to the accumulation of an aromatic substance 2‐acetyl‐1‐pyrroline (2‐AP). The use of genetic engineering to produce aroma in commercial and cultivated hybrids is a contemporary need for molecular breeding. The current study reports the generation of aroma in the three‐line hybrid restorer line Shu‐Hui‐313 (SH313). We created knock‐out ( KO ) lines of OsBADH2 through the CRISPR/Cas9. The analysis of KO lines revealed a significantly increased content of 2AP in the grains compared with the control. However, other phenotypic traits (plant height, seed setting rate, and 1000‐grain weight) were significantly decreased. These KO lines were crossed with a non‐aromatic three‐line hybrid rice male sterile line (Rong‐7‐A) to produce Rong‐7‐You‐626 (R7Y626), R7Y627 and R7Y628. The measurement of 2‐AP revealed significantly increased contents in these cross combinations. We compared the content of 2‐AP in tissues at the booting stage. Data revealed that young spike stalk base contained the highest content of 2‐AP and can be used for identification (by simple chewing) of aromatic lines under field conditions. In conclusion, our dataset offers a genetic source and illustrates the generation of aroma in non‐aromatic hybrids, and outlines a straightforward identification under field conditions.
In eggplant, two kinds of fruit‐bearing habits can be observed, that is, cluster and solitary. The fruits grown in cluster are small in size but contribute towards yield and productivity. In this investigation, we have studied the genetics of fruit‐bearing habit and identified the SSR markers linked to this trait. Pusa Safed Baingan 1 (cluster bearer) and Pusa Hara Baingan 1 (solitary bearer) having contrasting traits were crossed to generate F 1 (20 plants), F 2 (215 plants), B 1 (73 plants) and B 2 (64 plants). Genetic analysis of the trait was carried out using Chi‐square test. The segregation of plants in F 2 suggested recessive epistasis with ratio of 9:3:4 (solitary: mixed: cluster) indicating involvement of two genes for controlling the fruit‐bearing habit. The parental polymorphic 6 SSR markers were used for genotyping F 2 plants and co‐segregated with the fruit‐bearing habit ( Bh 1 ) locus in 1:2:1. A linkage map covering 63.86 cm distance was developed, the SSR marker emf11A03 was closest with a distance of 4.37 cm, emk03O04 at 12.5 cm distance, and emf21O06 at 14.5 cm. The physical positions of the SSR markers emb01J19 and emf11A03 flanking Bh 1 gene were located at 4.77 mb position of contig Sme2.5_05721.1 and 5.18 Mb position of contig Sme2.5_00994.1, respectively. The findings in the present study will be helpful in marker‐assisted breeding to enhance yield in eggplant.
Grain size is an important appearance quality trait in rice, which also affects grain yield. In this study, a recombinant inbred line (RIL) population derived from a cross between indica variety 9311 and japonica variety Cypress was constructed. And 181 out of 600 RILs were sequenced, and a high-density genetic map containing 2842 bin markers was constructed, with a total map length of 1500.6 cM. A total of 10 quantitative trait loci (QTL) related to grain length (GL), grain width (GW), grain length-to-width ratio (LWR), and 1000-grain weight (TGW) were detected under two environments. The genetic effect of qGL4, a minor QTL for GL and TGW, was validated using three heterogeneous inbred family (HIF) segregation populations. It was further dissected into two closed linked QTL, qGL4.1 and qGL4.2. By progeny testing, qGL4.1 and qGL4.2 were successfully delimited to intervals of 1304-kb and 423-kb, respectively. Our results lay the foundation for the map-based cloning of qGL4.1 and qGL4.2 and provide new gene resources for the improvement of grain yield and quality in rice.
