S组山羊草属Psy1基因片段的克隆与序列分析

Cloning and Sequence Analysis of Phytoene Synthase 1 (Psy1) Gene Fragment in Aegilops Species of S Genome

类胡萝卜素含量是影响小麦面制品色泽的主要因素,而Psy1基因则是类胡萝卜素生物合成中的关键基因。为了了解小麦近缘种S组山羊草的Psy1基因多样性分布规律,以高大山羊草、双角山羊草及西尔斯山羊草为材料,应用同源基因克隆技术对其进行Psy1基因克隆。结果表明,利用分子标记YP7B-5检测获得的8个新Psy1-S1等位变异的部分序列覆盖了Psy1基因的第二外显子和第二内含子的全部以及第三外显子93.1%的区域。在与包括来自普通小麦的Psy1-B1d、栽培二粒小麦的Psy1-B1k以及拟斯卑尔脱山羊草的Psy1-S1a、Psy1-S1b、Psy1-S1c对应部分的序列综合分析发现一致性高达93.4%~98.5%,而且编码区序列差异均为同义突变,没有造成编码氨基酸的变化。而第二内含子序列则存在数个SNP及InDel,尤其是Psy1-S1h的161bp大片段插入和Psy1-S1i的177bp大片段缺失,导致与其他Psy1-B1/Psy1-S1等位变异产生较大的长度差异。分析表明,内含子的差异是造成Psy1基因多样性的主要原因。

Carotenoids content（YPC） is the main factor influencing wheat flour color. The phytoene synthase 1 （Psyl） gene encodes a key enzyme to determine the carotenoids content of endosperm in wheat. Aegilops species are important resources for wheat genetic improvement. In the present study, three Aegilops species including Ae. longissima （SS） , Ae. bicornis （SS） and Ae. searsii （SS） were used to clone Psyl genes in order to disclose their genetic diversity. Using homologous gene cloning ap- proach,the partialPsyl gene sequences of eight novel allelic variants at Psyl-S1 locus were obtained by the marker YP7B-5, covering the whole region of the second exon and the second intron and the 93.1% region of the third exon. High sequence similarity of 93.4% to 98.5% was shown between these eight Psyl-S1 alleles and other five Psyl-B1/Psyl-S1 alleles, including Psyl-Bld from common wheat, Psy1-Blk from T. dicoccum and Psyl-Sla, Psyl-Slb, Psyl-Slc from Ae. speltoides. Moreover,they showed higher similarity in the coding sequences with only a few synonymous SNPs. There were several SNPs and InDels in the second intron,whieh make a valuable contribution to the genetic diversity of thePsyJ gene,partieularly a 163 bp insertion from PsyJ-Slh and a 177 bp deletion from Psyl-Sli, which were mainly attributed to the length difference, compared with other Psyl-B1/Psyl-S1 alleles.