Molecular diversity and association of SSR markers to rust and late leaf spot resistance in cultivated groundnut (Arachis hypogaea L.)

Molecular diversity and association of simple sequence repeat (SSR) markers with rust and late leaf spot (LLS) resistance were detected in a set of 20 cultivated groundnut genotypes differing in resistance against both diseases. Out of 136 bands amplified from 26 primers, 104 were found polymorphic (76.5%). Cluster analysis (UPGMA) revealed two main clusters separated at 52% Jaccard's similarity coefficient according to disease reaction to rust and LLS. Based on the Kruskal–Wallis one-way anova and simple regression analysis three and four SSR alleles were found associated with rust and LLS resistance, respectively.     

Components of resistance to late leaf spot and rust among interspecific derivatives and their significance in a foliar disease resistance breeding in groundnut (Arachis hypogaea L.)

Late leaf spot (LLS) and rust cause substantial yield losses and reduce the fodder and seed quality in groundnut (Arachis hypogaea L.). Adoption of resistant cultivars by the semi-arid tropic farmers is the best option to overcome yield losses. Knowledge on components of resistance to these diseases should facilitate the development of groundnut cultivars with enhanced resistance to LLS and rust. The objectives of the experiments were to study the genetic variability and relationships among components of resistance to LLS and rust, and assess their significance in disease resistance breeding. Fifteen interspecific derivatives for LLS and 14 for rust and a susceptible control, TMV 2, were evaluated in a randomised complete block design with two or three replications under greenhouse conditions. The experiments were repeated twice. Genotypic differences were highly significant for all the traits studied. Resistance to LLS is due to longer incubation and latent periods, lesser lesions per leaf, smaller lesion diameter, lower sporulation index, and lesser leaf area damage and disease score. Selection based on components of resistance to LLS may not lead to plants with higher retained green leaf area. The remaining green leaf area on the plant should, therefore, be the major selection criteria for resistance to LLS in breeding programs. Resistance to rust is due to longer incubation and latent periods, fewer pustules per leaf, smaller pustule diameter, lower sporulation index, and lesser leaf area damage and disease score. Rust resistant components appear to work additively, therefore, selection based on resistance components together with green leaf area retained on the plant should be the basis of selecting for resistance to rust in breeding programs. ICGV 99005, 99003, 99012, and 99015 for rust and ICGV 99006, 99013, 99004, 99003, and 99001 for LLS are the better parents for use in resistance breeding programs. This revised version was published online in July 2006 with corrections to the Cover Date.     

RAPD markers linked to a rust resistance gene in cultivated groundnut (Arachis hypogaea L.)

Groundnut rust (Puccinia arachidis Speg.) is an important air borne pathogen, which causes substantial losses in groundnut yield and quality. Although large numbers of accessions were identified as rust resistant in wild, interspecific derivative and cultivated groundnut species, transfer of resistance to well-adapted cultivars is limited due to linkage drag, which worsens yield potential and market acceptance. A F₂ mapping population comprising 117 individuals was developed from a cross between the rust resistant parent VG 9514 and rust susceptible parent TAG 24. Rust resistance was governed by single dominant gene in this cross. We identified 11 (out of 160) RAPD primers that exhibited polymorphism between these two parents. Using a modified bulk segregant analysis, primer J7 (5′CCTCTCGACA3′) produced a single coupling phase marker (J7₁₃₅₀) and a repulsion phase marker (J7₁₃₀₀) linked to rust resistance. Screening of the entire F₂ population using primer J7 revealed that the coupling phase marker J7₁₃₅₀ was linked with the rust resistance gene at a distance of 18.5 cM. On the other hand, the repulsion phase marker J7₁₃₀₀ was completely linked with rust resistance. Additionally, both J7₁₃₀₀ (P = 0.00075) and J7₁₃₅₀ (P < 0.00001) were significantly associated with the rust resistance. Marker J7₁₃₀₀ identified all homozygous rust resistant genotypes in the F₂ population and was present in all the eight susceptible genotypes tested for validation. Thus, J7₁₃₀₀ should be applicable for marker-assisted selection (MAS) in the groundnut rust resistance breeding programme in India. To the best of our knowledge this is the first report on the identification of RAPD markers linked to rust resistance in groundnut.     