Stripe rust, caused by Puccinia striiformis f. sp. tritici (Pst), is one of the most destructive fungal diseases of wheat. Cultivated einkorn (Triticum monococcum ssp. monococcum, 2n=2x=14, AmAm), one of the founder crops of agriculture, harbors unexploited genetic sources for wheat improvement. An advanced wheat line Z15-1949 with 42 chromosomes, selected from the hybrids of Pst-susceptible common wheat cultivar Crocus and resistant T. monococcum accession 10-1, exhibits high resistance to a mixture of the Chinese prevalent Pst races. Genetic analysis on F1, F2, and F2:3 generations of the cross between Z15-1949 and Pst-susceptible common wheat SY95-71 indicated that the resistance of Z15-1949 was conferred by a recessive gene, tentatively designated as YrZ15-1949. This gene was mapped to the short arm of chromosome 7D using the Wheat 55K SNP array, flanked by markers KASP-1949-2 and KASP-1949-10 within a 3.3 cM genetic interval corresponding to 1.12 Mb physical region in the Chinese Spring reference genome V2.0. The gene differs from previously reported Yr genes on 7D based on their physical positions, and is probably a novel gene. YrZ15-1949 would be a valuable resource for developing Pst-resistant wheat cultivars and the linked markers could be used for the marker-assisted selection.
Tobacco (Nicotiana tabacum L.) is the non-food commercially important industrial crop valued for its leaf. Size and thickness are the very important factors in tobacco quality, which is affected by many biotic and abiotic stresses. Among production-limiting pathogens, plant-parasitic nematodes are a major concern among growers, of which Meloidogyne spp. (root-knot nematode) obligate root parasites are the most economically important, causing extensive 15 to 28% yield losses. Looking at this importance, the present study was put forth for identification of marker linked to leaf thickness as well as root-knot nematode (RKN) resistance through marker-assisted breeding with an aim to speed up the varietal development program of tobacco. A cross was made between the resistant ABT-10 and susceptible A119 genotype, and F2 mapping population was developed. Genotyping was carried out from F2 individuals, and phenotyping for RKN was carried out in F2-derived F3 family. The ANOVA revealed significant genotypic components of variance for the F2:3 families and the broad-sense heritability. The low difference between GCV and PCV was observed, which indicated the little influence of the environment on both traits. Single marker analysis was employed for identification of marker-trait association. A total of five SSR markers, namely PT30157, PT30346, PT20149, TM11008, and TM10211, showed linkage with leaf thickness, whereas four SSR markers, namely TM10083, TM10820, TM816, and PT20213, were found to be linked with RKN. The marker identified in this investigation can be exploited for speedy development of tobacco varieties with maximum leaf thickness and RKN resistant traits.
Gray leaf spot (GLS) caused by Cercospora zeina or C. zeae‐maydis is a major maize disease throughout the world. Although more than 100 QTLs resistant against GLS have been identified, very few of them have been cloned.
Here, we identified a major resistance QTL against GLS, qRglsSB, explaining 58.42% phenotypic variation in SB12×SA101 BC1F1 population. By fine‐mapping, it was narrowed down into a 928 kb region. By using transgenic lines, mutants and complementation lines, it was confirmed that the ZmWAK02 gene, encoding an RD wall‐associated kinase, is the responsible gene in qRglsSB resistant against GLS.
The introgression of the ZmWAK02 gene into hybrid lines significantly improves their grain yield in the presence of GLS pressure and does not reduce their grain yield in the absence of GLS.
In summary, we cloned a gene, ZmWAK02, conferring large effect of GLS resistance and confirmed its great value in maize breeding.
Key message
By integrating QTL fine mapping and transcriptomics, a candidate gene responsible for oil content in rapeseed was identified. The gene is anticipated to primarily function in photosynthesis and photosystem metabolism pathways.
Abstract
Brassica napus is one of the most important oil crops in the world, and enhancing seed oil content is an important goal in its genetic improvement. However, the underlying genetic basis for the important trait remains poorly understood in this crop. We previously identified a major locus, OILA5 responsible for seed oil content on chromosome A5 through genome-wide association study. To better understand the genetics of the QTL, we performed fine mapping of OILA5 with a double haploid population and a BC3F2 segregation population consisting of 6227 individuals. We narrowed down the QTL to an approximate 43 kb region with twelve annotated genes, flanked by markers ZDM389 and ZDM337. To unveil the potential candidate gene responsible for OILA5, we integrated fine mapping data with transcriptome profiling using high and low oil content near-isogenic lines. Among the candidate genes, BnaA05G0439400ZS was identified with high expression levels in both seed and silique tissues. This gene exhibited homology with AT3G09840 in Arabidopsis that was annotated as cell division cycle 48. We designed a site-specific marker based on resequencing data and confirmed its effectiveness in both natural and segregating populations. Our comprehensive results provide valuable genetic information not only enhancing our understanding of the genetic control of seed oil content but also novel germplasm for advancing high seed oil content breeding in B. napus and other oil crops.
