Limits...
Development of genome-wide informative simple sequence repeat markers for large-scale genotyping applications in chickpea and development of web resource.

Parida SK, Verma M, Yadav SK, Ambawat S, Das S, Garg R, Jain M - Front Plant Sci (2015)

Bottom Line: These physically mapped SSR markers exhibited robust amplification efficiency (73.9%) and high intra- and inter-specific polymorphic potential (63.5%), thereby suggesting their immense use in various genomics-assisted breeding applications.The SSR markers particularly derived from intergenic and intronic sequences revealed high polymorphic potential.The intra-specific polymorphism (47.6%) and functional molecular diversity (65%) potential of polymorphic SSR markers developed in our study is much higher than that of previous documentations.

View Article: PubMed Central - PubMed

Affiliation: Functional and Applied Genomics Laboratory, National Institute of Plant Genome Research New Delhi, India.

ABSTRACT
Development of informative polymorphic simple sequence repeat (SSR) markers at a genome-wide scale is essential for efficient large-scale genotyping applications. We identified genome-wide 1835 SSRs showing polymorphism between desi and kabuli chickpea. A total of 1470 polymorphic SSR markers from diverse coding and non-coding regions of the chickpea genome were developed. These physically mapped SSR markers exhibited robust amplification efficiency (73.9%) and high intra- and inter-specific polymorphic potential (63.5%), thereby suggesting their immense use in various genomics-assisted breeding applications. The SSR markers particularly derived from intergenic and intronic sequences revealed high polymorphic potential. Using the mapped SSR markers, a wider functional molecular diversity (16-94%, mean: 68%), and parentage- and cultivar-specific admixed domestication pattern and phylogenetic relationships in a structured population of desi and kabuli chickpea genotypes was evident. The intra-specific polymorphism (47.6%) and functional molecular diversity (65%) potential of polymorphic SSR markers developed in our study is much higher than that of previous documentations. Finally, we have developed a user-friendly web resource, Chickpea Microsatellite Database (CMsDB; http://www.nipgr.res.in/CMsDB.html), which provides public access to the data and results reported in this study. The developed informative SSR markers can serve as a resource for various genotyping applications, including genetic enhancement studies in chickpea.

No MeSH data available.


Validation of amplification and polymorphic potential of selected SSRs and allelic variations in chickpea genotypes. (A) Polymerase chain reaction based validation of amplification and polymorphic potential of selected SSRs in chickpea genotypes. Only five selected examples of SSRs have been represented. Representative gels showing PCR amplification of polymorphic SSRs (labeled on right side) validating the length polymorphism between desi and kabuli chickpea genotypes (genotype details are available in Supplementary Table S3). M, 50 bp DNA ladder as size standard. (B) Eight different representative allele types identified based on the fragment length polymorphism pattern of 160 SSR markers across desi and kabuli chickpea genotypes. The SSR marker-alleles are illustrated according to their lower to higher fragment size (bp).
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4543896&req=5

Figure 5: Validation of amplification and polymorphic potential of selected SSRs and allelic variations in chickpea genotypes. (A) Polymerase chain reaction based validation of amplification and polymorphic potential of selected SSRs in chickpea genotypes. Only five selected examples of SSRs have been represented. Representative gels showing PCR amplification of polymorphic SSRs (labeled on right side) validating the length polymorphism between desi and kabuli chickpea genotypes (genotype details are available in Supplementary Table S3). M, 50 bp DNA ladder as size standard. (B) Eight different representative allele types identified based on the fragment length polymorphism pattern of 160 SSR markers across desi and kabuli chickpea genotypes. The SSR marker-alleles are illustrated according to their lower to higher fragment size (bp).