SSR markers associated for late leaf spot disease resistance by bulked segregant analysis in groundnut (Arachis hypogaea L.)

Late leaf spot (LLS) caused by Phaeoisariopsis personata is the major foliar disease that reduces the pod yield and severely affects the fodder and seed quality in groundnut. Molecular markers linked with LLS can improve the process of identification of resistant genotypes. In the present study, a LLS susceptible genotype (TMV 2) and the LLS resistant genotype (COG 0437) were crossed and their F₂ population was used for marker analysis. The phenotypic mean data on F_2:3 progenies were used as phenotype. Parents were surveyed with 77 SSR (Simple Sequence Repeat) primers to identify polymorphic markers. Among SSR markers, nine primers were found polymorphic between the parents TMV 2 and COG 0437. These markers were utilized for bulked segregant analysis (BSA). Among the polymorphic SSR markers, three primers viz., PM 375₁₆₂, pPGPseq5D5₂₂₀ and PM 384₁₀₀ were able to distinguish the resistant and susceptible bulks and individuals for LLS. In single marker analysis, the markers PM 375, PM 384, pPGPseq5D5, PM 137, PM 3, PMc 588 and Ah 4-26 were linked with LLS severity score. The phenotypic variation explained by these markers ranged from 32 to 59?%. The markers identified through BSA were also confirmed with single marker analysis. While validating the three primers over a set of resistant and susceptible genotypes, the primer PM 384₁₀₀ allele had association with resistance. Hence PM 384 could be utilized in the marker assisted breeding programme over a wide range of genetic background.     

Development of genic molecular markers linked to a rust resistance gene in cultivated groundnut (Arachis hypogaea L.)

Development of molecular markers for different economically important traits in cultivated groundnut has progressed at slow pace. Although many genomic SSR markers were developed in both the wild and cultivated groundnut, the genetic linkage map in the species is still not saturated. Availability of a large number of ESTs in GenBank opened up the possibility of integrating new markers and to identify markers closely linked to agronomic traits. EST-SSR markers are also considered as genic molecular markers. In this study, 259 EST-SSR markers were developed by mining 5,184 Arachis hypogaea ESTs from NCBI database. These EST-SSRs and 34 resistance gene candidate markers were used for association and genetic mapping of rust resistance in cultivated groundnut. From these, Cer2, SSR_GO340445, SSR_HO115759, SSR_GO341324 and RGC 2 had a significant association with rust resistance based on locus-by-locus AMOVA and/or Kruskal?CWallis ANOVA. Some of these associated markers also had protein activity related to biotic stress responses. Through genetic mapping, EST-SSR markers SSR_GO340445 and SSR_HO115759 were found closely linked to a rust resistance gene at 1.9 and 3.8?cM distances, respectively. These markers are thus suitable candidates for marker assisted selection in groundnut. The tight linkage of SSR_GO340445 would be helpful to screen BAC clones and to isolate rust resistance gene in groundnut.     

Foliar fungal disease‐resistant introgression lines of groundnut (Arachis hypogaea L.) record higher pod and haulm yield in multilocation testing

Introgression lines (ILs) of groundnut with enhanced resistance to rust and late leaf spot (LLS) recorded increased pod and haulm yield in multilocation testing. Marker‐assisted backcrossing (MABC) approach was used to introgress a genomic region containing a major QTL that explains >80% of phenotypic variance (PV) for rust resistance and 67.98% PV for LLS resistance. ILs in the genetic background of TAG 24, ICGV 91114 and JL 24 were evaluated for two seasons to select 20 best ILs based on resistance, productivity parameters and maturity duration. Multilocation evaluation of the selected ILs was conducted in three locations including disease hot spots. Background genotype, environment and genotype × environment interactions are important for expression of resistance governed by the QTL region. Six best ILs namely ICGV 13192, ICGV 13193, ICGV 13200, ICGV 13206, ICGV 13228 and ICGV 13229 were selected with 39–79% higher mean pod yield and 25–89% higher mean haulm yield over their respective recurrent parents. Pod yield increase was contributed by increase in seed mass and number of pods per plant.     

