Combination of newly developed SNP and InDel markers for genotyping the <Emphasis Type="Italic">Cf-9</Emphasis> locus conferring disease resistance to leaf mold disease in the tomato

The Cf-9 gene in the tomato is known to confer resistance against leaf mold disease caused by Cladosporium fulvum, and a gene-based marker targeted to the Cf-9 allele has been widely used as a crop protection approach. However, we found this marker to be misleading in genotyping. Therefore, we developed new single-nucleotide polymorphism (SNP) and insertion and deletion (InDel) markers targeted to the Cf-9 allele in order to increase genotyping accuracy and facilitate high-throughput screening. The DNA sequences of reported Cf-9, cf-9, Cf-0, and closely related Cf-4 alleles were compared, and two functional and non-synonymous SNPs were found to distinguish the Cf-9 resistance allele from the cf-9, Cf-0, and Cf-4 alleles. An SNP marker including these two SNPs was developed and applied to the genotyping of 33 tomato cultivars by high-resolution melting analysis. Our SNP marker was able to select all three Cf-9 genotypes (resistant, heterozygous, and susceptible alleles). Interestingly, two cultivars were grouped separately from these three genotypes. To further examine this outgroup, we preformed polymerase chain reaction (PCR) on two InDel regions identified by sequence comparison of the Cf-9 and Cf-4 genes. The band patterns revealed that these two cultivars carried Cf-4 rather than Cf-9 alleles and that three cultivars classified in the Cf-9 resistance group actually carried both Cf-9 and Cf-4 genes. To determine whether these genotyping results were consistent with disease resistance phenotypes, we examined the induction of a hypersensitive response by transiently expressing the corresponding effector genes, and found that the results matched perfectly with the genotyping results. These findings indicate that the combination of our SNP and InDel markers allows resistant Cf-9 alleles to be distinguished from cf-9 and Cf-4 alleles, which will be useful for marker-assisted selection of tomato cultivars resistant to C. fulvum.     

Identification of a molecular marker tightly linked to bacterial wilt resistance in tomato by genome-wide SNP analysis

Key message

Genotyping of disease resistance to bacterial wilt in tomato by a genome-wide SNP analysis

Abstract

Bacterial wilt caused by Ralstonia pseudosolanacearum is one of the destructive diseases in tomato. The previous studies have identified Bwr-6 (chromosome 6) and Bwr-12 (chromosome 12) loci as the major quantitative trait loci (QTLs) contributing to resistance against bacterial wilt in tomato cultivar ‘Hawaii7996’. However, the genetic identities of two QTLs have not been uncovered yet. In this study, using whole-genome resequencing, we analyzed genome-wide single-nucleotide polymorphisms (SNPs) that can distinguish a resistant group, including seven tomato varieties resistant to bacterial wilt, from a susceptible group, including two susceptible to the same disease. In total, 5259 non-synonymous SNPs were found between the two groups. Among them, only 265 SNPs were located in the coding DNA sequences, and the majority of these SNPs were located on chromosomes 6 and 12. The genes that both carry SNP(s) and are near Bwr-6 and Bwr-12 were selected. In particular, four genes in chromosome 12 encode putative leucine-rich repeat (LRR) receptor-like proteins. SNPs within these four genes were used to develop SNP markers, and each SNP marker was validated by a high-resolution melting method. Consequently, one SNP marker, including a functional SNP in a gene, Solyc12g009690.1, could efficiently distinguish tomato varieties resistant to bacterial wilt from susceptible varieties. These results indicate that Solyc12g009690.1, the gene encoding a putative LRR receptor-like protein, might be tightly linked to Bwr-12, and the SNP marker developed in this study will be useful for selection of tomato cultivars resistant to bacterial wilt.

     

Graphical genotyping as a method to map <Emphasis Type="Italic">Ny</Emphasis><Subscript><Emphasis Type="Italic">(o,n)sto</Emphasis></Subscript> and <Emphasis Type="Italic">Gpa5</Emphasis> using a reference panel of tetraploid potato cultivars

Key message

The method of graphical genotyping is applied to a panel of tetraploid potato cultivars to visualize haplotype sharing. The method allowed to map genes involved in virus and nematode resistance. The physical coordinates of the amount of linkage drag surrounding these genes are easily interpretable.

