Effectiveness of quantitative resistance conferred by the genetic background of pepper in the control of root‐knot nematodes and influence onto durability of Me1‐ and Me3‐resistant genes in greenhouse conditions

In pepper (Capsicum annuum), the major genes (R‐genes) Me1 and Me3 confer resistance against root‐knot nematodes (Meloidogyne spp.). The combination of R‐genes and quantitative resistance factors in the same genotype is considered a good breeding strategy for increasing the durability of R‐genes. To ascertain this hypothesis, five pepper inbred lines, differing in their quantitative resistance level, were combined with Me1 or Me3 genes in F₁ hybrids. The resistance of inbred lines and F₁ hybrids was evaluated in a greenhouse with soil naturally infected by M. incognita in two successive growing years. In both years, lines carrying Me3 were less infected by the nematode when combined with quantitative resistance. An increase in nematode infection was observed in the second growing year in lines carrying Me1 or Me3, independently of quantitative resistance. The infection level recorded in inbred lines without R‐genes was similar in both years. The effectiveness of quantitative resistance controlling M. incognita is confirmed in greenhouse conditions, although the durability of Me1 and Me3 when combined with quantitative resistance factors was not seen to increase.     

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.     

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.     

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.     

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.     

Development of gene‐based markers for the Turnip mosaic virus resistance gene retr02 in Brassica rapa

Turnip mosaic virus (TuMV) is responsible for a serious disease that affects the production of Chinese cabbage. Previous studies have cloned a series of TuMV resistance genes and developed molecular markers. In this study, a derived cleaved amplified polymorphism sequence (dCAPS) marker and a Kompetitive Allele Specific PCR (KASP) marker were developed based on a single recessive gene, retr02, which confers broad‐spectrum TuMV resistance in Chinese cabbage by means of an additional G at the junction of exon 1 and intron 1. The two markers were able to detect the retr02 allele in Chinese cabbage accessions used in breeding programmes. Compared with the dCAPS marker, the KASP marker was flexible, cost‐effective and quick to process, which is likely to be beneficial in establishing high‐throughput assays for marker‐assisted selection.     

Identification and marker‐assisted introgression of QTL conferring resistance to bacterial leaf blight in cowpea (Vigna unguiculata (L.) Walp.)

Bacterial leaf blight (BLB), caused by Xanthomonas axonopodis pv. vignicola (Xav), is widespread in major cowpea [Vigna unguiculata (L.) Walp.] growing regions of the world. Considering the resource poor nature of cowpea farmers, development and introduction of cultivars resistant to the disease is the best option. Identification of DNA markers and marker‐assisted selection will increase precision of breeding for resistance to diseases like bacterial leaf blight. Hence, an attempt was made to detect QTL for resistance to BLB using 194 F_2 : 3 progeny derived from the cross ‘C‐152’ (susceptible parent) × ‘V‐16’ (resistant parent). These progeny were screened for resistance to bacterial blight by the leaf inoculation method. Platykurtic distribution of per cent disease index scores indicated quantitative inheritance of resistance to bacterial leaf blight. A genetic map with 96 markers (79 SSR and 17 CISP) constructed from the 194 F₂ individuals was used to perform QTL analysis. Out of three major QTL identified, one was on LG 8 (qtlblb‐1) and two on LG 11 (qtlblb‐2 and qtlblb‐3). The PCR product generated by the primer VuMt337 encoded for RIN2‐like mRNA that positively regulate RPM1‐ and RPS2‐dependent hypersensitive response. The QTL qtlblb‐1 explained 30.58% phenotypic variation followed by qtlblb‐2 and qtlblb‐3 with 10.77% and 10.63%, respectively. The major QTL region on LG 8 was introgressed from cultivar V‐16 into the bacterial leaf blight susceptible variety C‐152 through marker‐assisted backcrossing (MABC).     

