| Flour color exerts a significant effect on wheat quality, which is controlled by Yellowpigment(YP) content, Lipoxygenase(LOX) and Polyphenol oxidase(PPO) activity. It isimportant to develop multiplex PCR systems as effective and economical methods for theidentification and selection of favorable alleles related to flour color in MAS, provideinformation on allelic compositions in wheat cultivars, to facilitate breeders, both in Chinaand abroad, in designing crosses aimed at improving wheat color quality, as well as to helpChinese farmers to choose elite varieties for cultivation. This study was classified into twoparts.(1) the alleles at nine loci related to flour color in176Chinese autumn-sown cultivarswere genotyped by twelve functional markers.(2)20cultivars and lines were used as controlsto established three multiplex PCR systems which covered seven loci related to flour color,and the multiplex PCR developed in this study were validated by106cultivars, then49Chinese autumn-sown cultivars were genotyped by the newly developed multiplex PCR.The results of single PCR amplification by twelve markers showed that (1) at Psy-A1locus, the frequencies of alleles Psy-A1a and Psy-A1b were44.9%and54.0%, respectively,and two cultivars without any fragment may carry the novel allele, accounting for1.1%of thetotal,(2) at Psy-B1locus, the frequencies of alleles Psy-B1a, Psy-B1b and Psy-B1c, were26.7%,65.3%and8.0%, respectively,(3) at Psy1-D1locus, the frequencies of allelesPsy1-D1a and a novel allele, were99.4%and0.6%, respectively,(4) at TaZds-A1locus, thefrequencies of alleles TaZds-A1a and TaZds-A1b were42.6%and57.4%respectively,(5) atTaZds-D1locus, only TaZds-D1a were found in all the tested cultivars and lines,(6) atQLpx.caas-1AL locus, the alleles Xwmc312-227, Xwmc312-235and Xwmc312-247were found inthe cultivars and lines, accounting for31.8%,18.2%and50.0%, respectively,(7) at TaLox-B1locus, the alleles TaLox-B1a and TaLox-B1b were detected, respectively taking up thefrequencies of29.0%and71.0%,(8) at Ppo-A1locus, the alleles Ppo-A1a and Ppo-A1b wererespectively found in47.2%and52.8%cultivars and lines,(9) at Ppo-D1locus, the frequencies of alleles Ppo-D1a and Ppo-D1b were36.4%and63.6%, respectively.In this study, three multiplex PCR systems including seven target loci were developedfor genotyping15alleles related to flour color. Multiplex PCR-I were developed byfunctional markers YP7B-1, YP7B-2and YP2A-1, which was able to identify five allelesPsy-B1a, Psy-B1b, Psy-B1c, TaZds-A1a and TaZds-A1b at the Psy-B1and TaZds-A1loci.Multiplex PCR-II were developed by YP2D-1and PPO18. It can investigate four allelesPpo-A1a, Ppo-A1b, TaZds-D1a and TaZds-D1b at two loci, Ppo-A1and TaZds-D1, in onePCR amplification. Multiplex PCR-III were established by YP7A, LOX18and STS01, whichcan genotype six alleles Psy-A1a, Psy-A1b, TaLox-B1a, TaLox-B1b, Ppo-D1a, and Ppo-D1bat three loci, Psy-A1, TaLox-B1and Ppo-D1, simultaneously. The results of these multiplexPCRs in20controls and106validated cultivars completely matched with those of eight singlePCR amplifications, showing that the multiplex PCR systems were accurate and reliable.Then the allelic combinations in other49Chinese autumn-sown cultivars were investigatedby the newly developed multiplex PCRs. So the information of altogether155cultivars atseven loci related to flour color were investigated. It can be seen that the frequencies of mostof the alleles related to lower YP content and higher LOX activity (Psy-A1b, Psy-B1b,TaZds-D1b, TaLox-B1a) were higher in new cultivars than those from earlier cultivars, and tenof elevenths allelic combinations related to lower YP content and higher LOX activity(TaZds-D1b was not considered because of the overly small proportion) in new cultivars werehigher than those from earlier cultivars.The identification and selection were first covered nearly all the known loci related toflour color in this study, providing cultivars and lines with ideal alleles at multiplex loci inMAS to meet flour color requirements. Three multiplex PCR systems developed providing aneffective tool for screening the cultivars with target alleles, which were first included YPcontent genes (Psy-1and TaZds-1), LOX and PPO genes, simultaneously. |
