Distribution Of Dwarfing And Vernalization Genes In Chinese Wheats And Development Of Multiplex-PCR For Improving Quality Properties

The use of dwarfing genes to reduce plant, increasing harvest index, improve lodging resistance, and increased grain yield has been the major strategy in developing modern bread wheat cultivars. The adaptability of wheat cultivars to environmental conditions is highly associated with their vernalization requirement which is mainly controlled by three major genes, Vrn-A1, Vrn-B1 and Vrn-D1. The vernalization genes determine growth habit (winter vs. spring) of wheat cultivars. Quality improvement has become an important objective for wheat breeding and production in China. Wheat quality properties are strongly affected by the compositions of high-and low-molecular-weight glutenin subunits, presence or absence of 1B/1R translocation, amylase content, polyphenol oxidase (PPO) activity, kernel hardness and pre-harvest sprouting tolerance. Knowledge on the distribution of dwarfing genes and vernalization genes in Chinese wheat will be very important for understanding adaptability of wheat to environmental conditions. It is very useful to develop multiplex PCR for wheat quality improvement in molecular marker assisted breeding. In this study, molecular markers were used to detect the presence of Rht-Blb (Rht1), Rht-Dlb (Rht2) and Rht8 genes in 263 Chinese wheats. Allelic variations at Vrn-A1, Vrn-B1, Vrn-D1 and Vrn-B4 loci in 278 leading cultivars from eight main China wheat zones were detected by molecular markers, and growth habit of 266 cultivars was also identified in greenhouse condition. In addition, five types of multiplex PCRs were developed, which were validated with many wheat cultivars and advanced lines with known genotypes.1. The results indicated that the PCR-based markers could be used to test the presence of Rht-Blb, Rht-Dlb and Rht8 in Chines wheat cultivars and advanced lines. Rht-Dlb and Rht8 genes take a dominant position with frequencies of 42.6% and 41.8%, respectively, followed by Rht-Blb with a frequency of 23.6%. Frequencies of thses three dwarfing genes among various regions were different. In four main autumn-sown wheat zonesⅠ(Northern Winter Wheat Zone),Ⅱ(Yellow & Huai River Facultative Winter Wheat Zone),Ⅲ(Middle & Low Yangtze Valley Winter Wheat Zone) andⅣ(Southwestern Winter Wheat Zone), average frequencies of Rht-Blb and Rht-Dlb have increased, from 8.6 to 32.2%, 36.2 to 53.4%, respectively, while frequency of Rht8 remains constant over time, when compared varieties released before and after 1990. The different distribution of three dwarfing genes in Chinese cultivars and advanced lines from various wheat zones were largely determined by their genetic background and utilization of dwarfing genes in wheat breeding. Rht-Blb in Chinese wheats has two sources, i.e., Norin 10 and Italian introduction St2422/464. Suwon 86, Norin 10 and two Chinese varieties Huixianhong and Yaobaomai are major original source of Rht-Dlb in Chinese wheats. Introductions such as Funo, Abbondanza, Villa Glori, St1472/506, St2422/464, Bezostaya 1 and Lovrin are major sources of Rht8 gene.2. The results indicated that the PCR-based markers could be used to detect allelic variations of Vrn-A1, Vrn-B1, Vrn-D1 and Vrn-B4 loci in Chines wheat cultivars. Averaged frequency of dominant gene Vrn-D1 in Chinese wheat was the highest (37.8%), followed by dominant genes Vrn-A1 (27.3%) and Vrn-B1 (26.3%), and dominant gene Vrn-B4 was the lowest (0.7%). Frequencies of spring and winter cultivars in tested genotypes were 65.4% and 34.6%, respectively. Spring cultivars were mostly presented in ZonesⅢ(Middle & Low Yangtze Valley Winter Wheat Zone),Ⅳ(Southwestern Winter Wheat Zone),Ⅵ(Northeastern Spring-sown Spring Wheat Zone),Ⅶ(Northern Spring-sown Spring Wheat Zone),Ⅷ(Northwestem Spring-sown Spring Wheat Zone) andⅩ(Xinjiang Winter & Spring Wheat Zone), while winter cultivars were presented inⅠ(Northem Winter Wheat Zone),Ⅱ(Yellow & Huai River Facultative Winter Wheat Zone) andⅩ(Xinjiang Winter & Spring Wheat Zone). Winter cultivars identified in the greenhouse possessed all recessive alleles at the four vernalization loci detected by PCR, and cultivars with at least one dominant gene identified by PCR were spring. However, distributions of the four dominant vernalization genes and growth habits in various wheat zones were different. All cultivars released in ZoneⅠ(Northern Winter Wheat Zone) were winter and carried recessive alleles at the four vernalization loci. Both winter (57.5%) and spring (42.5%) cultivars were present in ZoneⅡ(Yellow & Huai River Facultative Winter Wheat Zone) and most spring cultivars identified in the greenhouse carried single dominant vernalization gene either Vrn-B1 or Vrn-D1, with frequencies of 6.5% or 64.5%. In ZonesⅢ(Middle & Low Yangtze Valley Winter Wheat Zone) andⅣ(Southwestern Winter Wheat Zone), spring cultivars were predominant, and dominant gene Vrn-D1 was the most frequent. In three spring-sown ZonesⅥ(Northeastern Spring-sown Spring Wheat Zone),Ⅶ(Northern Spring-sown Spring Wheat Zone) andⅧ(Northwestern Spring-sown Spring Wheat Zone), all cultivars were spring and frequently possessed the strongest dominant vernalization gene Vrn-A1, along with another dominant gene(s). Both winter and spring cultivars were present in ZoneⅩ(Xinjiang Winter & Spring Wheat Zone) with autumn and spring-sown. Composition of vernalization genotypes with spring type in ZoneⅩ(Xinjiang Winter & Spring Wheat Zone) was very similar to that in spring-sown zones. Growth habits and vernalization genotypes of wheat cultivars released in same wheat zone were also determined by their parents.3. The five multiplex PCR types were developed. The first multiplex PCR was used to detect genesω-secalin (1B/1R), VplB3 and Pinb-Dlb, simultaneously, for improving wheat processing quality. The second one included molecular markers for six genesω-secalin, By8, Glu-A3d, Dx5 Pinb-Dlb and Glu-B3. The third multiplex PCR was used to detect the genesω-secalin, Ax2~*, Bx17 and Dx5, simultaneously. The second and third multiplex PCRs were to improve gluten quality. The fourth comprised functional markers for PPO-2A, PPO-2D and Wx-B1 loci for improving noodle quality, and the fifth included three markers for Wx-7A, Wx-4A and Wx-7D for breeding waxy wheat cultivars. The five multiplex PCRs were highly effective in the test of Chinese wheat cultivars and advanced lines, demonstrating good repeatability and low cost for the evaluation of wheat quality properties in wheat breeding program.It is the first to survey extensively three dwarfing genotpes and four vernalization genotypes in Chinese wheats from main eight regions, and develop five highly effective multiplex PCR types for evaluation of wheat quality characters. These information are very useful to breed and extend wheat cultivars with high-yield, good quality and wide adaptability in China.