ISSR and SCAR markers linked to the mungbean yellow mosaic virus (MYMV) resistance gene in blackgram [Vigna mungo (L.) Hepper]

A recombinant inbred line (RIL) mapping population (F₈) was generated by crossing Vigna mungo (cv. TU 94‐2) with Vigna mungo var. silvestris and screened for mungbean yellow mosaic virus (MYMV) resistance. The inter simple sequence repeat (ISSR) marker technique was employed to identify markers linked to the MYMV resistance gene. Of the 100 primers screened, 54 showed amplification of which 36 exhibited polymorphism between the parents TU 94‐2 (resistant) and V. mungo var. silvestris (susceptible). Individual plants from 53 RIL populations were analysed and one marker (ISSR811₁₃₅₇) was identified as tightly linked to the MYMV resistant gene at 6.8 cM. Both the phenotype as well as the ISSR811₁₃₅₇ marker segregated in a 1 : 1 ratio. The ISSR811₁₃₅₇ marker was sequenced and sequence characterized amplified region (SCAR) primers were designed (YMV1‐F and YMV1‐R) to amplify the marker. Screening for the SCAR marker in the RIL population distinguished the MYMV resistant and susceptible plants, agreeing well with the phenotypic data. The ISSR811₁₃₅₇ marker was validated using diverse blackgram genotypes differing in their MYMV reaction. The marker will be useful for the development of MYMV‐resistant genotypes in blackgram.     

QTL mapping of a partial resistance to the corn delphacid‐transmitted viruses in Lepidopteran‐resistant maize line Mp705

A partial resistance to maize mosaic virus (MMV) and maize stripe virus (MStV) was mapped in a RILs population derived from a cross between lines MP705 (resistant) and B73 (susceptible). A genetic map constructed from 131 SSR markers spanned 1399 cM with an average distance of 9.6 cM. A total of 10 QTL were detected for resistance to MMV and MStV, using composite interval mapping. A major QTL explaining 34–41% of the phenotypic variance for early resistance to MMV was detected on chromosome 1. Another major QTL explaining up to 30% of the phenotypic variation for all traits of resistance to MStV was detected in the centromeric region of chromosome 3 (3.05 bin). After adding supplementary SSR markers, this region was found to correspond well to the one where a QTL of resistance to MStV already was located in a previous mapping study using an F₂ population derived from a cross between Rev81 and B73. These results suggested that these QTL of resistance to MStV detected on chromosome 3 could be allelic in maize genome.     

User‐friendly markers linked to Fusarium wilt race 1 resistance Fw gene for marker‐assisted selection in pea

Fusarium wilt is one of the most widespread diseases of pea. Resistance to Fusarium wilt race 1 was reported as a single gene, Fw, located on linkage group III. The previously reported AFLP and RAPD markers linked to Fw have limited usage in marker‐assisted selection due to their map distance and linkage phase. Using 80 F₈ recombinant inbred lines (RILs) derived from the cross of Green Arrow × PI 179449, we amplified 72 polymorphic markers between resistant and susceptible lines with the target region amplified polymorphism (TRAP) technique. Marker–trait association analysis revealed a significant association. Five candidate markers were identified and three were converted into user‐friendly dominant SCAR markers. Forty‐eight pea cultivars with known resistant or susceptible phenotypes to Fusarium wilt race 1 verified the marker–trait association. These three markers, Fw_Trap_480, Fw_Trap_340 and Fw_Trap_220, are tightly linked to and only 1.2 cM away from the Fw locus and are therefore ideal for marker‐assisted selection. These newly identified markers are useful to assist in the isolation of the Fusarium wilt race 1 resistance gene in pea.     

Genetic analysis of gene‐specific resistance to mungbean yellow mosaic virus in mungbean (Vigna radiata)

Six intervarietal crosses involving two resistant and three susceptible genotypes of mungbean were attempted with the objectives to determine the mode of inheritance of gene‐specific Mungbean Yellow Mosaic Virus (MYMV) resistance. An infector row technique along with artificial inoculation was used for evaluating parents, F₁, F₂ and F₃ plants for MYMV resistance. Disease scoring for MYMV indicated that F₁s were highly susceptible as were the susceptible parents while resistant parent exhibited resistant reaction. The F₂ progeny segregated in the ratio of 9 S:3 MS:3 MR:1 R suggesting that the resistance was governed by digenic recessive genes (r_m1 and r_m2). When one gene (r_m1) was present in the homozygous recessive condition in different plants, it conferred moderately susceptible (MS) reaction, whereas when other gene (r_m2) was in homozygous condition, moderately resistant (MR) reaction was obvious. When both genes (r_m1 and r_m2) were present together in the homozygous recessive condition, resistant reaction (R) was observed. The F₂ segregation explained on the basis of phenotypic expression was further confirmed by F₃ segregation.     