Abstract

Graphical genotyping is a visually attractive and easily interpretable method to represent genetic marker data. In this paper, the method is extended from diploids to a panel of tetraploid potato cultivars. Application of filters to select a subset of SNPs allows one to visualize haplotype sharing between individuals that also share a specific locus. The method is illustrated with cultivars resistant to Potato virus Y (PVY), while simultaneously selecting for the absence of the SNPs in susceptible clones. SNP data will then merge into an image which displays the coordinates of a distal genomic region on the northern arm of chromosome 11 where a specific haplotype is introgressed from the wild potato species S. stoloniferum (CPC 2093) carrying a gene (Ny _(o,n)sto ) conferring resistance to two PVY strains, PVY^O and PVY^NTN. Graphical genotyping was also successful in showing the haplotypes on chromosome 12 carrying Ry-f _sto , another resistance gene derived from S. stoloniferum conferring broad-spectrum resistance to PVY, as well as chromosome 5 haplotypes from S. vernei, with the Gpa5 locus involved in resistance against Globodera pallida cyst nematodes. The image also shows shortening of linkage drag by meiotic recombination of the introgression segment in more recent breeding material. Identity-by-descent was found to be a requirement for using graphical genotyping, which is proposed as a non-statistical alternative method for gene discovery, as compared with genome-wide association studies. The potential and limitations of the method are discussed.

     

Developing Single Nucleotide Polymorphism (SNP) Markers for the Identification of Coffee Germplasm

Coffee is one of the most widely consumed beverages and represents a multibillion-dollar global industry. Accurate identification of coffee cultivars is essential for efficient management, exchange, and use of coffee genetic resources. To date, a universal platform that can allow data comparison across different laboratories and genotyping platforms has not been developed by the coffee research community. Using expressed sequence tags (EST) of Coffea arabica, C. canephora and C. racemosa from public databases, we developed 7538 single nucleotide polymorphism (SNP) markers and selected 180 for validation using 25 C. arabica and C. canephora accessions from Puerto Rico. Based on the validation result, we designated a panel of 55 SNP markers that are polymorphic across the two species. The average minor allele frequency and information index of this SNP panel are 0.281 and 0.690, respectively. This panel enabled the differentiation of all tested accessions of C. canephora, which accounts for 79.2 % of the total polymorphism in the samples. Only 21.8 % of the polymorphic SNPs were detected in the 12 C. arabica cultivars, which, nonetheless, were able to unambiguously differentiate the 12 Arabica cultivars into ten unique genotypes, including two synonymous groups. Several local Puerto Rican cultivars with partial Timor pedigree, including Limaní, Frontón, and TARS 18087, showed substantial genetic difference from the other common Arabica cultivars, such as Catuai, Borbón, and Mundo Nuevo. This coffee SNP panel provides robust and universally comparable DNA fingerprints, thus can serve as a genotyping tool to assist coffee germplasm management, propagation of planting material, and coffee cultivar authentication.     

<Emphasis Type="Italic">FaRCg1</Emphasis>: a quantitative trait locus conferring resistance to Colletotrichum crown rot caused by <Emphasis Type="Italic">Colletotrichum gloeosporioides</Emphasis> in octoploid strawberry

Colletotrichum crown rot (CCR) is an important disease of strawberry (Fragaria?×ananassa) throughout the Southeastern US and in subtropical climates around the world, where hot and humid conditions facilitate rapid disease development. Yet no resistance loci have been described to date, as genetic studies have been historically difficult in allo-octoploid (2n?=?8x?=?56) strawberry. In the present study, we investigate the genetic architecture of resistance to CCR. Four population sets from the University of Florida were inoculated in four different seasons from 2013–2014 to 2016–2017. Two large, multiparental discovery population sets were used for QTL discovery, and two validation sets of cultivars and advanced selections representing the parent pool of the breeding program were also assessed. Subgenome-specific single-nucleotide polymorphism (SNP) markers were mapped, and FlexQTL? software was utilized to perform a Bayesian, pedigree-based QTL analysis. A quantitative trait locus on linkage group 6B, which we name FaRCg1, accounts for most of the genetic variation for resistance in the discovery sets (26.8–29.8% in 2013–2014 and 17% in 2015–2016). High-throughput marker assays were developed for the most significant SNPs which correlated with the mode of the QTL region. The discovery and characterization of the FaRCg1 locus and the molecular tools developed from it will be utilized to achieve increased genetic gains for resistance.     

