烟草祖先种及重要野生种叶绿素酶基因CLH1的生物信息学分析

叶绿素酶(Chlorophyllase)是所有光合生物叶绿素代谢中最重要的酶之一。叶绿素酶催化叶绿素的水解,产生脱植基叶绿素和叶绿醇。以烟草祖先种和重要野生种为研究对象,以拟南芥CHLOROPHYLLASE 1 (CLH1)为探针,通过同源序列搜索,从数据库中获取普通烟草(Nicotiana tabacum)及祖先种、烟草属重要野生种中的CLH1基因及其蛋白序列,并通过生物信息学分析手段对其编码蛋白的理化特性、氨基酸序列特征、蛋白结构与功能、启动子序列及表达调控特征进行了分析,并以林烟草(Nicotiana sylvestris) Nsyl CLH1为代表,预测了该基因启动子的顺式作用元件,以渐狭叶烟草(Nicotiana attenuata) Natt CLH1为例,预测了该基因的编辑位点。结果表明,栽培烟草CLH1蛋白虽在理化特性上存在差异,但氨基酸序列分析表明这些序列高度相似;对林烟草Nsyl CLH1基因的启动子顺式作用元件分析显示,除光响应作用元件、抗病性作用元件、干旱和盐分等作用元件,还有与茉莉酸甲酯(Methyl jasmonate,Me JA)、赤霉素(Gibberellic acid,GA)、脱落酸(Abscisic acid,ABA)和水杨酸(Salicylic acid,SA)等激素响应相关的元件,表明Nsyl CLH1基因的表达水平受多种因素的影响。最后,基因编辑位点预测显示渐狭叶烟草Natt CLH1可能跨越外显子-外显子连接。为茄科烟草属植物CLH1基因的功能研究、基因编辑及品质育种等提供了理论基础。

Bioinformatics Analysis of Chlorophyllase Gene CLH1 in Tobacco Ancestor Species and Important Wild Species

Chlorophyllase is one of the most important enzymes in the chlorophyll metabolism of all photosynthetic organisms.Chlorophyllase catalyzes the hydrolysis of chlorophyll to produce chlorophyllin and phytol. In this work, we utilized tobacco ancestral species and important wild species as the research objective,by using Arabidopsis CHLOROPHYLLASE 1(CLH1) as a probe,and we obtained the CLH1 genes in common tobacco and other genes in ancestral species and important wild species from the database through homologous sequence search, and analyzed the physical and chemical properties of the encoded protein, amino acid sequence characteristics, protein structure and function, promoter sequence and expression regulation characteristics through bioinformatics analysis methods. It was represented by Nicotiana sylvestris Nsyl CLH1. The cis-acting elements of the Nsyl CLH1 promoter were predicted. Taking Nicotiana attenuata Natt CLH1 as an example,the editing site of the gene was predicted. The results showed that although different CLH1 proteins in common tobacco plants had differences in physical and chemical properties,amino acid sequence analysis showed that these sequences are highly similar;analysis of cis-acting elements in the promoter of Nsyl CLH1 gene shows that in addition to light-responsive elements,disease-resistance acting elements,drought and salt acting elements,as well as other elements related to hormone response such as methyl jasmonate(Me JA), gibberellin acid(GA), abscisic acid(ABA) and salicylic acid(SA), indicating that the expression level of Nsyl CLH1 was affected by many factors. Finally, Natt CLH1 gene editing site predictions showed that the sequence may span the exon-exon junction. This study provided a theoretical basis for the functional study, gene editing and quality breeding of the CLH1 gene of Nicotiana in the Solanaceae family.