Characterization of broad‐spectrum resistance to Soybean mosaic virus in soybean [Glycine max (L.) Merr.] cultivar ‘RN‐9’

Soybean mosaic virus (SMV) can cause serious yield losses in soybean. Soybean cultivar ‘RN‐9’ is resistant to 15 of 21 SMV strains. To well‐characterize this invaluable broad‐spectrum SMV‐resistance, populations (F₁, F₂ and F_2:3) derived from resistant (R) × susceptible (S) and R × R crosses were tested for SMV‐SC18 resistance. Genetic analysis revealed that SC18 resistance in ‘RN‐9’ plus two elite SMV‐resistant genotypes (‘Qihuang No.1’ and ‘Kefeng No.1’) are controlled by independently single dominant genes. Linkage analysis showed that the resistance of ‘RN‐9’ to SMV strains SC10, SC14, SC15 and SC18 is controlled by more than one gene(s). Moreover, Rsc10‐r and Rsc18‐r were both positioned between the two simple sequence repeats markers Satt286 and Satt277, while Rsc14‐r was fine‐mapped in 136.8‐kb genomic region containing sixteen genes, flanked by BARCSOYSSR_06_0786 and BARCSOYSSR_06_0790 at genetic distances of 3.79 and 4.14 cM, respectively. Allelic sequence comparison showed that Cytochrome P450‐encoding genes (Glyma.06g176000 and Glyma.06g176100) likely confer the resistance to SC14 in ‘RN‐9’. Our results would facilitate the breeding of broad‐spectrum and durable SMV resistance in soybeans.     

The first genetic linkage map of crape myrtle (Lagerstroemia) based on amplification fragment length polymorphisms and simple sequence repeats markers

Lagerstroemia (crape myrtle) are famous ornamental plants with large pyramidal racemes, long flower duration and diverse colours. Genetic maps provide an important genomic resource of basic and applied significance. A genetic linkage map was developed by genotyping 192 F₁ progeny from a cross between L. caudata (female) and L. indica (‘Xiang Xue Yun’) (male) with a combination of amplification fragment length polymorphisms (AFLP) and simple sequence repeats (SSR) markers in a double pseudo‐testcross mapping strategy. A total of 330 polymorphic loci consisting of 284 AFLPs and 46 SSRs showing Mendelian segregation were generated from 383 AFLP primer combinations and 150 SSR primers. The data were analysed using JoinMap 4.0 (evaluation version) to construct the linkage map. The map consisted of 20 linkage groups of 173 loci (160 AFLPs and 13 SSRs) covering 1162.1 cM with a mean distance of 10.69 cM between adjacent markers. The 20 linkage groups contained 2–49 loci and ranged in length from 7.38 to 163.57 cM. This map will serve as a framework for mapping QTLs and provide reference information for future molecular breeding work.     

Molecular mapping of a locus controlling resistance to Albugo candida in Indian mustard

Brassica juncea (L.) Czern & Coss is widely grown as an oilseed crop in the Indian subcontinent. White rust disease caused by Albugo candida (Pers.) Kuntze is a serious disease of this crop causing considerable yield loss every year. The present study was undertaken to identify molecular markers for the locus controlling white rust resistance in a mustard accession, BEC‐144, using a set of 94 recombinant inbred lines (RILs). The screening of individual RILs using an isolate highly virulent on the popular Indian cultivar ‘Varuna’ revealed the presence of a major locus for rust resistance in BEC‐144. Based on screening of 186 decamer primers employing bulked segregant analysis (BSA), 11 random amplified polymorphic DNA markers were identified, which distinguished the parental lines and the bulks. Five of these markers showed linkage with the rust resistance locus. Two markers, OPN0_l000 and OPB06₁₀₀₀, were linked in coupling and repulsion phases at 9.9 cM and 5.5 cM, respectively, on either side of the locus. The presence of only two double recombinants in a population of 94 RILs suggested that the simultaneous use of both markers would ensure efficient transfer of the target gene in mustard breeding programmes.     

