中间锦鸡儿肉桂酰辅酶A还原酶基因的克隆和功能鉴定

发布时间：2017-12-28 13:01

本文关键词：中间锦鸡儿肉桂酰辅酶A还原酶基因的克隆和功能鉴定　出处：《内蒙古农业大学》2017年硕士论文　论文类型：学位论文

【摘要】：木质素是一种芳香杂聚物,它是自然界中在总量上仅次于纤维素的天然有机高分子物质,已得到广泛的研究。本研究从中间锦鸡儿中克隆得到两个木质素合成途径上的肉桂酰辅酶A还原酶基因,分别命名为CiCCR2和CiCCR3,并构建了植物表达载体,转化野生型拟南芥并获得纯合体株系,具体结果如下:(1)中间锦鸡儿CiCCR2基因开放阅读框长为897bp,gDNA共有4个内含子,分别长131bp、1351bp、176bp、222bp;编码299个氨基酸,推测的分子量为33.696KD,等电点为5.11,平均疏水指数(GRAVY)为-0.288;CiCCR3基因开放阅读框为966bp,gDNA 有 5 个内含子,分别长 829bp、856bp、197bp、522bp、1191bp,编码 322 个氨基酸,推测的分子量为35.91KD,等电点为6.01,平均疏水指数为-0.173。(2)构建过表达载体p35S::CiCCR2和p35S::CiCCR3,并转化野生型拟南芥Columbia-0,分别得到纯合体8个株系、9个株系。(3)过表达CiCCR2和CiCCR3基因的拟南芥,在成熟时期和幼苗时期木质素含量均高于野生型拟南芥组织化学染色也验证了转基因株系木质素积累较野生型拟南芥多。并且CiCCR2和CiCCR3过表达植株的鲜重和干重比野生型增加。
[Abstract]:Lignin is a kind of aromatic heterozygous. It is a natural organic polymer in the natural world, second only to cellulose, and has been widely studied. The cloning of cinnamoyl coenzyme A reductase gene two lignin biosynthesis pathway from Caragana intermedia, named CiCCR2 and CiCCR3, and constructed the plant expression vector and transformed into wild type Arabidopsis and obtain homozygous lines, the specific results are as follows: (1) Caragana CiCCR2 gene open reading frame 897bp long, gDNA has 4 introns, respectively 131bp, 1351bp, 176bp, long 222bp; encoding 299 amino acids. The predicted molecular weight of 33.696KD and isoelectric point is 5.11, the average hydrophobicity index (GRAVY) for -0.288; on the open reading frame of 966bp CiCCR3 gene has 5 introns, gDNA respectively, 829bp, 856bp, long 197bp, 522bp, 1191bp, encoding 322 amino acids. The predicted molecular weight of 35.91KD and isoelectric point is 6.01, the average hydrophobicity index was -0.173. (2) the over expression vector p35S:: CiCCR2 and p35S:: CiCCR3 were constructed and transformed into wild Arabidopsis Columbia-0, and 8 strains of homozygous lines and 9 strains were obtained. (3) the overexpression of CiCCR2 and CiCCR3 genes in Arabidopsis thaliana showed higher lignin content in mature and seedling stage than in wild Arabidopsis thaliana. Histochemical staining also confirmed that the accumulation of lignin in transgenic plants was more than that in wild Arabidopsis thaliana. The fresh weight and dry weight of the overexpressed plants of CiCCR2 and CiCCR3 increased as compared with the wild type.
【学位授予单位】：内蒙古农业大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：Q943.2

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