Molecular markers for adult plant leaf rust resistance gene <Emphasis Type="Italic">Lr48</Emphasis> in wheat

Leaf rust of wheat, caused by Puccinia triticina, is an important disease throughout the world. The adult plant leaf rust resistance gene Lr48 reported in CSP44 was previously mapped in chromosome 2B, but the marker–gene association was weak. In this study, we confirmed the location of Lr48 to be in the short arm of chromosome 2B and identified closely linked markers suitable for use in breeding. The CSP44/WL711 recombinant inbred line (RIL) population (90 lines) showed monogenic segregation for Lr48. Twelve resistant and 12 susceptible RILs were used for selective genotyping using an iSelect 90K Infinium SNP assay. Closely linked SNPs were converted into Kompetitive allele-specific primers (KASP) and tested on the parental lines. KASP markers giving clear clusters for alternate genotypes were assayed on the entire RIL population. SNP markers IWB31002, IWB39832, IWB34324, IWB72894 and IWB36920 co-segregated with Lr48 and the marker IWB70147 was mapped 0.3 cM proximal to this gene. Closely linked KASP markers were tested on a set of Australian and Nordic wheat genotypes. The amplification of SNP alleles alternate to those linked with Lr48 in the majority of the Australian and Nordic wheat genotypes demonstrated the usefulness of these markers for marker-assisted pyramiding of Lr48 with other rust resistance genes.     

Adult plant stripe rust resistance gene <Emphasis Type="Italic">Yr71</Emphasis> maps close to <Emphasis Type="Italic">Lr24</Emphasis> in chromosome 3D of common wheat

Australian cultivar Sunco carries three adult plant stripe rust resistance genes. One of these genes corresponded to Yr18 in chromosome 7DS; the second, YrCK, was mapped on chromosome 2D. Here, we describe the characterization of the third adult plant resistance (APR) gene from Sunco. Sunco/2*Avocet S-derived lines SA65 (resistant) and SA67 (susceptible) were crossed and a recombinant inbred line F₆ population was generated. Monogenic segregation among SA65/SA67-derived RIL population was demonstrated and the resistance locus was designated YrSA3. Selective genotyping using an iSelect 90 K Infinium SNP array and SSR markers located YrSA3 on chromosome 3D. Development of KASP markers for SNP loci showing association with YrSA3 allowed construction of a genetic map harboring the resistance gene. Ten KASP markers (KASP_8306, KASP_9142, KASP_10438, KASP_16434, KASP_17207, KASP_20836, KASP_23518, KASP_23615, KASP_57983 and KASP_63653), one SSR marker (gwm114b) and Lr24/Sr24 were mapped 1.8 cM distal to YrSA3. Comparison of marker data indicated that the previously named seedling stripe rust resistance gene Yr45 was located proximal to YrSA3, and therefore the latter was formally designated Yr71. Two recombinants carrying Lr24/Sr24 and Yr71 in combination were identified for use as donor sources in wheat breeding programs. The robustness of gwm114b, KASP_16434, KASP_17207 and KASP_20836 for marker-assisted selection of these genes was demonstrated through tests on 74 Australian wheat cultivars.     

