摘要
ANS为花青素植物花青素合成途径末端的关键酶基因。本研究从东方百合中克隆了3个ANS同源基因,并通过碱基和氨基酸的差异分析了该基因的转录水平和表型效应。为进一步研究百合ANS的特性,完善百合花色形成的分子机制提供理论依据。选取东方百合‘西伯利亚’品种为材料,通过同源克隆技术鉴定了LsANS1、LsANS2、LsANS3,并分别构建其过表达载体;利用花序侵染法转化拟南芥,经潮霉素抗性筛选及分子检测以获得阳性植株;利用电子解剖镜观察和RT-qPCR定量分析表达量差异,验证野生型与转基因植株后代的表型差异水平。结果显示,LsANS1与LsANS3高度同源,转基因阳性植株主茎明显为红色,可能是吗啡合成N-端的非血红素加氧酶(non-haem dioxygenase in morphine synthesis N-terminal,DIOX_(N))亚家族结构域上的个别碱基的变异导致基因表达水平变化从而引起表型差异;LsANS2与其他两个同源基因存在氨基酸序列上的差异,且基因表达水平也远远低于其他两个同源基因,在转基因植株中呈粉红色。
ANS is a significant enzyme gene at the end of the anthocyanin synthesis pathway in plant.The study clone three ANS homologous genes from Oriental lily,and analysis the gene transcription level and phenotypic effects by the difference of base and amino acid.The further study for the features of the ANS in lily,and improve the molecular mechanism of lily flower color formation theory basis.Using Lilium orientalis ’Siberia’ as material,LsANS1,LsANS2 and LsANS3 were identified by homologous cloning,and their overexpression vectors were constructed.Arabidopsis thaliana was transformed by inflorescence infection,and positive plants were obtained by hygromycin resistance screening and molecular detection.The phenotypic level of wild-type and transgenic plant progenies was verified by quantitative analysis of expression level by electron anatomical microscopy and RT-qPCR.The results showed that LsANS1 and LsANS3 were highly homologous,and the main stem of the transgenic positive plants was obviously red,which might be caused by the variation of individual bases in the non-haem dioxygenase in morphine synthesis N-terminal(DIOX_(N)),resulting in the change of gene expression level and the phenotypic difference.LsANS2 was different from the other two homologous genes in amino acid sequence,and the gene expression level was much lower than the other two homologous genes,showing pink in transgenic plants.
作者
杨成龙
张洁
方少忠
郑益平
林智敏
Yang Chenglong;Zhang Jie;Fang Shaozhong;Zheng Yiping;Lin Zhimin(Institute of Biotechnology,Fujian Academy of Agricultural Sciences,Fuzhou,350003)
出处
《分子植物育种》
CAS
北大核心
2021年第20期6741-6746,共6页
Molecular Plant Breeding
基金
福建省属公益类科研院所基本科研专项(2020R10270011)资助。