Identification of molecular markers associated with resistance to squash silverleaf disorder in summer squash (<Emphasis Type="Italic">Cucurbita</Emphasis> <Emphasis Type="Italic">pepo</Emphasis>)

Squash silverleaf (SSL), caused by the silverleaf whitefly [Bemisia argentifolii (formerly known as Bemisia tabaci Gennadius, B strain)], is an important physiological disorder that affects squash (Cucurbita spp.) by reducing yield potential. Breeding squash with resistance to SSL disorder can be facilitated by using marker-assisted selection (MAS). Resistance to SSL disorder, in Cucurbita pepo, is conferred by a single recessive gene (sl). The objective of this study was to identify molecular markers associated with resistance. A zucchini squash, SSL disorder resistant breeding line, ‘Zuc76’ (sl/sl) and a SSL disorder susceptible zucchini cultivar ‘Black Beauty’ (Sl/Sl) were screened with 1,152 randomly amplified polymorphic DNA (RAPD) primers and 432 simple sequence repeat (SSR) markers to identify polymorphisms. Using F₂ and BC₁ progeny segregating for SSL disorder resistance, three RAPD (OPC07, OPL07 and OPBC16) primers and one SSR (M121) marker were found associated with sl. Fragments amplified by RAPD primer OPC07 was linked in coupling phase to sl, whereas RAPD primer OPL07 was linked in repulsion phase. RAPD primer OPBC16 and SSR marker M121 were co-dominant. The allelic order of these loci was found to be M121–sl–OPC07–OPL07–OPBC16. The closest marker to sl is M121 with an estimated genetic distance of 3.3 cM. The markers identified in this study will be useful for breeding summer squash (C. pepo) for SSL disorder resistance derived from zucchini squash breeding line ‘Zuc76’.     

Efficient development of Haynaldia villosa chromosome 6VS‐specific DNA markers using a CISP‐IS strategy

Development of effective molecular markers linked to Pm21 deriving from Haynaldia villosa is critical for wheat breeding of powdery mildew resistance. In this study, we designed 12 pairs of conserved‐intron scanning primers (CISPs), using intron‐containing conserved genes located on the short arm of Brachypodium distachyon chromosome 3 (3BdS) aligned with cDNA or expressed sequence tags (ESTs) of Triticeae crops. Of 12 CISP primer pairs, 11 amplified DNA both in H. villosa and in wheat, and four displayed H. villosa chromosome 6VS‐specific polymorphisms. Six non‐polymorphic DNAs were further sequenced for designing internal primers, and five additional 6VS‐specific markers were obtained. Of the total nine 6VS‐specific co‐dominant markers, six could effectively trace Pm21 in F₂ population derived from the hybrid between the T6AL.6VS line and ‘Yangmai 158’. This study demonstrated that Brachypodium genomic information could be powerfully utilized to develop molecular markers in H. villosa or other Triticeae species.     

Mapping of male sterility gene ms10 in chilli pepper (Capsicum annuum L.)

Most of the hybrid seed in chilli are produced manually, but the use of male sterility (MS) can reduce the cost of hybrid seed production. MS‐12, a nuclear male‐sterile (NMS) line developed at Punjab Agricultural University, Ludhiana (India), has been utilized to develop commercial F₁ hybrids. A recessive gene, designated as ms10, governs MS in MS‐12. Due to recessive gene control, development of new NMS lines incorporating ms10 gene is tedious and time‐consuming. We identified SSR markers AVRDC‐PP12 and AVRDC_MD997* linked to the ms10 gene. A total of 558 primer pairs were screened following bulked segregant analysis (BSA). Linkage analysis in 210 F₂ plants indicated that the two SSR markers were linked to the ms10 gene and the marker AVRDC‐PP12 was closest to the gene at 7.2 cM distance. The marker was mapped to chromosome 1 at genome position 175 694 513 to 175 694 644. Until more closely linked markers are developed, the marker AVRDC‐PP12 would facilitate transfer of ms10 gene through marker‐assisted selection (MAS). Fine mapping would lead to cloning of the ms10 gene.     