Genetic research on insect natural enemies was, until relatively recently, rare and was essentially limited to parasitoid wasps and some coccinellid beetles.
Bean pod mottle virus (BPMV) is a comovirus that infects common bean and legumes in general. BPMV is distributed throughout the world and is a major threat on soybean, a closely related species of common bean. In common bean, BAT93 was reported to carry the R-BPMV resistance gene conferring resistance to BPMV and linked with the I resistance gene. To fine map R-BPMV, 182 recombinant inbred lines (RILs) derived from the cross BAT93 x JaloEEP558 were genotyped with polymerase-chain (PCR)-based markers developed using genome assemblies from G19833 and BAT93, as well as BAT93 BAC clone sequences. Analysis of RILs carrying key recombination events positioned R-BPMV to a target region containing at least 16 TIR-NB-LRR (TNL) sequences in BAT93. Because the I cluster presents a suppression of recombination and a large number of repeated sequences, none of the 16 TNLs could be excluded as R-BPMV candidate gene. The evolutionary history of the TNLs for the I cluster were reconstructed using microsynteny and phylogenetic analyses within the legume family. A single I TNL was present in Medicago truncatula and lost in soybean, mirroring the absence of complete BPMV resistance in soybean. Amplification of TNLs in the I cluster predates the divergence of the Phaseolus species, in agreement with the emergence of R-BPMV before the separation of the common bean wild centers of diversity. This analysis provides PCR-based markers useful in marker-assisted selection (MAS) and laid the foundation for cloning of R-BPMV resistance gene in order to transfer the resistance into soybean.
Soil salinity is one of the major environmental constraints that limits crop yield and nearly 7% of the total area worldwide is affected by salinity. Salinity-induced oxidative stress causes membrane damage during germination and seedling growth. Indian mustard is a major oilseed crop in India and its production and productivity are severely affected by salt stress. Breeding Brassica cultivars for salinity tolerance by conventional means is very difficult and time-consuming. Therefore, understanding the molecular components associated with salt tolerance is needed to facilitate breeding for salt tolerance in Brassica. In this investigation, quantitative trait loci (QTLs) associated with salt tolerance were identified using F2:3 developed from a cross between CS52 (salinity tolerant) and RH30 (salinity sensitive). Parents and F2:3 were evaluated under controlled and salinity stress conditions for 14 morpho-physiological traits for two consecutive generations (F2 and F2:3), explaining proportion of the phenotypic variance under control condition. Simple sequence repeat (SSR) markers were used for mapping studies. A genetic linkage map based on 42 simple sequence repeats (SSRs) markers was constructed covering 2298.5 cM (Haldane) to identify the loci associated with salt tolerance in Brassica juncea. Forty-one SSRs that showed polymorphism in the parents (CS52 and RH30) mapped on 8 linkage groups (C1-C8). One marker (nga 129) did not map to any of the linkage groups and was excluded from mapping. Linkage group 5 (C5; 317.9 cM) was longest and linkage group 1 (C1, 255.0 cM) was shortest. Further, we identified 15 QTLs controlling 8 traits using F2:3 population. These QTLs explained 12.44-60.63% of the phenotypic variation with a LOD score range of 3.62-5.97. Out of these QTLs, QMI4.1 related to membrane injury showed 51.28% phenotypic variance with a LOD score of 3.34. QTL QBYP8.1 related to biological yield per plant showed 60.63% phenotypic variance at a LOD score of 3.62. The highest LOD score of 5.97 was recorded for QTL related to seed yield per plant (QSYP4.1). Major QTL were biological yield per plant (QBYP8.1), QTL for siliquae per plant (QSP4.1), QTL for primary branches (QPB4.1), QTLs for seed per siliqua (QSS4.1, QSS4.2), QTL for seed yield per plant (QSYP4.1), and QTL for membrane injury (QMI8.1) which showed more than 50% phenotypic variance. These QTLs identified in our study need to be confirmed in other populations as well so that these can be used in marker-assisted selection and breeding to enhance salt tolerance in Brassica juncea.