Inheritance of resistance to broomrape (Orobanche cumana Wallr.) race F in a sunflower line derived from wild sunflower species

Genetic resistance to broomrape (Orobanche cumana Wallr.) race F in sunflower line J1, derived from the wild perennial species Helianthusgrosseserratus Martens and Helianthus divaricatus L., has been reported to be controlled by dominant alleles at a single locus, Or6. However, deviations from this monogenic inheritance have been observed. The objective of the present study was to gain insight into the inheritance of resistance to broomrape race F in the sunflower line J1. F₁, F₂, F₃ and BC generations from crosses between J1 and three susceptible lines, P21, NR5 and HA821 were evaluated. F₁ hybrids showed both resistant (R) and moderately resistant (MR) plants, the latter having a maximum of five broomrape stalks per plant compared with >10 in the susceptible parents. This indicated incomplete dominance of the Or6 alleles. F₂ plants were classified as R, MR or susceptible (more than five broomrape stalks per plant). Three different segregation ratios were observed: 3 : 1, 13 : 3 and 15 : 1 (R + MR : S), suggesting the presence of a second gene, Or7, whose expression was influenced by the environment. A digenic model was confirmed, based on the evaluation of F_2:3 families.     

QTL analysis for cooking traits of super rice with a high‐density SNP genetic map and fine mapping of a novel boiled grain length locus

Hybrid rice has contributed substantially to the improvement of grain production worldwide, yet its poor cooking and tasting characteristics have long been recognized. In this study, 132 recombinant inbred lines derived from LYPJ were used to identify quantitative trait loci (QTLs) for 12 cooking traits with the high‐density SNP linkage map recently developed by our team. We identified 17 QTLs on chromosomes 1, 2, 4, 5, 6, 7, 8, 9 and 11, which accounted for 7.50% to 23.50% of the phenotypic variations. A novel major QTL qBGL7 for boiled grain length was further fine‐mapped to an interval of 440 Kb between the two markers RM21906 and gl3 using a BC₃F₂ population. Two near‐isogenic lines with extreme boiled grain length, GX5‐176 and GX5‐101, could be directly used in improving cooking quality. We also identified a QTL for soaked grain width expansion rate, qSGWE6, in the Wx gene region on chromosome 6. The Wx differential regulation coincided with sequential variation between the two parents. Our work offered a theoretical basis for molecular breeding of high‐quality hybrid rice.     

Marker‐assisted selection for rice blast resistance genes Pi2 and Pi9 through high‐resolution melting of a gene‐targeted amplicon

The use of host resistance (R) genes is considered the most cost‐effective option to control the rice blast disease. The two allelic R genes Pi2 and Pi9 confer very broad‐spectrum resistance against blast isolates collected worldwide. However, the two genes have not yet been widely deployed in rice breeding programmes. Availability of specific markers for them would facilitate incorporating the two R genes into new rice lines through marker‐assisted selection. Herein, we report the development and utilization of a robust and specific marker for the Pi2 and Pi9. This marker was derived from polymorphisms within the target gene, and achieved simultaneously distinguish Pi2 and Pi9 from other alleles through high‐resolution melting of a small amplicon. With the marker, we were able to transfer the Pi2 into an elite restorer line through marker‐assisted backcrossing, successfully obtained effective resistance to blast disease, and we were also able to, respectively, incorporate the Pi2 and Pi9 with two other R genes. As the additive effect, blast resistance in these stacking lines harbouring three R genes were significantly improved.     

Pyramiding three rust‐resistance genes confers a high level of resistance in soybean (Glycine max)

Pyramiding Asian soybean rust (ASR) resistance (Rpp) genes in a single genotype has been shown to increase ASR resistance in soybean. However, it remains unclear which combinations of Rpp genes are superior. Therefore, here, we developed six new Rpp‐pyramided lines carrying different combinations of Rpp genes and evaluated their resistance against 13 Bangladeshi rust (Phakopsora pachyrhizi) isolates (BdRPs) alongside seven previously developed Rpp‐pyramided lines. We found that lines carrying one, two and three Rpp genes had high ASR resistance without sporulation in 13.8%, 35.2% and 73.1% of the assays, respectively. Among the new lines that were developed, those with Rpp3 + Rpp4 and Rpp3 + Rpp4 + Rpp5 had high levels of ASR resistance, while the line containing Rpp2 + Rpp4 + Rpp5 showed immunity phenotype at two weeks after inoculation by the BdRP‐22 infection. Thus, pyramiding larger numbers of Rpp genes confers soybean with a higher level of resistance to ASR pathogens and can produce an immunity phenotype at two weeks after inoculation.     