Identification and mapping of a novel Turnip mosaic virus resistance gene TuRBCS01 in Chinese cabbage (Brassica rapa L.)

We aimed to identify Turnip mosaic virus (TuMV) resistance genes in Chinese cabbage by analysing the TuMV resistance of 43 P₁ (resistant), 88 P₂ (susceptible), 26 F₁, 104 B₁ (F₁ × P₁), 108 B₂ (F₁ × P₂) and 509 F₂ individuals. All parents and progeny populations were mechanically inoculated with TuMV‐C4. Both F₁ and B₁ populations showed TuMV resistance. Resistant: susceptible ratios in the B₂ and F₂ populations were 1 : 1 and 3 : 1, respectively. TuMV resistance in P₁ was controlled by a dominant gene, TuRBCS01. Bulked segregation analysis was performed to identify simple sequence repeat or insertion or deletion markers linked to TuRBCS01. Data from 108 B₂ individuals with resistant or susceptible phenotypes were analysed using mapmake r/exp 3.0. Polymorphic marker sequences were blast searched on http://brassicadb.org/brad/ . TuRBCS01 was found to be linked to eight markers: SAAS_mDN192117a_159 (3.3 cM), SAAS_mDN192117b_196 (4.0 cM), SAAS_mDN192403_148 (13.0 cM), SAAS_mGT084561_233 (6.8 cM), BrID10723 (3.3 cM), mBr4041 (3.3 cM), SAAS_mBr4055_194 (2.6 cM) and mBr4068 (4.0 cM). Further, TuRBCS01 was mapped to a 1.98‐Mb region on chromosome A04 between markers BrID10723 and SAAS_mBr4055_194.     

Detection of RAPD markers linked to the everbearing gene in Japanese cultivated strawberry

To identify markers for the everbearing gene in strawberries, 199 F₁ progeny plants were produced from a cross between ‘Ever Berry’ (a Japanese everbearing strawberry) and ‘Toyonoka’ (a Japanese Junebearing strawberry) as the experimental population. The results of flowering tests produced 97 everbears and 102 Junebears. The chi‐square test gave a goodness of fit for the expected ratio of 1 : 1 for everbears to Junebears, suggesting the inheritance of the everbearing trait is controlled by a monogenic dominant gene. RAPD analyses on this trait were carried out using ‘Ever Berry’ and ‘Toyonoka’. Seventy‐one primers, which produced 89 polymorphic fragments between the two parents, were identified from a total of 175 primers. Five markers relating to the everbearing trait were selected from 26 of the 199 progeny plants. The remaining 173 seedlings were analysed with these five markers and a linkage map was constructed using all of the 199 F₁ progeny plants. The length of this linkage group is 39.7 cM. The closest markers found, OPE07‐1 and OPB05‐1, are respectively mapped at 11.8 and 15.8 cM on each side of the everbearing gene.     

Inheritance and chromosome location of photoperiod‐thermo sensitive male sterility in wheat line Xinong 291S

The present research aimed to study the inheritance and chromosomal location of a photoperiod‐thermo sensitive male sterility (PTSMS) gene in Xinong 291S (XN291S), which is a new PTSMS wheat line. The inheritance was studied in F₁ and F₂ populations derived from crosses between XN291S and eight wheat cultivars. All F₁ plants were fertile and the F₂ populations segregated in either 15 : 1 or 3 : 1 fertile : sterile ratios indicating that PTSMS was controlled by one or two recessive major genes. Five wheat cultivars carried homozygous dominant alleles for fertility, whereas the other three, including ‘Chinese Spring’ (CS), carried a single homozygous dominant allele. Chromosomal location of the PTSMS gene was studied by crossing a set of CS nulli‐tetrasomic lines to XN291S. Self‐fertility of F₁ hybrids XN291S/N5BT5A and XN291S/N5BT5D were significantly lower than the others. Therefore, the PTSMS gene in XN291S that differed from CS was located on chromosome 5B. Location of the second PTSMS gene needs further study.     