<Emphasis Type="Bold">A user guide to the</Emphasis><Emphasis Type="BoldItalic">Brassica</Emphasis><Emphasis Type="Bold">60K Illumina Infinium</Emphasis>™ <Emphasis Type="Bold">SNP genotyping array</Emphasis>

The Brassica napus 60K Illumina Infinium? SNP array has had huge international uptake in the rapeseed community due to the revolutionary speed of acquisition and ease of analysis of this high-throughput genotyping data, particularly when coupled with the newly available reference genome sequence. However, further utilization of this valuable resource can be optimized by better understanding the promises and pitfalls of SNP arrays. We outline how best to analyze Brassica SNP marker array data for diverse applications, including linkage and association mapping, genetic diversity and genomic introgression studies. We present data on which SNPs are locus-specific in winter, semi-winter and spring B. napus germplasm pools, rather than amplifying both an A-genome and a C-genome locus or multiple loci. Common issues that arise when analyzing array data will be discussed, particularly those unique to SNP markers and how to deal with these for practical applications in Brassica breeding applications.     

Development of species diagnostic SNP markers for quality control genotyping in four rice (<Emphasis Type="Italic">Oryza</Emphasis> L.) species

Species misclassification (misidentification) and handling errors have been frequently reported in various plant species conserved at diverse gene banks, which could restrict use of germplasm for correct purpose. The objectives of the present study were to (i) determine the extent of genotyping error (reproducibility) on DArTseq-based single-nucleotide polymorphisms (SNPs); (ii) determine the proportion of misclassified accessions across 3134 samples representing three African rice species complex (Oryza glaberrima, O. barthii, and O. longistaminata) and an Asian rice (O. sativa), which are conserved at the AfricaRice gene bank; and (iii) develop species- and sub-species (ecotype)-specific diagnostic SNP markers for rapid and low-cost quality control (QC) analysis. Genotyping error estimated from 15 accessions, each replicated from 2 to 16 times, varied from 0.2 to 3.1%, with an overall average of 0.8%. Using a total of 3134 accessions genotyped with 31,739 SNPs, the proportion of misclassified samples was 3.1% (97 of the 3134 accessions). Excluding the 97 misclassified accessions, we identified a total of 332 diagnostic SNPs that clearly discriminated the three indigenous African species complex from Asian rice (156 SNPs), O. longistaminata accessions from both O. barthii and O. glaberrima (131 SNPs), and O. sativa spp. indica from O. sativa spp. japonica (45 SNPs). Using chromosomal position, minor allele frequency, and polymorphic information content as selection criteria, we recommended a subset of 24 to 36 of the 332 diagnostic SNPs for routine QC genotyping, which would be highly useful in determining the genetic identity of each species and correct human errors during routine gene bank operations.     

Development of user-friendly markers for disease resistance to black root rot of tobacco through genotyping by sequencing

Black root rot (BRR), a disease caused by the hemibiotrophic fungus Thielaviopsis basicola, seriously compromises yield and leaf quality in tobacco (Nicotiana tabacum). Full resistance to black root rot, conferred by the resistance to BRR 1 (RBRR1) locus from Nicotiana debneyi Domin, was transferred to a burley tobacco cultivar through interspecific hybridization. Some undesirable traits potentially caused by linkage drag restrict wider application of RBRR1 in flue-cured tobacco. Therefore, user-friendly molecular markers are needed to assist selection for resistance to black root rot and to break the unfavorable linkage. Genotyping by sequencing (GBS) is a rapid and robust approach for reduced representation sequencing of multiplexed genomic DNA samples that combines genome-wide molecular marker discovery with genotyping. In the present study, we used GBS to identify single-nucleotide polymorphisms (SNPs) linked to the RBRR1 locus, and PCR-based assays for detection of these SNPs were also developed. Sequence analysis of the SNP markers suggested that RBRR1 is located on chromosome 17, providing a basis for map-based cloning of this valuable gene. Co-dominant CAPS markers that co-segregate with the disease-resistant phenotype offer user-friendly tools for tobacco breeding and variety improvement. Furthermore, tested with diverse N. tabacum germplasm, SS192650 displayed 100% selection accuracy for resistance to BRR, suggesting that this marker can be used in diverse tobacco populations.     