Evaluation of the Ph‐3 gene‐specific marker developed for marker‐assisted selection of late blight‐resistant tomato

Late blight caused by Phytophthora infestans is one of the most destructive diseases of tomato (Solanum lycopersicum L.) that mainly occurs in cool and wet environments. With the spread of the A2 mating type and new clonal lineages, fewer fungicides provide effective control of the disease, which has increased its worldwide threat. Host resistance could contribute significantly to sustainable disease control. Ph‐3 is a race‐specific late blight resistance gene commonly used in commercial tomato breeding. Availability of precise and easy to use gene‐based markers would facilitate selection. In this study, a Ph‐3 on‐gene cleaved amplified polymorphic sequence (CAPS) marker, Ph3.gsm/HincII, was developed based on the published gene sequence of Ph‐3. The effectiveness of the marker was evaluated along with other published Ph‐3 markers using an F₉ recombinant inbred line (RIL) population derived from NC 23E‐2(93) × L3708. Markers Ph3.gsm/HincII and TG328/BstNI accurately genotyped the RIL population for Ph‐3. In addition, Ph3.gsm/HincII was able to differentiate variable susceptible alleles. This reliable codominant DNA marker would be very useful in marker‐assisted selection, particularly for resistance gene pyramiding.     

A genetic linkage map in southern‐by‐spring oat identifies multiple quantitative trait loci for adaptation and rust resistance

We developed 178 recombinant inbred lines from a southern‐by‐spring oat population designated as “TxH.” These lines were genotyped to generate a high‐quality linkage map that resolved 6,902 markers into 21 linkage groups that matched closely with the latest hexaploid oat consensus map. Three major quantitative trait loci (QTLs) affecting heading date were found in locations that are consistent with known QTLs and candidate genes, and two other QTLs affecting heading date were found in novel locations. Five QTLs affecting plant height were found. Both sets of QTLs are responsible for transgressive segregation observed for these two traits. Four QTLs affecting resistance to crown rust, caused by the pathogen Puccinia coronata f. sp. avenae, were identified. Two of these QTLs are consistent with known clusters of rust resistance genes, while two may represent new locations of novel rust resistance genes. A complete set of SNP sequences suitable for generating markers for molecular selection is provided.     

Genomic prediction of sunflower hybrid performance

Predicting single‐cross performance is of high importance to improve the efficiency of sunflower (Helianthus annuus L.) hybrid breeding programmes. We used experimental data from inter‐ and intragroup sunflower hybrids and their parental lines adapted to Central Europe to (i) study the genetic diversity and combining ability and (ii) examine the accuracy to predict hybrid performance based on phenotypic and genomic data. We evaluated 133 intragroup and 104 intergroup hybrids with their parental lines in replicated trials at four environments for grain yield, oil yield and oil content. Furthermore, the parental lines were fingerprinted with 572 AFLP markers. Variance due to specific combining ability was comparable for intergroup and intragroup crosses. This suggested a lack of clearly defined heterotic groups for the sample of studied sunflower lines. Prediction accuracy of hybrid performance based on general combining ability effects was high and could not be increased using genomic selection approaches. For situations where no information on GCA effects of parental lines was available, hybrid prediction based on genomic selection methods was accurate for groups of related lines. For groups of unrelated lines, however, we observed a strong decrease in the prediction accuracy. This suggests that prediction of hybrid performance for crosses based on genetically distant parents remains challenging.     

Development of a SCAR marker linked with a MYMV resistance gene in mungbean (Vigna radiata L. Wilczek)

Yellow mosaic disease (YMD) caused by mungbean yellow mosaic virus (MYMV) is the most important disease of mungbean, causing great yield loss. The present investigation was carried out to study the inheritance and identify molecular markers linked with MYMV resistance gene by using F₁, F₂ and 167 F_2 : 8 recombinant inbred lines (RILs) developed from the cross ‘TM‐99‐37’ (resistant) × Mulmarada (susceptible). The F₁ was susceptible, F₂ segregated in 3S:1R phenotypic ratio and RILs segregated in 1S:1R ratio in the field screening indicating that the MYMV resistance gene is governed by a single recessive gene. Of the 140 RAPD primers, 45 primers showing polymorphism in parents were screened using bulked segregant analysis. Three primers amplified specific polymorphic fragments viz. OPB‐07_600, OPC‐06₁₇₅₀ and OPB‐12₈₂₀. The marker OPB‐07₆₀₀ was more closely linked (6.8 cM) with a MYMV resistance gene as compared to OPC‐06₁₇₅₀ (22.8 cM) and OPB‐12₈₂₀ (25.2 cM). The resistance‐specific fragment OPB‐07₆₀₀ was cloned, sequenced and converted into a sequence‐characterized amplified region (SCAR) marker and validated in twenty genotypes with different genetic backgrounds.     