Mapping powdery mildew resistance gene in V97‐3000 soybean

V97‐3000 is a maturity group (MG) V soybean breeding line derived from SS 516 × V90‐2592 (Vance × V81‐1325) with high stachyose, small seed and powdery mildew resistance. A total of 53 F_2:3 families were derived from a cross between V97‐3000 and a powdery mildew susceptible line V99‐5089. The 53 F_2:3 families, each with 30 plants, were grown in the greenhouse for powdery mildew evaluation, and the corresponding 53 F₂ plants were genotyped using simple sequence repeat (SSR) markers. Results showed that the 53 F_2:3 families segregated in ratio of one resistant : two segregating : one susceptible (13 : 26 : 14) and the 26 segregating F_2:3 families each exhibited a good fit to three resistant : one susceptible, indicating that resistance to powdery mildew is conditioned by a single dominant gene. The gene for powdery mildew resistance in V97‐3000 was mapped on chromosome 16 [linkage group (LG) J] flanked by Satt547 and Sat_396 on one side and Sat_393 on the other side with 3.8 cM and 3.9 cM distance, respectively. This study provides a new source of powdery mildew resistance and information of genetic location of the resistance gene and linked markers, which is useful for breeders selecting powdery mildew resistance through marker‐assisted selection (MAS) in soybean breeding programmes.     

Development of molecular markers linked to the Leptosphaeria maculans resistance gene Rlm6 and inheritance of SCAR and CAPS markers in Brassica napus × Brassica juncea interspecific hybrids

Leptosphaeria maculans causes blackleg disease on Brassica napus, an economically important oilseed crop. Brassica juncea has high resistance to blackleg and is a source for the development of resistant B. napus varieties. To transfer the Rlm6 resistance gene from B. juncea into B. napus, an interspecific cross between B. napus “Topas DH16516” and B. juncea “Forge” was produced, followed by the development of F₂ and F₃ generations. Sequence characterized amplified region (SCAR) and cleaved amplified polymorphic sequence (CAPS) markers linked to the L. maculans resistance gene Rlm6 were developed. Segregation of SCAR and CAPS markers linked to Rlm6 were confirmed by genotyping of F₂ and F₃ progeny. Segregation of CAPS markers and phenotypes for blackleg disease severity in F₂ plants had a Mendelian ratio of 3:1 in resistant vs. susceptible plants, respectively, supporting the assumption that genetic control of resistance was by a single dominant gene. The molecular markers developed in this study, which show linkage with the L. maculans resistance gene Rlm6, would facilitate marker‐assisted backcross breeding in a variety development programme.     

Genetic analysis and fine mapping of tms9, a novel thermosensitive genic male‐sterile gene in rice (Oryza sativa L.)

Two‐line hybrid rice as a novel hybrid breeding method has huge potential for yield increasing and utilization of intersubspecific heterosis, and it is of major significance for the food security of rice‐consuming populations. Zhu1S is a thermosensitive genic male‐sterile line of rice with low critical temperature and excellent combining ability, which has been widely exploited as a female parent in Chinese two‐line hybrid rice breeding. Here, genetic mapping in F₂ populations was used to show that its male sterility is inherited as a single recessive gene and that responsible gene (termed tms9) lies on the short arm of chromosome 2. A high‐resolution linkage analysis which was based on the Zhu1S/R173 F₂ population found that the thermosensitive genic male‐sterile gene tms9 of Zhu1S was fine mapped between insertion–deletion (Indel) markers Indel 37 and Indel 57, and the genetic distance from the tms9 to the two markers was 0.12 and 0.31 cM, respectively. The physical distance between the two markers was about 107.2 kb. Sequence annotation databases showed that the two Indel markers (Indel 37 and Indel 57) were located on two BAC clones (B1307A11 and P0027A02). There are sixteen open reading frames (ORF) present in this region. The results of this study are of great significance for further cloning tms9 and molecular marker–assisted selection.     

Integration of AFLPs,SSRs and SNPs markers into a new genetic map of industrial chicory (Cichorium intybus L. var. sativum)

A chicory genetic map of 1208 cM has been created using 247 F₂ plants and 237 markers (170 AFLP, 28 SSR, 27 EST‐SNP and 12 EST‐SSR). This map covers 84% of the chicory genome. The chicory‐genic‐markers‐associated sequences were used to find potential orthologs in mapped lettuce ESTs from the Compositae Genome Project Database. Twenty‐seven putative orthologous pairs were retained, pinpointing seven putative blocks of synteny that covered 11% of the chicory genome and 13% of the lettuce genome, opening new perspectives for the analysis of these two species.     