Global warming poses a threat to rice production. Breeding heat-tolerant rice is an effective and economical approach to addressing this challenge. African rice is a valuable genetic resource for developing heat-tolerant crops due to its intricate mechanism for adapting to high temperatures. Oryza longistaminata, a wild rice species widely distributed in Africa, may harbor an even richer gene pool for heat tolerance, which remains untapped. In this study, we identified three heat-tolerant QTLs from O.longistaminata at the seedling stage. Among these, qTT4 and qTT5 are novel heat-tolerant loci identified in O. longistaminata. Our findings demonstrated that the O. longistaminata alleles for these two QTLs can enhance the heat tolerance of rice seedlings. Remarkably, qTT5 was mapped to a region spanning approximately 287.2 kb, containing 46 expressing genes. Through the analysis of Gene Ontology and expression differences under heat induction, we identified four candidate genes. Our results lay the foundation for the discovery of heat-tolerant genes underlying O. longistaminata and the development of new genetic resources for heat tolerant rice breeding.
Key words: wild rice; Oryza longistaminata; heat tolerance; QTL
Plant breeding plays a crucial role in improving plant genotypes and ensuring global food security. However, traditional breeding methods have limitations in effectively selecting traits, particularly quantitative traits, and can be time-consuming and unreliable. Breeders have introduced molecular markers into plant breeding practices to address these problems, resulting in the fields of “smart breeding,” “molecular breeding,” and “marker-assisted breeding.” To pinpoint the genetic areas that regulate desired features, a number of DNA-based molecular marker systems, including RFLP, RAPD, AFLP, SSR, and SNP, have been created. These markers eliminate the need for phenotype-based selection by streamlining the identification of target genomic areas based on genotypes.
The advances in genomics and bioinformatics have made possible the study in non-model plants of phenotypes associated to flower development. Floriculture crops are an interesting source of traits associated to flower development such as the transition between zygomorphic and actinomorphic flowers or the production of flowers with double and triple corollas. In this chapter, we summarize the material and methods for the use of floriculture crops to study flower development using genomic tools, from the sequencing and assembly of a reference genome to QTL and RNA-Seq analysis to search candidate genes associated to specific traits.
As modern crop cultivars have narrow genetic base which is a real barrier to maintain and improve crop productivity, are more vulnerable to possibly new biotic and abiotic stresses. Plant germplasm resources are valuable repositories of natural genetic differences that can be used to broaden the genetic base of the cultivars. Even though, number of agriculturally essential traits i.e., yield, quality, resistance to disease and environmental stresses are quantitative in nature. The genetic variation of a quantitative trait is governed by the combined effects of several genes, referred as quantitative trait loci (QTLs). Detection of QTLs of economic value and its application in a crop breeding involves mapping of these QTLs using molecular markers with in the genome of different crops. Due to developments in molecular markers and statistical genomics, QTL mapping in crop plants is now a regular practice. As a result, multiple QTLs have been located in various crops for a wide range of attributes. There are now a number of statistical programs that are appropriate for the type of mapping population, the trait(s) being investigated for QTL studies and the objective of the study. Simpler techniques like single marker analysis and simple interval mapping as well as more extensive ones like composite interval mapping and multiple interval mapping are included among these methods. The comparative importance of each of these strategies differs enormously. This chapter offers a summary of the various approaches to linkage-based QTL analysis.
Background
The structural basis of chloroplast and the regulation of chloroplast biogenesis remain largely unknown in maize. Gene mutations in these pathways have been linked to the abnormal leaf color phenotype observed in some mutants. Large scale structure variants (SVs) are crucial for genome evolution, but few validated SVs have been reported in maize and little is known about their functions though they are abundant in maize genomes.