An integrative AmpSeq platform for highly multiplexed marker-assisted pyramiding of grapevine powdery mildew resistance loci

Resistance breeding often requires the introgression and tracking of resistance loci from wild species into domesticated backgrounds, typically with the goal of pyramiding multiple resistance genes, to provide durable disease resistance to breeding selections and ultimately cultivars. While molecular markers are commonly used to facilitate these efforts, high genetic diversity and divergent marker technologies can complicate marker-assisted breeding strategies. Here, amplicon sequencing (AmpSeq) was used to integrate SNP markers with dominant presence/absence markers derived from genotyping-by-sequencing and other genotyping technologies, for the simultaneous tracking of five loci for resistance to grapevine powdery mildew. SNP haploblocks defined the loci for REN1, REN2 and REN3, which confer quantitative resistance phenotypes that are challenging to measure via field ratings of natural infections. Presence/absence markers for RUN1 and REN4 were validated to predict qualitative resistance phenotypes and corresponded with previous presence/absence fluorescent electrophoretic assays. Thus, 37 AmpSeq-derived markers were identified for the five loci, and markers for REN1, REN2, REN4 and RUN1 were used for multiplexed screening and selection within diverse breeding germplasm. Poor transferability of SNP markers indicated imperfect marker-trait association in some families. Together, AmpSeq SNP haploblocks and presence/absence markers provide a high-throughput, cost-effective tool to integrate divergent technologies for marker-assisted selection and genetic analysis of introgressed disease resistance loci in grapevine.     

Identification of loci controlling forage yield and nutritive value in diploid alfalfa using GBS-GWAS

Key message

We attempted to identify genomic regions controlling forage yield and nutritive value in alfalfa. Several candidate genes and associated genetic markers were identified that could potentially be useful for alfalfa breeding to more efficiently develop improved cultivars.

Abstract

Alfalfa is one of the most widely cultivated forage legumes worldwide and improving alfalfa forage yield and nutritive value is a major global breeding goal. Genotyping-by-sequencing (GBS) provides cost-effective molecular marker genotyping for genome-wide association studies (GWAS). Using more than 15,000 genome-wide single nucleotide polymorphisms (SNP) identified from GBS, we conducted a GWAS to investigate forage yield and nutritive value-related traits. We have detected a number of associations for all the traits evaluated and a number of associations detected were located on the Medicago truncatula genome. The SNP in a coding region of a cell wall biosynthesis gene was associated with several cell wall-related traits, and we suggest that it may be the causative polymorphism. Two other SNPs residing in meristematic development and early growth genes were found to associate with the total biomass yield. None of the SNPs associated with regrowth after harvest or with spring regrowth were mapped to the M. truncatula genome, possibly reflecting the fact that M. truncatula is an annual species related to alfalfa that typically has limited ability to regrow. The alleles we identify with the major impact on forage yield and nutritive value can be rapidly incorporated into our breeding program.

     

Molecular mapping of powdery mildew resistance gene <Emphasis Type="Italic">PmSGD</Emphasis> in Chinese wheat landrace Shangeda using RNA-seq with bulk segregant analysis

Wheat powdery mildew, caused by the fungal pathogen Blumeria graminis f. sp. tritici (Bgt), is one of the most devastating diseases of wheat in China and causes serious yield losses. Resistance genes are urgently needed by wheat breeding programs to combat this disease. In the present study, genetic analysis of powdery mildew resistance was conducted on segregated F₂ and F_2:3 populations derived from the cross of Shangeda (providing good resistance to powdery mildew) and Chancellor (susceptible to powdery mildew). The results showed that the resistance of Shangeda to E09 was controlled by a single recessive gene, tentatively designated as PmSGD. In addition, RNA sequencing of the parental lines Shangeda and Chancellor and the corresponding bulked pools derived from homozygous resistant or susceptible F_2:3 lines was implemented to identify single-nucleotide polymorphisms (SNPs). The PmSGD gene was estimated to be located in the 240–250-Mb region of chromosome 7B based on the characteristics of putative SNP loci distributed on 21 wheat chromosomes. Among the developed SNP markers, 17 (57%) markers were linked to PmSGD flanked by SNP2-57 and SNP2-46, with genetic distances of 0.4 and 0.8 cM, respectively. The reaction patterns of Shangeda and cultivars (lines) carrying the Pm5e, Pmhym, mlxbd, and PmTm4 genes to 22 Bgt isolates indicated that PmSGD may be allelic or very closely linked to those genes. All of the SNP loci linked to PmSGD were used to test 38 cultivars with known Pm gene(s), and the results suggested that these SNP loci are useful for pyramiding PmSGD by marker-assisted selection.     