Mentions: We utilized 341 SSR markers (revealing ≥4-bp fragment length polymorphism between desi and kabuli chickpea) in total located in different components of genes (upstream, 5′-UTRs, CDS, introns and 3′-UTRs) and intergenic regions to evaluate their amplification efficiency as well as potential for detecting polymorphism among 31 desi and 15 kabuli chickpea genotypes (Supplementary Table S3). Two hundred fifty-two of the 341 markers gave successful PCR amplification in all 46 chickpea genotypes with an amplification success rate of 73.9% (Supplementary Table S2). One hundred sixty (63.5%) of 252 amplified markers showed polymorphism in at least two combinations of chickpea genotypes (Figure 5A). It included 130 (73.9%, mean PIC: 0.76) of 176 class I and 30 (39.5%, 0.69) of 76 class II SSR markers. The remaining 92 (36.5%) markers exhibited monomorphic amplification among chickpea genotypes used. A total number of 764 alleles were amplified by 160 polymorphic SSR markers with a mean allele number of 4.8. The number of alleles amplified per locus varied from 2 to 12. The PIC ranged from 0.23 to 0.86 with an average of 0.75, while gene diversity varied from 0.25 to 0.89 with a mean of 0.77 (Supplementary Table S2). The polymorphic potential of markers in different sequence components of genes and intergenic regions was analyzed in detail based on the percent polymorphism, PIC and polymorphic alleles amplified among chickpea genotypes. We were able to detect polymorphism in 55 (62.5%, allele number from 2–9 and PIC 0.74) of 88 markers derived from the different coding and non-CDS components of genes between desi and kabuli chickpea. The remaining 105 (64%, 2–12 and 0.63) of 164 markers derived from intergenic regions also showed polymorphism between the two chickpea types (Supplementary Table S2). Within genes, maximum potential of polymorphism was detected by the markers developed from intronic sequences (28 of 38 markers, 73.7%, allele number 2–9 and PIC 0.74) followed by 3′-UTRs (5 of 8 markers, 62.5%, 2–4 and 0.66), upstream (19 of 35 markers, 54.3%, 2–5 and 0.76) and 5′-UTRs (2 of 6 markers, 33.3%, 4–5 and 0.62) of genes (Supplementary Table S2, Figure 5A). Remarkably, 120 (47.6%, 2–10 and 0.62) and 107 (42.5%, 2–8 and 0.57) markers revealed polymorphism within desi and kabuli chickpea genotypes too, respectively. Overall, eight different representative allele types were detected based on fragment length polymorphism patterns of all the 160 SSR markers in 46 desi and kabuli genotypes (Figure 5B).


Development of genome-wide informative simple sequence repeat markers for large-scale genotyping applications in chickpea and development of web resource.

Parida SK, Verma M, Yadav SK, Ambawat S, Das S, Garg R, Jain M - Front Plant Sci (2015)

Validation of amplification and polymorphic potential of selected SSRs and allelic variations in chickpea genotypes. (A) Polymerase chain reaction based validation of amplification and polymorphic potential of selected SSRs in chickpea genotypes. Only five selected examples of SSRs have been represented. Representative gels showing PCR amplification of polymorphic SSRs (labeled on right side) validating the length polymorphism between desi and kabuli chickpea genotypes (genotype details are available in Supplementary Table S3). M, 50 bp DNA ladder as size standard. (B) Eight different representative allele types identified based on the fragment length polymorphism pattern of 160 SSR markers across desi and kabuli chickpea genotypes. The SSR marker-alleles are illustrated according to their lower to higher fragment size (bp).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4543896&req=5