Results
In this research, a spontaneous maize mutant, pale green leaf-shandong (pgl-sd), was studied. Genetic analysis showed that the phenotype of pale green leaf was controlled by a recessive Mendel factor mapped to a 156.8-kb interval on the chromosome 1 delineated by molecular markers gy546 and gy548. There were 7 annotated genes in this interval. Reverse transcription quantitative PCR analysis, SV prediction, and de novo assembly of pgl-sd genome revealed that a 137.8-kb deletion, which was verified by Sanger sequencing, might cause the pgl-sd phenotype. This deletion contained 5 annotated genes, three of which, including Zm00001eb031870, Zm00001eb031890 and Zm00001eb031900, were possibly related to the chloroplast development. Zm00001eb031870, encoding a Degradation of Periplasmic Proteins (Deg) homolog, and Zm00001eb031900, putatively encoding a plastid pyruvate dehydrogenase complex E1 component subunit beta (ptPDC-E1-β), might be the major causative genes for the pgl-sd mutant phenotype. Plastid Degs play roles in protecting the vital photosynthetic machinery and ptPDCs provide acetyl-CoA and NADH for fatty acid biosynthesis in plastids, which were different from functions of other isolated maize leaf color associated genes. The other two genes in the deletion were possibly associated with DNA repair and disease resistance, respectively. The pgl-sd mutation decreased contents of chlorophyll a, chlorophyll b, carotenoids by 37.2%, 22.1%, and 59.8%, respectively, and led to abnormal chloroplast. RNA-seq revealed that the transcription of several other genes involved in the structure and function of chloroplast was affected in the mutant.
Conclusions
It was identified that a 137.8-kb deletion causes the pgl-sd phenotype. Three genes in this deletion were possibly related to the chloroplast development, which may play roles different from that of other isolated maize leaf color associated genes.
Powdery mildew, caused by Blumeria graminis f. sp. tritici (Bgt), is a destructive disease worldwide. Host resistance is the preferred method for limiting the disease epidemic, protecting environment, and minimizing the economic losses. In the present study, the reactions to powdery mildew for a collection of 600 wheat cultivars and breeding lines from different wheat growing regions were tested using the Bgt isolate E09. One hundred and sixteen resistant genotypes were identified and then crossed with susceptible wheat cultivars/lines to produce segregating populations for genetic analysis. Among them, 87, 19, and 10 genotypes displayed single, dual, and multiple genic inheritance, respectively. To identify the Pm gene(s) in those resistant genotypes, 16 molecular markers for 13 documented Pm genes were used to test the resistant and susceptible parents and their segregating populations. Of the 87 wheat genotypes that fitted the monogenic inheritance, 75 ones carried the Pm2a allele. Three, two, one, and two genotypes carried Pm21, Pm6, Pm4, and the recessive genes pm6 and pm42, respectively. Four genotypes did not carry any of the tested genes, suggesting that they might have other uncharacterized or new genes. The other 29 wheat cultivars/lines carried two or more of the tested Pm genes and/or other untested genes, including Pm2, Pm5, Pm6, and/or pm42 . It was obvious that Pm2 was widely used in wheat production, whereas Pm1, Pm24, Pm33, Pm34, Pm35, Pm45, and Pm47 were not detected in any of these resistant wheat genotypes. This study clarified the genetic basis of the powdery mildew resistance of these wheat cultivars/lines to provide information for their rational utilization in different wheat growing regions. Moreover, some wheat genotypes which may have novel Pm gene(s) were mined to enrich the diversity of resistance source.
The history of the modern oat may be traced to its intergeneric contamination of wheat and barley in southwest Asia, particularly in Mesopotamia and due to its use in only forage crop rather than a staple food in most of the countries it’s been neglected. There are just a few economically cultivated species of this self-pollinated crop, including A. sativa, A. nuda, and A. byzantina. After rice, wheat, maize, sorghum, and barley, oat production comes in at number six and in terms of oat production (FAO 2019). The present review was undertaken in order to address the present scenario of oats utilization and its consideration in research. The work majorly focussed in present are β glucan (due to its nutritional status) and disease resistance (majorly powdery mildew, pyrenophora leaf spot and crown rust) throughout the world but some drawbacks is that oats is usually neglected because of its limited use as forage value and also of its hexaploid nature. The different breeding methods employed till date for crop improvement are described in the chapter viz, traditional methods, diversity studies, mapping strategies, Marker aided breeding Association mapping, Genomic assisted breeding etc. Some of the qtls mapped by various researchers for different traits is also been listed in this article. Apart from these studies, the novel techniques which are still lacking in oats has much scope now to get it utilized through various improvement programmes like in other cereal crops.
