【作者】 陈其皎；

【导师】 刘登才；

【作者基本信息】 四川农业大学 ， 生物化学与分子生物学， 2006， 硕士

【摘要】 黑麦属(Secale L．，染色体组RR，2n=14)属于小麦族(Triticeae)，是小麦(Triticum aestivum L．，染色体组AABBDD，2n=42)的近缘属植物。因其蕴藏着丰富的抗逆性(抗旱、耐寒、耐贫瘠)、抗多种病虫害、多分蘖、多小穗等丰产性和高赖氨酸含量等突出优点而作为普通小麦重要的三级基因资源，特别是黑麦的1RS臂以1BL／1RS易位的方式已在普通小麦产量、抗病、抗逆等改良中发挥了重要作用。研究表明，黑麦的1RS、1RL、2RS染色体臂上有控制蛋白质品质的相关基因。其中1RS上的secalin基因对小麦的烘烤品质有负面作用。尽管2RS上的75K γ-secalin基因的编码产物占黑麦种子储藏蛋白的一半以上，但是对该基因的遗传信息还缺乏了解。 本文用3份不同来源地的栽培黑麦材料，通过SDS-PAGE确认了75K γ-secalin亚基的差异，进一步从DNA序列上分析了其遗传差异，并根据DNA序列信息找到了75K γ-secalin基因的分子标记。其主要结果如下： 1．75K γ-secalin亚基的分离与克隆 根据GenBank中登录的75K γ-secalin编码基因的序列特征，设计了针对该基因完整编码区的特异引物，对3份材料(Clse 20，Clse 35，PI168199)进行PCR扩增，得到大小分别为1.2kb、1.2kb和1.0kb左右的1条特异扩增条带，与引物设计时预期的75K γ-secalin亚基白基因编码区的长度接近。阳性克隆分别命名为pSec-1、pSec-2和pSec-3，经测序后获得的实际编码区长度分别为1194bp、1230bp和1017bp，DNAMAN推导的编码产物分别为包含了397个、409个和338个氨基酸残基的成熟表达蛋白。 2．75K γ-secalin亚基编码序列的信息发掘 使用NCBI提供的BLAST在线分析软件比对发现，pSec-1、pSec-2和pSec-3的氨基酸序列与已报道的75K γ-secalin亚基的编码产物具有较高一致性。使用DNAMAN、SignalP等软件对pSec-1、pSec-2和pSec-3编码产物进行理化性质(保守的信号肽序列、典型氨基酸模体等参数)预测，发现与75K γ-secalin亚基一致。更多还原

【Abstract】 Secale L, one of the relative genera of Triticum, has been playing an important tertiary gene pool in wheat breeding for its superior traits in many fields, such as multiple tillers, supernumerary spikelets, high content of lysine and good tolerance to diseases, pests, drought and cold environment. The wheat-rye 1BL/1RS translocation lines, derived from the introgression of rye 1RS into wheat 1BL, have made great contributions in wheat breeding.The rye chromosome arms of 1RS, 1RL and 2RS carry some genes which control the quality of storage protein. And the coding products of secalin genes in 1RS have negative effects on the baking quality of dough. The 75K 7-secalin, coded by the corresponded genes on 2RS, accounts for half contents of the total seed protein, however we still are lack of awareness of both the features of the coding genes and the role of its expression products.In order to investigate the genetic diversity and analysis the sequence features of the 75K 7-secalin gene family, three ryes (Secale cereale L.) were studied by SDS-PAGE and molecular cloning. The results are as following: 1. Cloning and sequencing of the coding sequences of 75K γ-secalinWe designed a pair of special primers based on the known sequence of 75K 7-secalin in GenBank. The rye cultivars Clse 20, Clse 35 and PI 168199 were respectively amplified a 1.1 kb c.a. product using this primer pairs. Cloned these fragments to T-Vector and the positive clones of Clse 20, Clse 35 and PI168199, were named as pSec-1, pSec-2 and pSec-3, respectively. The sequencing results showed that the real length of the three clones were 1194 bp, 1230 bp and 1017 bp, respectively. The predicted amino-acid sequences were 397 aa, 409 aa and 338 aa, deduced by DNAMAN software. 2. Genetic information of the 75K γ-secalin更多还原