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High-throughput SNP discovery and genotyping for constructing a saturated linkage map of chickpea (Cicer arietinum L.).

Gaur R, Azam S, Jeena G, Khan AW, Choudhary S, Jain M, Yadav G, Tyagi AK, Chattopadhyay D, Bhatia S - DNA Res. (2012)

Bottom Line: Of these, 697 SNPs could be successfully used for genotyping, demonstrating a high success rate of 90.75%.The map was used for the synteny analysis of chickpea, which revealed a higher degree of synteny with the phylogenetically close Medicago than with soybean.The first set of validated SNPs and map resources developed in this study will not only facilitate QTL mapping, genome-wide association analysis and comparative mapping in legumes but also help anchor scaffolds arising out of the whole-genome sequencing of chickpea.

View Article: PubMed Central - PubMed

Affiliation: National Institute of Plant Genome Research, Aruna Asaf Ali Marg, PO Box 10531, New Delhi 110067, India.

ABSTRACT
The present study reports the large-scale discovery of genome-wide single-nucleotide polymorphisms (SNPs) in chickpea, identified mainly through the next generation sequencing of two genotypes, i.e. Cicer arietinum ICC4958 and its wild progenitor C. reticulatum PI489777, parents of an inter-specific reference mapping population of chickpea. Development and validation of a high-throughput SNP genotyping assay based on Illumina's GoldenGate Genotyping Technology and its application in building a high-resolution genetic linkage map of chickpea is described for the first time. In this study, 1022 SNPs were identified, of which 768 high-confidence SNPs were selected for designing the custom Oligo Pool All (CpOPA-I) for genotyping. Of these, 697 SNPs could be successfully used for genotyping, demonstrating a high success rate of 90.75%. Genotyping data of the 697 SNPs were compiled along with those of 368 co-dominant markers mapped in an earlier study, and a saturated genetic linkage map of chickpea was constructed. One thousand and sixty-three markers were mapped onto eight linkage groups spanning 1808.7 cM (centiMorgans) with an average inter-marker distance of 1.70 cM, thereby representing one of the most advanced maps of chickpea. The map was used for the synteny analysis of chickpea, which revealed a higher degree of synteny with the phylogenetically close Medicago than with soybean. The first set of validated SNPs and map resources developed in this study will not only facilitate QTL mapping, genome-wide association analysis and comparative mapping in legumes but also help anchor scaffolds arising out of the whole-genome sequencing of chickpea.

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Inter-specific linkage map of chickpea. The inter-specific linkage map of chickpea based on RILs of C. arietinum (ICC4958) × C. reticulatum (PI489777) was generated using JoinMap version 4.0. The name of the linkage groups is mentioned at the top of each LG. Distances of the loci (cM) are shown to the left and the names of loci are shown to the right side of the linkage groups. SNP markers are represented in black, while previously published markers are shown in colour: red, genic markers; green, genomic markers; blue, STMS markers of Winter et al.58 used for anchoring.
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DSS018F2: Inter-specific linkage map of chickpea. The inter-specific linkage map of chickpea based on RILs of C. arietinum (ICC4958) × C. reticulatum (PI489777) was generated using JoinMap version 4.0. The name of the linkage groups is mentioned at the top of each LG. Distances of the loci (cM) are shown to the left and the names of loci are shown to the right side of the linkage groups. SNP markers are represented in black, while previously published markers are shown in colour: red, genic markers; green, genomic markers; blue, STMS markers of Winter et al.58 used for anchoring.

