A wheat (Triticum aestivum L.) immature spike culture system was used to expeditiously generate mutations for use in wheat improvement programs. Wheat immature spikes in culture were treated with three concentrations of ethylmethane sulphonate (EMS) to generate a spike culture derived variant (SCDV) population. EMS in a concentration dependent manner affected seed development in wheat immature spike cultures. Based on the number of seeds produced, inclusion of EMS (25 mM) for three hours in immature spike culture medium generated variants in wheat cv. AC Nanda. The wheat AC Nanda SCDV population showed variation in several phenotypic characters. Flag leaf (length, angle and sheath length), length of first and second internode, spike length, number of spikes, number of seeds per spike and seed weight, showed variation below and above the non-treated controls. A molecular screening technique combining simple sequence repeat (SSR) oligonucleotide primers with high resolution melt (HRM) PCR with EvaGreen was used to identify the variants. Screening for starch branching enzyme IIb (SbeIIb) revealed 75 lines with point mutations. Combining SSR and SbeIIb, a total of 100 Kbp portion of wheat DNA was screened. The estimated mutation frequency in SbeIIb was one per 20.8 Kbp. The spike culture system utilizes very small amounts of EMS for a brief period, thus needs minimal handling of EMS and saves one generation of plant growth in a greenhouse. The morphological variants observed are similar to those reported for seed-derived variants using EMS.
Authors:R. Singh, P. Hucl, M. Båga, and R. Chibbar
Pre-harvest sprouting (PHS) in bread wheat (Triticum aestivum L.) is one of the major abiotic constraints influencing production of high quality grain. Selection for pre-harvest sprouting (PHS) resistance in bread wheat (Triticum aestivum L.) in early generations is difficult because it is expressed as a quantitatively inherited trait and subject to environmental effects. The objectives of this study were to validate a major quantitative trait locus (QTL) for PHS resistance on chromosome 4A in bread wheat and to isolate near-isogenic lines for this QTL using marker-assisted selection. A total of 60 Canadian wheat cultivars and experimental lines were screened with three SSR markers in a QTL region for PHS resistance. The SSR markers DuPw004, barc170 and wmc650 explained 67%, 75% and 60% of total variation in germination (%), respectively, among different wheat genotypes. Marker assisted back crossing with DuPw004 reduced the population size in BC1F1 and BC2F1 generation by 41% and 59%, respectively. A survey of pedigrees of different genotypes revealed that the parental line RL4137 is a major source of increased PHS resistance in a number of western Canadian wheat cultivars. Microsatellite markers (DuPw004, barc170 and wmc650) will be useful for plant breeders to pyramid QTL from different PHS resistance sources.