Stripe rust resistance gene Yr18 and its suppressor gene in Chinese wheat landraces

The slow‐rusting and mildewing gene Yr18/Lr34/Pm38/Sr57 confers partial, durable resistance to multiple fungal pathogens and has its origins in China. A number of diagnostic markers were developed for this gene based on the gene sequence, but these markers do not always predict the presence of the resistant phenotype as some wheat varieties with the gene are susceptible to stripe rust in China. We hypothesized that these varieties have a suppressor of Yr18. This study was undertaken to determine the presence of Yr18, the suppressor and/or another resistance gene in 144 Chinese wheat landraces using molecular markers and stripe rust field data. Forty‐three landraces were predicted to have Yr18 based on the presence of the markers, but had final disease severities higher than 70%, indicating that this gene may be under the influence of a suppressor. Four of these landraces, ‘Sichuanyonggang 2’, ‘Baikemai’, ‘Youmai’ and ‘Zhangsihuang’, were chosen for genetic studies. Crosses were made between the lines and ‘Avocet S’, with further crosses of Sichuanyonggang 2 ×  ‘Huixianhong’ and Sichuanyonggang 2 ×  ‘Chinese Spring’. The F₁ plants of Sichuanyonggang 2/Chinese Spring was susceptible indicating the presence of a dominant suppressor gene. The results of genetic analyses of F_2:3 and BC₁F₂ families derived from these crosses indicated the presence of Yr18, a Yr18 suppressor and another additive resistance gene. The Yr18 region in Sichuanyonggang 2 was sequenced to ensure that it contained the functional allele. This is the first report of a suppressor of Yr18/Lr34/Pm38/Sr57 gene with respect to stripe rust response.     

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.     

Marker‐assisted breeding of Pi‐1 and Piz‐5 genes imparting resistance to rice blast in PRR78, restorer line of Pusa RH‐10 Basmati rice hybrid

Rice blast, caused by fungus Magnaporthe grisea, is a serious disease causing considerable economic damage worldwide. Best way to overcome disease is to breed for disease‐resistant cultivars/parental lines of hybrids. Pusa RH10, first aromatic, fine‐grain rice hybrid released and cultivated extensively in India. Hybrid and its parental lines, Pusa 6A and PRR78, are highly susceptible to blast. CO39 pyramid carrying two dominant, broad‐spectrum blast‐resistance genes, viz. Pi‐1 and Piz‐5, used as a donor parent to introgress these genes into PRR78 using marker‐assisted backcrossing (MABC). Microsatellite markers RM5926 and AP5659‐5 tightly linked to Pi‐1 and Piz‐5 genes, respectively, were used for foreground selection to derive introgression lines. Further, these lines were evaluated for agronomic performance, disease reaction and cooking quality traits along with PRR78. Most of the improved lines were on par with PRR78 for all traits evaluated except gelatinization temperature. Recurrent parent genome percentage (RPG) study also revealed similarity of these lines with PRR78. Hybrids derived using improved PRR78 lines were superior over Pusa RH10 in terms of yield.     

Phenotypic and marker‐assisted pyramiding of genes for resistance to zucchini yellow mosaic virus in oilseed pumpkin (Cucurbita pepo)

We report on the identification of phenotypic and molecular markers for genes introgressed into oilseed pumpkin Cucurbita pepo from C. moschata germplasm originating in Nigeria, Portugal and Puerto Rico, which provide resistance against zucchini yellow mosaic virus (ZYMV) and on pyramiding these genes for improved and long‐lasting field protection of oilseed pumpkins. One SCAR and two SSR markers have been found for three dominant resistance genes, Zym‐0, Zym‐1 and Zym‐2. Characteristic reactions to ZYMV inoculation of plants carrying the recessive genes for resistance zym‐4* and zym‐6 have been defined. Described are procedures and results of pyramiding various combinations of these genes in oilseed pumpkin using the three markers and the specific phenotypic reactions to infection of some of these genes. The putative combination of all six resistance genes in one genotype resulted in a resistance that appeared to be at least as strong as or even stronger than that of the resistance source germplasm in C. moschata.     

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.