Mentions: For construction of a dense genetic map of chickpea, the genotyping data of 697 polymorphic SNPs obtained from 129 RILs of the reference inter-specific mapping population (C. arietinum ICC4958 x C. reticulatum PI489777) were utilized. Further, to generate an integrated and advanced genetic map of chickpea, the genotyping data of the 697 SNPs were compiled along with the genotyping data of 368 co-dominant markers published previously by our group.48 These markers included 238 gSSRs (genomic SSRs), 52 EST-SSRs, 51 ITPs (Intron Targeted Polymorphism), 25 ESTPs and 2 MtESTs (Table 1), which served as a framework for anchoring the SNP loci. Hence, data of 1065 markers (697 SNPs and 368 previously mapped co-dominant markers) were compiled together for linkage analysis and map generation using JoinMap ver. 4.0.67 The resulting integrated linkage map of chickpea defined map positions of 1063 markers distributed over 8 linkage groups (Fig. 2). The current map spanned 1808.7 cM with an average inter-marker distance of 1.7 cM. The map contained an average of 1.43 map positions per Mb of genome (1063 map positions/740 Mb) and had one marker per 696 kb (740 Mb/1063) when considering the chickpea genome to be 740 Mb.44 The LGs were numbered (1 to 8) based on the common marker positions shared between corresponding LGs from previous studies.48,58 The genetic length of the LGs ranged from 186.8 cM (LG2) to 269.5 cM (LG1) (Table 2). LG3 was the most saturated, having 205 markers with an average marker density of 0.95 cM, whereas LG8 had the least number of markers (only 84) (Table 2). On an average, one linkage group contained 132.9 markers that spanned an average of 226 cM. All categories of markers including genic, genomic and SNPs were found on each of the LGs. However, the number of SNP markers in each of the LGs was maximum and varied in the range 65–75% except in LG7, which had only 38% SNP markers. The markers were unevenly and non-randomly distributed in the LGs and 25 clusters of markers (≥5 markers within 1 cM distance) were identified of which 20 clusters comprise only SNP markers, whereas a single cluster on LG3 contained only STMS markers. Moreover, nine gaps ranging from 20 to 35 cM were observed in four LGs (LG1, LG5, LG7 and LG8) located near the distal ends. The χ2 test performed resulted in segregation distortion for 42% (448) of the marker loci. However, 446 of these markers were finally integrated into the map so that the loss of genetic information related to these markers was minimized. Moreover, the distorted markers were found to be widely distributed throughout all the LGs even though the ratios varied from one LG to another. For example in LG1 and LG4, <25% markers showed distortion, whereas in others >30% of the mapped loci were distorted. The overall segregation distortion of 42% observed in this study was similar to the 41.3 and 38% reported earlier for the same mapping population.48,58Table 2.


High-throughput SNP discovery and genotyping for constructing a saturated linkage map of chickpea (Cicer arietinum L.).

Gaur R, Azam S, Jeena G, Khan AW, Choudhary S, Jain M, Yadav G, Tyagi AK, Chattopadhyay D, Bhatia S - DNA Res. (2012)

Inter-specific linkage map of chickpea. The inter-specific linkage map of chickpea based on RILs of C. arietinum (ICC4958) × C. reticulatum (PI489777) was generated using JoinMap version 4.0. The name of the linkage groups is mentioned at the top of each LG. Distances of the loci (cM) are shown to the left and the names of loci are shown to the right side of the linkage groups. SNP markers are represented in black, while previously published markers are shown in colour: red, genic markers; green, genomic markers; blue, STMS markers of Winter et al.58 used for anchoring.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3473369&req=5

