【目的】纤维素合酶(cellulose synthase,Ces A)在植物纤维素合成途径中发挥主要调节作用,是控制木材纤维品质和产量的重要基因。从日本落叶松中分离克隆与纤维素合成相关的Lk Ces A基因,并对其进行核苷酸多样性以及连锁不平衡分析,为在日本落叶松中开展基于Lk Ces A基因的连锁不平衡作图及其辅助日本落叶松木材纤维性状的分子育种提供理论依据。【方法】依据日本落叶松转录组数据库检测到的纤维素合酶(Ces A)基因ESTs序列设计引物,从日本落叶松中分离获得Lk Ces A基因片段。在此基础上,利用Dna SP5.0软件对日本落叶松40株基因型个体的Lk Ces A序列进行核苷酸多样性和连锁不平衡分析。【结果】从日本落叶松中成功克隆了Ces A基因片段:该片段长1 209 bp,包含部分开放阅读框,长度为1 053 bp,可编码350个氨基酸,所推导的蛋白质氨基酸序列与火炬松PtCes A2的蛋白质氨基酸序列同源性为95.4%。在日本落叶松40株基因型个体的Lk Ces A序列中共检测到83个SNP位点,SNP发生频率为1/21 bp,多样性指数πT为0.006 05。在这些SNPs中,69个属于转换,14个属于颠换,其中19个为常见SNPs,64个为罕见SNPs。在外显子区域,共检测到54个SNP位点,其中34个为错义突变,20个为同义突变。进一步的连锁不平衡分析显示,随着核苷酸序列长度的增加,SNP连锁不平衡程度逐渐减弱。【结论】克隆到的Lk Ces A为植物Ces A基因家族中的一员。Lk Ces A基因的连锁不平衡在基因内部就已衰退,说明选择该基因作为候选基因,在日本落叶松中开展连锁不平衡作图用于指导日本落叶松的定向培育及木材品质改良是可行的。此外,在Lk Ces A基因中检测到多个常见SNP位点,为进一步开展该基因的连锁不平衡作图提供了材料。
Molecular biological research into wood development and formation has been the focus in recent years, but the pace of discovery of related genes and their functions in the control of wood properties has been slow. The microarray technique--with its advantages of high throughput capacity, sensitivity, and reliability over other tools developed for investigating genes expression patterns-is capable of rapidly assaying thousands of genes. In this study, a cDNA microarray prepared from two cDNA libraries of developing poplar xylem tissues was used to assay gene expression patterns in immature xylem tissues at different heights from the main stem of Populus deltoides （15 years old）, which was confirmed to have distinct wood properties （microfibrillar angle, woody density） by X-ray. Two hundred seventy-four transcripts with differ- ential expression profles between the chips were screened out, and the individual clones were subjected to 5＇ sequencing. Using bioinformatic analysis, we identified candidate genes that may influence poplar wood properties, many of which belong to various regulatory and signal transduction gene families, such as zinc finger protein transcription factor, DNA-binding transcription factor, ethylene response factors, and so on. The results suggest that these genes may regulate enzymes involved in wood formation. Further work will be performed to clone these genes and determine how they influence poplar wood properties.
Qinjun HuangChangjun DingWeixi ZhangBingyu ZhangYanguang ChuDahai WangXiaohua Su