DNA markers and sampling of three-generation families can be used to construct complete linkage maps of human chromosomes. This is important in mapping disease loci and in determining the genetic or environmental component of a disease.
Assuming random mating and random sampling of pedigrees, the likelihood of a set of pedigree data is developed in terms of: (1) the population distribution of the different genotypes; (2) the phenotypic distributions for the different genotypes, and (3) the genotypic distribution of offspring given the parents’ genotypes. This last is given for any number of unlinked autosomal loci, two linked autosomal loci, an X-linked locus, and combinations of these possibilities. Methods are given for using this likelihood to test specific genetic hypotheses and for genetic counselling.Copyright © 1971 S. Karger AG, Basel
A detailed linkage map of lettuce was constructed using 53 genetic markers including 41 restriction fragment length polymorphism (RFLP) loci, five downy mildew resistance genes, four isozyme loci and three morphological markers. The genetic markers were distributed into nine linkage groups and cover 404 cM which may be 25-30% of the lettuce genome. The majority (31 of 34) of the RFLP probes detected single segregating loci, although seven of these may have been homologous to further monomorphic loci. When several loci were detected by a single probe, the loci were generally linked, suggesting tandem duplications. One probe, however, detected loci in three linkage groups suggesting translocations. The five downy mildew resistance genes (Dm1, Dm3, Dm4, Dm5/8 and Dm13), segregating in the Calmar x Kordaat cross, represented each of the four resistance gene linkage groups. Dm5/8 is flanked by two cDNA loci, each located 10 cM away. These flanking markers will be used to study the source of variation in downy mildew genes and are also part our strategy to clone resistance genes.
High-density genetic maps of individual human chromosomes will permit
accurate localization of disease-associated genes and provide signposts
that will be useful in the construction of physical maps. We have
constructed a genetic map of chromosome 7 with 63 polymorphic DNA
markers by using segregation data from 23 three-generation families and
recently developed multilocus linkage-analysis techniques. The map spans
250 centimorgans in females and 170 centimorgans in males, with much of
the difference being concentrated in a few intervals. The density and
informativeness of the markers are such that there is a high probability
of detecting linkage to any disease gene on this chromosome for which 20
phase-known meioses are available.
The articles published by the Annals of Eugenics (1925–1954) have been made available online as an historical archive intended for scholarly use. The work of eugenicists was often pervaded by prejudice against racial, ethnic and disabled groups. The online publication of this material for scholarly research purposes is not an endorsement of those views nor a promotion of eugenics in any way.
High-density genetic maps of individual human chromosomes will permit accurate localization of disease-associated genes and provide signposts that will be useful in the construction of physical maps. We have constructed a genetic map of chromosome 7 with 63 polymorphic DNA markers by using segregation data from 23 three-generation families and recently developed multilocus linkage-analysis techniques. The map spans 250 centimorgans in females and 170 centimorgans in males, with much of the difference being concentrated in a few intervals. The density and informativeness of the markers are such that there is a high probability of detecting linkage to any disease gene on this chromosome for which 20 phase-known meioses are available.
A formula is given for the advantage of n-point sampling, which approaches infinity with n. However, 2-point and 3-point analyses extract nearly all the information in such samples and at the same time communicate this information as lods and chi 2, which can be combined with other data by simple addition without reevaluation of the likelihood. When null interference is assumed, map distances and multiple recombination frequencies are inflated, and there is substantial loss of efficiency and of support for the correct order.
The principles of linkage detection and measurement are traced from the first discovery of linkage to its present-day use in human genetics. Some indications are given of their success and of the present problems and challenges facing them.
Human genetic linkage maps are most accurately constructed by using information from many loci simultaneously. Traditional methods for such multilocus linkage analysis are computationally prohibitive in general, even with supercomputers. The problem has acquired practical importance because of the current international collaboration aimed at constructing a complete human linkage map of DNA markers through the study of three-generation pedigrees. We describe here several alternative algorithms for constructing human linkage maps given a specified gene order. One method allows maximum-likelihood multilocus linkage maps for dozens of DNA markers in such three-generation pedigrees to be constructed in minutes.