SSR标记与花生抗黄曲霉性状的关联分析

本文选用100对SSR引物对12个抗感黄曲霉花生品种的基因组DNA进行扩增,结果表明共有41对引物在不同品种间检测出2～4个等位基因,多态性信息量(PIC)为0.153～0.750.供试品种SSR标记基因型与黄曲霉侵染指数间的关联分析表明有5个标记与抗性相关,其中标记pPGSseq19D9与抗性关联度最高,Pearson相关系数达0.913.进一步观察发现pPGSseq19D9的扩增带型能直接区分抗感品种,初步推断pPGSseq19D9可能与一个贡献率较大的抗黄曲霉基因连锁.     

花生SSR标记与农艺性状的相关性

以农家品种四粒红和冀农黑3号构建的包含有251个家系的重组自交系(RIL)群体为材料, 在保定市和邯郸大名县两地进行表型鉴定, 利用Pearson’s相关和逐步多元回归分析了花生农艺性状之间及其与标记的相关性。结果表明, 多数农艺性状间存在显著或极显著相关, 其中相关性最高的为单株生产力和单株仁重(r=0.970), 其次是主茎高和第一侧枝长(r=0.918); 77对SSR标记与18个农艺性状显著相关, 每个性状相关标记数在2~16之间; 14个SSR标记与13个农艺性状关联, 解释的表型变异为5.2%~11.5%。以上结果为今后花生的常规育种和分子标记辅助选择奠定了基础。     

High level of natural variation in a groundnut (Arachis hypogaea L.) germplasm collection assayed by selected informative SSR markers

The ability to identify genetic variation is indispensable for effective management and use of genetic resources in crop breeding. Genetic variation among 189 groundnut ( Arachis hypogaea L.) accessions comprising landraces, cultivars, a mutant, advanced breeding lines and others (unknown genetic background) representing 29 countries and 10 geographical regions was assessed at 25 microsatellite or simple sequence repeat loci. A high number of alleles (265) were detected in the range of 3 (Ah1TC6G09) to 20 (Ah1TC11H06) with an average of 10.6 alleles per locus. The polymorphism information content value at these loci varied from 0.38 (Ah1TC6G09) to 0.88 (Ah1TC11H06) with an average of 0.70. A total of 59 unique alleles and 127 rare alleles were detected at almost all the loci assayed. Cluster analysis grouped 189 accessions into four clusters. In general, genotypes of South America and South Asia showed high level of diversity. Extraordinary level of natural genetic variation reported here provides opportunities to the groundnut community to make better decisions and define suitable strategies for harnessing the genetic variation in groundnut breeding.     

Marker‐assisted backcrossing to develop foliar disease‐resistant genotypes in TMV 2 variety of peanut (Arachis hypogaea L.)

TMV 2 is a very popular peanut variety among the Indian farmers, but it is highly susceptible to fungal foliar diseases like late leaf spot (LLS) and rust. Marker‐assisted backcrossing (MABC) in TMV 2 using foliar disease‐resistant donor, GPBD 4 and the disease resistance‐linked markers (GM2009, GM2079, GM2301, GM1839 and IPAHM103) resulted in a large number of backcross populations and also straight cross populations. Foreground selection followed by field evaluation under disease epiphytotic conditions could identify a few superior genotypes. Two homozygous backcross lines TMG‐29 and TMG‐46 showed enhanced resistance to LLS and rust diseases (score of 3.00 for both) along with 71.0% and 62.7% increase in the pod yield per plot, respectively, over the check, TMV 2. These foliar disease‐resistant and productive lines can be released as commercial varieties or can be used as genetic resources in the peanut improvement.     

Simple sequence repeat markers for botanical varieties of cultivated peanut (Arachis hypogaea L.)

Cultivated peanut (Arachis hypogaea L.) consists of six botanical varieties. Identification of DNA markers associated with botanical varieties would be useful in plant genotyping, germplasm management, and evolutionary studies. We have developed 130 simple sequence repeat (SSR) markers in peanut, 38 of which were used in this study because of their ability in detecting genetic polymorphism among 24 peanut accessions. Eight SSR markers were found useful to classify botanical varieties. Among them, six SSR markers were specific to botanical varieties fastigiata and vulgaris, one to botanical varieties hypogaea and hirsuta, and one to botanical varieties peruviana, and aequatoriana. Also, three of them derived from peanut expressed sequence tags (ESTs) were associated with putative genes. As botanical varieties have different morphological traits and belong to different subspecies in A. hypogaea, these markers might be associated with genes involved in the expression of morphological traits. The results also suggested that SSRs (also called microsatellites) might play a role in shaping evolution of cultivated peanut. Multiplex PCR of botanical variety-specific markers could be applied to facilitate efficient genotyping of the peanut lines.     