Genome-wide association study on chicken carcass traits using sequence data imputed from SNP array

Chicken carcass traits are economically important for the chicken industry. Detecting which genes affect chicken carcass traits is of great benefit to the genetic improvement of this important agricultural species. To investigate the genetic mechanism of carcass traits in chickens, we carried out a genome-wide association study (GWAS). A total of 435 Chinese indigenous chickens were phenotyped for carcass weight (CW), eviscerated weight with giblets (EWG), and eviscerated weight (EW) after slaughter at 91 days and were genotyped using a 600-K single nucleotide polymorphism (SNP) genotyping array. Twenty-four birds were selected for sequencing, and the 600 K SNP panel data were imputed to sequence data with the 24 birds as the reference. Univariate GWASs were performed with GEMMA software using the whole genome sequence data imputed from SNP chip data. Finally, 3, 25, and 63 suggestively significant SNPs were identified to be associated with carcass weight (CW), eviscerated weight with giblets (EWG), and eviscerated weight (EW), respectively. Six candidate genes, RNF219, SCEL, MYCBP2, ETS1, APLP2, and PRDM10 were detected. SCEL and MYCBP2 were potentially associated with these three traits, RNF219 and APLP2 were potentially associated with EWG and EW, and ETS1 and PRDM10 were only potentially associated with EWG and EW, respectively. Compared with forefathers’ research, 10 reported QTLs associated with CW were located within a 5-Mb distance near the SNPs with P value lower than 1×10^?5. This study enriched the knowledge of the genetic mechanisms of chicken carcass traits.     

High-throughput SNP discovery and genotyping in durum wheat (<Emphasis Type="Italic">Triticum durum</Emphasis> Desf.)

We describe the application of complexity reduction of polymorphic sequences (CRoPS^®) technology for the discovery of SNP markers in tetraploid durum wheat (Triticum durum Desf.). A next-generation sequencing experiment was carried out on reduced representation libraries obtained from four durum cultivars. SNP validation and minor allele frequency (MAF) estimate were carried out on a panel of 12 cultivars, and the feasibility of genotyping these SNPs in segregating populations was tested using the Illumina Golden Gate (GG) technology. A total of 2,659 SNPs were identified on 1,206 consensus sequences. Among the 768 SNPs that were chosen irrespective of their genomic repetitiveness level and assayed on the Illumina BeadExpress genotyping system, 275 (35.8%) SNPs matched the expected genotypes observed in the SNP discovery phase. MAF data indicated that the overall SNP informativeness was high: a total of 196 (71.3%) SNPs had MAF >0.2, of which 76 (27.6%) showed MAF >0.4. Of these SNPs, 157 were mapped in one of two mapping populations (Meridiano × Claudio and Colosseo × Lloyd) and integrated into a common genetic map. Despite the relatively low genotyping efficiency of the GG assay, the validated CRoPS-derived SNPs showed valuable features for genomics and breeding applications such as a uniform distribution across the wheat genome, a prevailing single-locus codominant nature and a high polymorphism. Here, we report a new set of 275 highly robust genome-wide Triticum SNPs that are readily available for breeding purposes.     

High resolution DNA melting markers for identification of <Emphasis Type="Italic">H1</Emphasis>-linked resistance to potato cyst nematode

