‘宁杞1号’与‘宁杞3号’S-RNase基因的克隆与表达分析

Cloning and Expression Analysis of S-RNase Genes from ’Ningqi1’ and ’Ningqi3’

本研究通过‘宁杞1号’和‘宁杞3号’S-RNase基因全长序列的克隆和荧光定量表达分析,为分子手段调控宁夏枸杞自交不亲和性状提供了参考,为‘宁杞1号’自交亲和的分子机理研究以及自交亲和品种的选育提供科学依据。以‘宁杞1号’和‘宁杞3号’为试材,利用特异性PCR扩增技术,从花柱基因组DNA中克隆S-RNase基因全长,并进行生物信息学分析;运用实时荧光定量(qRT-PCR)技术分析S-RNase表达量的变化。克隆到S-RNase基因,暂分别命名为S-BARB2n和S-BARB8n,两者cDNA全长为672 bp,编码223个氨基酸,二者均属于RNase T2基因家族。预测2个S-RNase蛋白的二级结构均以无规则卷曲、α-螺旋和延伸链等组成。随着花柱的发育S-RNase表达量呈上升趋势。获得两个S-RNase基因的全长序列和不同发育阶段花柱S-RNase的表达量。

The full-length sequences of‘Ningqi1’and‘Ningqi3’S-RNase genes were cloned and analyzed by fluorescence quantitative exp ression,which provides a reference for the regulation of self-incompatibility of Lycium halimifolium and a scientific basis for the study of molecular mechanism of self-compatibility of‘Ningqi1’and the breeding of self-compatible varieties.‘Ningqi1’and‘Ningqi3’were used as test materials,and the whole length of S-RNase gene was cloned from the genomic DNA of the flower column by using the specific PCR amplification technique,and the bioinformatics analysis was carried out.The change of S-RNase expression was analyzed by real-time fluorescence quantitative technique.The S-RNase gene was cloned and tentatively named S-BARB2n and S-BARB8n,respectively.The total length of cDNA was 672 bp,which encoded 223 amino acids,both of which belonged to the RNase T2 gene family.It is predicted that the secondary structure of the two S-RNase proteins is composed of random coil,α-helix and extended strand.With the development of style,the expression of S-RNase increased.Finally,the full-length sequences of two S-RNase genes and the expression of style S-RNase at different developmental stages were obtained.