Figure 5: Validation of amplification and polymorphic potential of selected SSRs and allelic variations in chickpea genotypes. (A) Polymerase chain reaction based validation of amplification and polymorphic potential of selected SSRs in chickpea genotypes. Only five selected examples of SSRs have been represented. Representative gels showing PCR amplification of polymorphic SSRs (labeled on right side) validating the length polymorphism between desi and kabuli chickpea genotypes (genotype details are available in Supplementary Table S3). M, 50 bp DNA ladder as size standard. (B) Eight different representative allele types identified based on the fragment length polymorphism pattern of 160 SSR markers across desi and kabuli chickpea genotypes. The SSR marker-alleles are illustrated according to their lower to higher fragment size (bp).
Mentions: We utilized 341 SSR markers (revealing ≥4-bp fragment length polymorphism between desi and kabuli chickpea) in total located in different components of genes (upstream, 5′-UTRs, CDS, introns and 3′-UTRs) and intergenic regions to evaluate their amplification efficiency as well as potential for detecting polymorphism among 31 desi and 15 kabuli chickpea genotypes (Supplementary Table S3). Two hundred fifty-two of the 341 markers gave successful PCR amplification in all 46 chickpea genotypes with an amplification success rate of 73.9% (Supplementary Table S2). One hundred sixty (63.5%) of 252 amplified markers showed polymorphism in at least two combinations of chickpea genotypes (Figure 5A). It included 130 (73.9%, mean PIC: 0.76) of 176 class I and 30 (39.5%, 0.69) of 76 class II SSR markers. The remaining 92 (36.5%) markers exhibited monomorphic amplification among chickpea genotypes used. A total number of 764 alleles were amplified by 160 polymorphic SSR markers with a mean allele number of 4.8. The number of alleles amplified per locus varied from 2 to 12. The PIC ranged from 0.23 to 0.86 with an average of 0.75, while gene diversity varied from 0.25 to 0.89 with a mean of 0.77 (Supplementary Table S2). The polymorphic potential of markers in different sequence components of genes and intergenic regions was analyzed in detail based on the percent polymorphism, PIC and polymorphic alleles amplified among chickpea genotypes. We were able to detect polymorphism in 55 (62.5%, allele number from 2–9 and PIC 0.74) of 88 markers derived from the different coding and non-CDS components of genes between desi and kabuli chickpea. The remaining 105 (64%, 2–12 and 0.63) of 164 markers derived from intergenic regions also showed polymorphism between the two chickpea types (Supplementary Table S2). Within genes, maximum potential of polymorphism was detected by the markers developed from intronic sequences (28 of 38 markers, 73.7%, allele number 2–9 and PIC 0.74) followed by 3′-UTRs (5 of 8 markers, 62.5%, 2–4 and 0.66), upstream (19 of 35 markers, 54.3%, 2–5 and 0.76) and 5′-UTRs (2 of 6 markers, 33.3%, 4–5 and 0.62) of genes (Supplementary Table S2, Figure 5A). Remarkably, 120 (47.6%, 2–10 and 0.62) and 107 (42.5%, 2–8 and 0.57) markers revealed polymorphism within desi and kabuli chickpea genotypes too, respectively. Overall, eight different representative allele types were detected based on fragment length polymorphism patterns of all the 160 SSR markers in 46 desi and kabuli genotypes (Figure 5B).

Bottom Line: These physically mapped SSR markers exhibited robust amplification efficiency (73.9%) and high intra- and inter-specific polymorphic potential (63.5%), thereby suggesting their immense use in various genomics-assisted breeding applications.The SSR markers particularly derived from intergenic and intronic sequences revealed high polymorphic potential.The intra-specific polymorphism (47.6%) and functional molecular diversity (65%) potential of polymorphic SSR markers developed in our study is much higher than that of previous documentations.

View Article: PubMed Central - PubMed

Affiliation: Functional and Applied Genomics Laboratory, National Institute of Plant Genome Research New Delhi, India.

ABSTRACT
Development of informative polymorphic simple sequence repeat (SSR) markers at a genome-wide scale is essential for efficient large-scale genotyping applications. We identified genome-wide 1835 SSRs showing polymorphism between desi and kabuli chickpea. A total of 1470 polymorphic SSR markers from diverse coding and non-coding regions of the chickpea genome were developed. These physically mapped SSR markers exhibited robust amplification efficiency (73.9%) and high intra- and inter-specific polymorphic potential (63.5%), thereby suggesting their immense use in various genomics-assisted breeding applications. The SSR markers particularly derived from intergenic and intronic sequences revealed high polymorphic potential. Using the mapped SSR markers, a wider functional molecular diversity (16-94%, mean: 68%), and parentage- and cultivar-specific admixed domestication pattern and phylogenetic relationships in a structured population of desi and kabuli chickpea genotypes was evident. The intra-specific polymorphism (47.6%) and functional molecular diversity (65%) potential of polymorphic SSR markers developed in our study is much higher than that of previous documentations. Finally, we have developed a user-friendly web resource, Chickpea Microsatellite Database (CMsDB; http://www.nipgr.res.in/CMsDB.html), which provides public access to the data and results reported in this study. The developed informative SSR markers can serve as a resource for various genotyping applications, including genetic enhancement studies in chickpea.

No MeSH data available.