Although Sequence-Characterized Amplified Region (SCAR) markers linked to the potato H1 locus, which confers resistance to pathotypes Ro1 and Ro4 of the potato cyst nematode (PCN) Globodera rostochiensis, have been reported, robust markers that enable estimation of allele dosage would improve the quality of information obtained from genotyping parental accessions (cultivars/breeding lines) and progeny populations within breeding programmes. With this in mind, we have developed single nucleotide polymorphism (SNP)-based molecular markers flanking the H1 resistance gene, using genomic re-sequence data from five elite tetraploid accessions. The published TG689 and 57R primer sequences were used in a Basic Local Alignment Search Tool (BLAST) examination of the reference potato genome, and SNPs within the vicinity of these primer regions were identified and targeted for designing probe-based High Resolution Melting (HRM) SNP assays. Evaluation of the subsequently developed HRM markers, TG689_1P and 57R_1P, against the publicly available SCAR markers, TG689 and 57R, indicated that the HRM markers enabled more reliable marker-trait association than the SCARs. Additionally, allelic dosage estimates for the H1 locus were also derived using the TG689_1P marker, providing a tool to optimise parental and progeny selections in PCN resistance breeding.     

QTL mapping for bacterial wilt resistance in peanut (<Emphasis Type="Italic">Arachis hypogaea</Emphasis> L.)

Bacterial wilt (BW) caused by Ralstonia solanacearum is a serious, global, disease of peanut (Arachis hypogaea L.), but it is especially destructive in China. Identification of DNA markers linked to the resistance to this disease will help peanut breeders efficiently develop resistant cultivars through molecular breeding. A F₂ population, from a cross between disease-resistant and disease-susceptible cultivars, was used to detect quantitative trait loci (QTL) associated with the resistance to this disease in the cultivated peanut. Genome-wide SNPs were identified from restriction-site-associated DNA sequencing tags using next-generation DNA sequencing technology. SNPs linked to disease resistance were determined in two bulks of 30 resistant and 30 susceptible plants along with two parental plants using bulk segregant analysis. Polymorphic SSR and SNP markers were utilized for construction of a linkage map and for performing the QTL analysis, and a moderately dense linkage map was constructed in the F₂ population. Two QTL (qBW-1 and qBW-2) detected for resistance to BW disease were located in the linkage groups LG1 and LG10 and account for 21 and 12 % of the bacterial wilt phenotypic variance. To confirm these QTL, the F₈ RIL population with 223 plants was utilized for genotyping and phenotyping plants by year and location as compared to the F₂ population. The QTL qBW-1 was consistent in the location of LG1 in the F₈ population though the QTL qBW-2 could not be clarified due to fewer markers used and mapped in LG10. The QTL qBW-1, including four linked SNP markers and one SSR marker within 14.4-cM interval in the F₈, was closely related to a disease resistance gene homolog and was considered as a candidate gene for resistance to BW. QTL identified in this study would be useful to conduct marker-assisted selection and may permit cloning of resistance genes. Our study shows that bulk segregant analysis of genome-wide SNPs is a useful approach to expedite the identification of genetic markers linked to disease resistance traits in the allotetraploidy species peanut.     

<Emphasis Type="Italic">LIG1</Emphasis> polymorphisms: the Indian scenario

Elucidation of the genetic diversity and relatedness of the subpopulations of India may provide a unique resource for future analysis of genetic association of several critical community-specific complex diseases. We performed a comprehensive exploration of single nucleotide polymorphisms (SNPs) within the gene DNA ligase 1 (LIG1) among a multiethnic panel of Indian subpopulations representative of the ethnic, linguistic and geographical diversity of India using a two-stage design involving DNA resequencing-based SNP discovery followed by SNP validation using sequenom-based genotyping. Thirty SNPs were identified in LIG1 gene using DNA resequencing including three promoter SNPs and one coding SNP. Following SNP validation, the SNPs rs20580/C19008A and rs3730862/C8804T were found to have the most widespread prevalence with noticeable variations in minor allele frequencies both between the Indian subpopulation groups and also from those reported on other major world populations. Subsequently, SNPs found in Indian subpopulations were analysed using bioinformatics-based approaches and compared with SNP data available on major world populations. Further, we also performed genotype–phenotype association analysis of LIG1 SNPs with publicly available data on LIG1 mRNA expression in HapMap samples. Results showed polymorphisms in LIG1 affect its expression and may therefore change its function. Our results stress upon the uniqueness of the Indian population with respect to the worldwide scenario and suggest that any epidemiological study undertaken on the global population should take this distinctiveness in consideration and avoid making generalized conclusions.