DSS018F2: Inter-specific linkage map of chickpea. The inter-specific linkage map of chickpea based on RILs of C. arietinum (ICC4958) × C. reticulatum (PI489777) was generated using JoinMap version 4.0. The name of the linkage groups is mentioned at the top of each LG. Distances of the loci (cM) are shown to the left and the names of loci are shown to the right side of the linkage groups. SNP markers are represented in black, while previously published markers are shown in colour: red, genic markers; green, genomic markers; blue, STMS markers of Winter et al.58 used for anchoring.
Mentions: For construction of a dense genetic map of chickpea, the genotyping data of 697 polymorphic SNPs obtained from 129 RILs of the reference inter-specific mapping population (C. arietinum ICC4958 x C. reticulatum PI489777) were utilized. Further, to generate an integrated and advanced genetic map of chickpea, the genotyping data of the 697 SNPs were compiled along with the genotyping data of 368 co-dominant markers published previously by our group.48 These markers included 238 gSSRs (genomic SSRs), 52 EST-SSRs, 51 ITPs (Intron Targeted Polymorphism), 25 ESTPs and 2 MtESTs (Table 1), which served as a framework for anchoring the SNP loci. Hence, data of 1065 markers (697 SNPs and 368 previously mapped co-dominant markers) were compiled together for linkage analysis and map generation using JoinMap ver. 4.0.67 The resulting integrated linkage map of chickpea defined map positions of 1063 markers distributed over 8 linkage groups (Fig. 2). The current map spanned 1808.7 cM with an average inter-marker distance of 1.7 cM. The map contained an average of 1.43 map positions per Mb of genome (1063 map positions/740 Mb) and had one marker per 696 kb (740 Mb/1063) when considering the chickpea genome to be 740 Mb.44 The LGs were numbered (1 to 8) based on the common marker positions shared between corresponding LGs from previous studies.48,58 The genetic length of the LGs ranged from 186.8 cM (LG2) to 269.5 cM (LG1) (Table 2). LG3 was the most saturated, having 205 markers with an average marker density of 0.95 cM, whereas LG8 had the least number of markers (only 84) (Table 2). On an average, one linkage group contained 132.9 markers that spanned an average of 226 cM. All categories of markers including genic, genomic and SNPs were found on each of the LGs. However, the number of SNP markers in each of the LGs was maximum and varied in the range 65–75% except in LG7, which had only 38% SNP markers. The markers were unevenly and non-randomly distributed in the LGs and 25 clusters of markers (≥5 markers within 1 cM distance) were identified of which 20 clusters comprise only SNP markers, whereas a single cluster on LG3 contained only STMS markers. Moreover, nine gaps ranging from 20 to 35 cM were observed in four LGs (LG1, LG5, LG7 and LG8) located near the distal ends. The χ2 test performed resulted in segregation distortion for 42% (448) of the marker loci. However, 446 of these markers were finally integrated into the map so that the loss of genetic information related to these markers was minimized. Moreover, the distorted markers were found to be widely distributed throughout all the LGs even though the ratios varied from one LG to another. For example in LG1 and LG4, <25% markers showed distortion, whereas in others >30% of the mapped loci were distorted. The overall segregation distortion of 42% observed in this study was similar to the 41.3 and 38% reported earlier for the same mapping population.48,58Table 2.

Bottom Line: Of these, 697 SNPs could be successfully used for genotyping, demonstrating a high success rate of 90.75%.The map was used for the synteny analysis of chickpea, which revealed a higher degree of synteny with the phylogenetically close Medicago than with soybean.The first set of validated SNPs and map resources developed in this study will not only facilitate QTL mapping, genome-wide association analysis and comparative mapping in legumes but also help anchor scaffolds arising out of the whole-genome sequencing of chickpea.

View Article: PubMed Central - PubMed

Affiliation: National Institute of Plant Genome Research, Aruna Asaf Ali Marg, PO Box 10531, New Delhi 110067, India.

ABSTRACT
The present study reports the large-scale discovery of genome-wide single-nucleotide polymorphisms (SNPs) in chickpea, identified mainly through the next generation sequencing of two genotypes, i.e. Cicer arietinum ICC4958 and its wild progenitor C. reticulatum PI489777, parents of an inter-specific reference mapping population of chickpea. Development and validation of a high-throughput SNP genotyping assay based on Illumina's GoldenGate Genotyping Technology and its application in building a high-resolution genetic linkage map of chickpea is described for the first time. In this study, 1022 SNPs were identified, of which 768 high-confidence SNPs were selected for designing the custom Oligo Pool All (CpOPA-I) for genotyping. Of these, 697 SNPs could be successfully used for genotyping, demonstrating a high success rate of 90.75%. Genotyping data of the 697 SNPs were compiled along with those of 368 co-dominant markers mapped in an earlier study, and a saturated genetic linkage map of chickpea was constructed. One thousand and sixty-three markers were mapped onto eight linkage groups spanning 1808.7 cM (centiMorgans) with an average inter-marker distance of 1.70 cM, thereby representing one of the most advanced maps of chickpea. The map was used for the synteny analysis of chickpea, which revealed a higher degree of synteny with the phylogenetically close Medicago than with soybean. The first set of validated SNPs and map resources developed in this study will not only facilitate QTL mapping, genome-wide association analysis and comparative mapping in legumes but also help anchor scaffolds arising out of the whole-genome sequencing of chickpea.

Show MeSH
Related in: MedlinePlus