Methods are given for efficient calculation of the likelihood for multilocus linkage in families comprised of grandparents, parents, and children. Such families are being used in large-scale cooperative efforts to build a detailed linkage map of the human genome. The methods are illustrated by an application to loci on chromosome 13.
We describe a new basis for the construction of a genetic linkage map of the human genome. The basic principle of the mapping scheme is to develop, by recombinant DNA techniques, random single-copy DNA probes capable of detecting DNA sequence polymorphisms, when hybridized to restriction digests of an individual's DNA. Each of these probes will define a locus. Loci can be expanded or contracted to include more or less polymorphism by further application of recombinant DNA technology. Suitably polymorphic loci can be tested for linkage relationships in human pedigrees by established methods; and loci can be arranged into linkage groups to form a true genetic map of "DNA marker loci." Pedigrees in which inherited traits are known to be segregating can then be analyzed, making possible the mapping of the gene(s) responsible for the trait with respect to the DNA marker loci, without requiring direct access to a specified gene's DNA. For inherited diseases mapped in this way, linked DNA marker loci can be used predictively for genetic counseling.
A computer program that calculates lod scores and genetic risks for a wide variety of both qualitative and quantitative genetic traits is discussed. An illustration is given of the joint use of a genetic marker, affection status, and quantitative information in counseling situations regarding Duchenne muscular dystrophy.
AsstRAcT: LINKAGE-1 is a PASCAL computer program Designed to aid the geneticist in the detection and esti--nation of linkage in segregating progenies. Loci segregating for both dominant and codominant genes ;an be simultaneously analyzed. Goodness-of-fit to expected ratios for single-factor segregation are tested mIy chi-square analyses. Contingency chi-square anal-yses are used to test for independent assortment be-Tween all pairs of jointly segregating loci. If significant deviation is detected, recombination percentages and 'heir standard errors are calculated. The program and ,nstructions for ts use are available from the authors. IN GENE SEGREGATION studies the estimation of recombination intensities between linked loci is most accurately provided by the method of maximum likelihood as discussed by Allard and Mather 3 . In progenies where numerous loci are simultaneously segregating, the number of locus pairs to be sorted and tested for linkage can be quite large. The required data manipulations resulting from such progenies are tedious and time consuming. In order to facilitate segregation and linkage analyses, we have developed LINKAGE-I, a computer program that performs many of the steps routinely employed in the detection of linkage and the calculation of its intensity.
A linkage map in tomato has been developed based on isozyme and random cDNA clones derived from mRNA. Interspecific backcross and F(2) populations of Lycopersicon esculentum and L. pennellii were employed in the linkage analysis. Allelic differences in cDNA markers were based on restriction fragment length polymorphisms detected through Southern analysis. A total of 57 unique cDNA clones have been analyzed. The majority of cDNA markers correspond to single loci and are dispersed throughtout the genome. Of those clones that hybridize to two or more loci, most show genetic independence (ie., they are unlinked). The combination of isozyme, cDNA and previously mapped DNA markers total 112 loci. It is estimated that approximately 92% of the genome can be monitored during segregation with these markers. Molecular maps, such as the one being constructed in tomato, may allow genetic and breeding experiments that previously were not possible.
Analysis of Human Genetic Linkage Identification of more than 566 RFLPs by random screening: 8th Human Gene Mapping Workshop The development of human linkage analysis
- J Ott
- Md Schumm
- J Knowlton
- R Braman
- J Barker
- D Vovis
- G Akots
- G Brown
- V Gravius
- T Helms
- C Rjzdiker
- K Thurston
- J Botstein
- D And Donis-Keller
OTT, J. (1985) " Analysis of Human Genetic Linkage, " Johns Hopkins Press, Baltimore, MD. SCHUMM, J., KNOWLTON, R., BRAMAN, J., BARKER, D., VOVIS, G., AKOTS, G., BROWN, V., GRAVIUS, T., HELMS, C., RJZDIKER, K., THURSTON, J., BOTSTEIN, D., AND DONIS-KELLER, H. (1985). Identification of more than 566 RFLPs by random screening: 8th Human Gene Mapping Workshop. Cytogenet. Cell Genet. 40: 576. SMITH, C. A. B. (1986). The development of human linkage analysis. Ann. Hum. Genet. 50: 293-311.