Inheritance of resistance to the peach root‐knot nematode (Meloidogyne floridensis) in interspecific crosses between peach (Prunus persica) and its wild relative (Prunus kansuensis)

The peach root‐knot nematode, Meloidogyne floridensis (MF), infects majority of available nematode‐resistant peach rootstocks which are mostly derived from peach (Prunus persica) and Chinese wild peach (P. davidiana). Interspecific hybridization of peach with its wild relative, Kansu peach (P. kansuensis), offers potential for broadening the resistance spectrum in standard peach rootstocks. We investigated the inheritance of resistance to MF in segregating populations of peach (‘Okinawa’ or ‘Flordaguard’) × P. kansuensis. A total of 379 individuals from 13 F₂ and BC₁F₁ families were challenged with a pathogenic MF isolate “MFGnv14” and were classified as resistant (R) or susceptible (S) based on root galling intensity. Segregation analyses in F₂ progeny revealed the involvement of a major locus with a dominant or recessive allele determining resistance in progeny segregating 3R:1S and 1R:3S, respectively. Testcrosses with a homozygous‐susceptible peach genotype (‘Flordaguard’ or ‘UFSharp’) confirmed P. kansuensis as a source of new resistance and the heterozygous allelic status of P. kansuensis at the locus conferring resistance to MF. We propose a single‐locus dominant/recessive model for the inheritance of resistance.     

Broadening the genetic base of lentil cultivars through inter‐sub‐specific and interspecific crosses of Lens taxa

Wild Lens taxa are invaluable sources of useful traits for broadening genetic base of cultivated lentil. Nine inter‐sub‐specific and interspecific crosses were made successfully between cultivated (Lens culinaris ssp. culinaris) and wild lentils (L. culinaris ssp. orientalis, odemensis, lamottei and ervoides). The effect of species groups, day length and temperature on crossability in lentils was evident under normal winter sowing in New Delhi and in summer Himalayan nursery at Sangla in Himachal Pradesh, India, although pollen fertility assessed in all the cross‐combinations showed no significant variation. True hybridity of nine inter‐sub‐specific and interspecific crosses was confirmed through morphological and molecular (ISSR) markers, in which three of 120 primers could confirm the hybridity of all the crosses. All cross‐combinations were also studied for important quantitative traits related to yield. The range, mean and coefficient of variation were estimated in parental lines, F₁ and F₂ generations to determine the extent of variability generated in cultivated lentils through the introgression of genes from wild L. taxa. A high level of heterosis was observed in F₁ crosses for important traits studied. Substantially higher variations for seed yield and its attributing traits were exhibited in F₂ generations indicating transgressive segregation. The results of the present investigation revealed that wild L. taxa can be successfully exploited for lentil improvement programmes, and the variations generated could be easily utilized for broadening the genetic base of cultivated lentil gene pool for improving the yield as well as wider adaptation.     

A novel aphid resistance locus in cowpea identified by combining SSR and SNP markers

The utility of combining simple sequence repeat (SSR) and single nucleotide polymorphism (SNP) marker genotyping was determined for genetically mapping a novel aphid (Aphis craccivora) resistance locus in cowpea breeding line SARC 1‐57‐2 and for introgressing the resistance into elite cultivars by marker‐assisted backcrossing (MABC). The locus was tagged with codominant SSR marker CP 171F/172R with a recombination fraction of 5.91% in an F₂ population from ‘Apagbaala’ x SARC 1‐57‐2. A SNP‐genotyped biparental recombinant inbred line population was genotyped for CP 171F/172R, which was mapped to position 11.5 cM on linkage group (LG) 10 (physical position 30.514 Mb on chromosome Vu10). Using CP 171F/172R for foreground selection and a KASP‐SNP‐based marker panel for background selection in MABC, the resistance from SARC 1‐57‐2 was introduced into elite susceptible cultivar ‘Zaayura’. Five BC₄F₃ lines of improved ‘Zaayura’ that were isogenic except for the resistance locus region had phenotypes similar to SARC 1‐57‐2. This study identified a novel aphid resistance locus and demonstrated the effectiveness of integrating SSR and SNP markers for trait mapping and marker‐assisted breeding.