文献类型：期刊文献

中文题名：日本落叶松纤维素合酶基因片段的克隆及单核苷酸多态性分析

英文题名：Isolation and Single Nucleotide Polymorphisms Analysis of Cellulose Synthase Gene Fragment in Larix kaempferi

作者：易敏[1] 张守攻[1] 谢允慧[1] 孙晓梅[1]

第一作者：易敏

机构：[1]林木遗传育种国家重点实验室,中国林业科学研究院林业研究所,北京100091

年份：2015

卷号：28

期号：3

起止页码：303-310

中文期刊名：林业科学研究

外文期刊名：Forest Research

收录：CSTPCD;;Scopus;北大核心:【北大核心2014】；CSCD:【CSCD2015_2016】；

基金：国家“973”计划项目(2012CB114506)

语种：中文

中文关键词：日本落叶松;纤维素合酶基因;基因克隆;单核苷酸多态性

外文关键词：Larix kaempferi; cellulose synthase; gene cloning; single nucleotide polymorphism

分类号：S791.223

摘要：【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。
Objective Cellulose synthase （CesA） plays a key role in the biosynthesis pathway of plant cellulose, which controls the quality and yield of wood fiber. This work aims at cloning the homologous genes cellulose-related from Larix kaempferi, and further analyzing the nucleotide diversity and linkage disequilibrium, which may provide important genetic foundation associated with LkCesA gene and gene-assisted breeding of new germplasms with desira- ble wood fiber traits in L. kaempferi. Method According to the expressed sequence tags （ESTs） of cellulose syn- thase from L. kaempferi transcriptome database, a cDNA fragment encoding CesA was isolated from L. gene-specific PCR amplification. The genomic sequences of LkCesA in 40 individuals were cloned and then the single nucleotide polymorphisms （SNPs） diversity and linkage disequilibrium of LkCesA were kaempferi by sequenced, analyzed using DnaSPS. 0 software. Result The CesA fragment was 1 209 bp in length with a partial open reading frame （ORF, 1 053 bp） which would be capable to encode a predicted protein of 350 AA. The sequence indicated that the deduced amino acids shared 95.4% identity with PtCesA2 from Pinus taeda. A total of 83 SNPs was detected, and the frequency and diversity of SNPs （~rT） were 1/21 bp and 0.006 05, respectively. There were 69 SNPs be- longing to transition type and 24 belonging to transversion type, including 19 common SNPs and 64 rare SNPs. In total, 54 SNPs were detected in the coding regions of LkCesA, among which 20 were synonymous mutation and 34 were missense mutation. The results of linkage disequilibrium analysis showed that the LD declined rapidly with the nucleotide length. Conclusion The results of this study indicate that LkCesA is a member of CesA gene family. The linkage disequilibrium declined rapidly within the gene regions of LkCesA suggests that the gene could be used as the candidate gene for LD mapping, which contribute to the directional cultivation and wood quality improvement in Japanese larch. In addition, many common SNPs were detected in LkCesA which could be used in further LD mapping.