花生栽培种EST-SSRs分布特征及应用研究

利用自行开发的20 160条花生栽培种荚果EST, 通过序列拼接, 获得8 289条无冗余EST。经搜索, 共检测出740个SSR位点, 分布于651条EST中, 发生频率为7.8%, 平均每6.8 kb EST序列含一个SSR位点。功能注释结果表明具生物过程、分子功能和细胞组分的EST分别为73、111和56条。在花生荚果EST-SSR中, 三核苷酸重复类型出现频率最高, 占总SSR的62.8%, 其次是二核苷酸重复类型, 占总SSR的33.6%。在出现的26类重复基序中, AG/TC重复基序出现频率最高, AAG/TTC次之。利用Primer premier 5从651条含有SSR的EST中共设计引物233对, 从中随机选取100对引物检测EST-SSR在花生栽培种中的多态性及在野生种中的可转移性。结果表明, 有86对引物在供试的22个花生栽培品种中得到有效扩增, 其中10对在栽培种中具有多态性, 每对引物检测出的等位基因数2~3个, 平均2.2个。可扩增引物在野生种中的可转移率为12.5%~100%,平均96%。在野生种间检测出多态性的引物76对,每对引物检测出等位基因2~9个, 平均4.06个。     

Genetics,genomics and breeding of groundnut (Arachis hypogaea L.)

Groundnut is an important food and oil crop in the semiarid tropics, contributing to household food consumption and cash income. In Asia and Africa, yields are low attributed to various production constraints. This review paper highlights advances in genetics, genomics and breeding to improve the productivity of groundnut. Genetic studies concerning inheritance, genetic variability and heritability, combining ability and trait correlations have provided a better understanding of the crop's genetics to develop appropriate breeding strategies for target traits. Several improved lines and sources of variability have been identified or developed for various economically important traits through conventional breeding. Significant advances have also been made in groundnut genomics including genome sequencing, marker development and genetic and trait mapping. These advances have led to a better understanding of the groundnut genome, discovery of genes/variants for traits of interest and integration of marker‐assisted breeding for selected traits. The integration of genomic tools into the breeding process accompanied with increased precision of yield trialing and phenotyping will increase the efficiency and enhance the genetic gain for release of improved groundnut varieties.     

Identification and inheritance of male sterility in groundnut (Arachis hypogaea L.)

Summary Some plants without pods but with gynophores were observed in two F₄ progenies of two crosses of goundnut (Arachis hypogaea L.). The flowers on these plants had translucent white anthers with no or a few sterile pollen grains. Three such plants in the succeeding generation were hand pollinated with pollen from a short-duration Indian cv. JL 24. The resulting F₁ hybrid plants (male sterile x JL 24) were normal. Chi-square tests for segregation for male fertile and male sterile plants in F₂ and F₃ generations indicated that the male sterility in these crosses of groundnut is governed by two recessive genes. We designate these genes as ms₁ and ms₂ with ms₁ms₁ms₂ms₂ being a male sterile genotype.Submitted as ICRISAT J. A. No. 1812.     

Characterization of resistance to angular leaf spot of common bean (Phaseolus vulgaris L.) breeding line SPS50HB

The common bean (Phaseolus vulgaris L.) makes an important contribution to the human diet, particularly in Africa and Latin America. Because angular leaf spot (ALS), caused by the fungal pathogen Pseudocercospora griseola, is one of the most severe foliar diseases of the bean plant, an important priority is to identify genes encoding resistance. The present study focused on the resistance shown by the Mesoamerican common bean breeding line SPS50HB. From the pattern of segregation for resistance displayed in an F₂ population bred from a cross between SPS50HB and the ALS-susceptible Ethiopian variety Red Wolaita, it was concluded that the resistance of SPS50HB is controlled by two unlinked dominant genes. An allelism test indicated that one of these genes was either identical with, allelic to, or closely linked to the major gene Phg-2 carried by variety Mexico 54. The sequence-characterized amplified region assays OPEO4 and PF13, which are diagnostic for an ALS resistance gene carried by the germplasm accession G10909, both tracked a possible second gene present in SPS50HB.