文献类型：期刊文献

中文题名：梨花粉自交不亲和基因克隆及其评价分析

英文题名：Cloning and evaluation of S locus F-box brother genes in genus Pyrus plants

作者：梁文杰[1] 乌云塔娜[2,3] 谭晓风[3]

第一作者：梁文杰

机构：[1]温州科技职业学院,浙江温州325006;[2]中国林业科学研究院经济林研究所,郑州450003;[3]中南林业科技大学经济林育种与栽培国家林业和草原局重点实验室,长沙410004

年份：2022

卷号：39

期号：6

起止页码：905-919

中文期刊名：果树学报

外文期刊名：Journal of Fruit Science

收录：CSTPCD;;Scopus;北大核心:【北大核心2020】；CSCD:【CSCD2021_2022】；

基金：国家自然科学基金项目(31000309)。

语种：中文

中文关键词：梨;自交不亲和性;SFBB基因;共同进化

外文关键词：Pear;Self-incompatibility;S-locus F-box brother genes;Co-evolution

分类号：S661.2

摘要：【目的】获得梨SFBB新基因全长序列,研究不同SFBB基因参与梨自交不亲和的作用。【方法】设计特异性引物扩增梨SFBB22-gamma基因组DNA全长序列,并对所有已鉴定、能够与梨S-RNase基因一一对应的SFBB基因全长序列进行比较分析。【结果】梨SFBB22-gamma基因编码区序列大小为1191 bp,编码396个氨基酸,ProtParam预测分子质量为45.47 ku,理论等电点4.67,为酸性亲水不稳定蛋白。其中N端包含由50个氨基酸组成的F-box序列,蛋白质二级结构有5个α-螺旋和24个β-折叠。不同梨SFBB基因和对应S-RNase基因在序列变异、基因多态性以及进化特性等方面存在差异。【结论】现有序列数据显示,梨SFBB基因的序列多态性和变异位点均小于S-RNase。梨SFBB基因4个群体中单个群体SFBB-beta基因和对应S-RNase基因在序列特征上最接近;4个梨SFBB基因个体作为整体与对应S-RNase基因序列特征比SFBB-beta基因和对应S-RNase基因序列特征更接近。SFBB-beta基因存在独自参与自交不亲和的可能性,SFBB-alpha以及SFBB-gamma基因不存在独自参与自交不亲和的可能性。
【Objective】In order to avoid self-pollination in the evolution process of Rosaceae,self-incompatibility is common.Studies have shown that the self-incompatibility of Rosaceae Maloideae Pyrus is achieved by the interaction of the pistil S-RNase gene and the pollen SFBB gene.In order to obtain the full-length sequence of the new pear SFBB gene,the situation of different pear SFBB genes involved in self-incompatibility was clarified,and the SFBB gene type that was more compatible with pear S-RNase gene was found.【Methods】DNA was extracted from the leaves of Yunnan Baozhu pear,Fu’an Jianba pear,and Pitai pear varieties,and specific primers were designed based on the conserved characteristics of the SFBB-gamma gene sequence of the Maloideae Pyrus,and the pear SFBB22-gamma genomic DNA Full-length sequence was amplified by PCR.The full-length sequences of all the isolated and identified SFBB-alpha,SFBB-beta,SFBB-gamma and SFBB-epsilon genes that can correspond one-to-one with the pear S-RNase gene were compared and analyzed.The two are respectively based on the physical and chemical properties of the gene,the proportion of variant sites,the proportion of different nucleotide compositions,the ratio of conversion and transversion,the pairwise genetic distance of the coding region sequence and the genetic diversity index,Tajima’s Neutrality Test D value,positive position The proportion of points,the proportion of neutral loci and the number of consecutive positive loci were compared and analyzed.【Results】The size of the coding region of pear SFBB22-gamma gene was 1191 bp,encoding 396 amino acids;the predicted molecular weight of ProtParam was 45.47 ku;the theoretical isoelectric point was 4.67 and thus it might be an acid-hydrophilic unstable protein.The N-terminus contained an F-box sequence consisting of 50 amino acids.The protein secondary structure had 5α-helices and 24β-sheets.Theα-helix was mainly distributed in the F-box region,and the β-sheet was scattered throughout the protein.At present,a total of 52 full-length CDS sequences of SFBB gene corresponding to pear S-RNase gene have been registered in the NCBI database,including 15 SFBB-alpha,10 SFBB-beta,24 SFBB-gamma,and 3 SFBB-epsilon,corresponding to 30 full-length CDS sequences of pear S-RNase genes.Different SFBB gene in NJ phylogenetic trees were clustered into 4 subgroups,and there was no clustering with species as the boundary;the corresponding pear SRNase gene did not have obvious clustering with species as the unit either.The prediction of protein physicochemical properties showed that the proteins encoded by the pear SFBB-alpha,SFBB-beta,SFBB-gamma and SFBB-epsilon genes were all acidic proteins,and the basic proteins encoded by the pear S-RNase gene were theoretically attracted by positive and negative charges.At the same time,the proteins encoded by both were hydrophilic proteins with similar thermal stability.The molecular weight of the protein encoded by SFBB was about 1.75 times that of the protein encoded by the corresponding S-RNase gene.There was a theoretical possibility of interaction.The SFBB-alpha,SFBB-beta,SFBB-gamma and SFBB-epsilon genes of pear plants were highly conserved.Further research and analysis showed that the pear SFBB-alpha,SFBB-gamma and SFBB-epsilon genes had the same correlation with the corresponding S-RNase.There was less evidence of gene co-evolution,and these SFBB genes exhibited lower sequence variation,less positive selection,and lower gene polymorphisms.The SFBB-beta gene showed a higher level of sequence diversity,although it was still lower than that shown by the corresponding S-RNase gene.If the four subtypes of pear SFBB gene are considered as a whole,the above indicators and corresponding S-RNase conditions are better than those of SFBB-alpha,SFBB-gamma,SFBB-epsilon and SFBB-beta genes.【Conclusion】The design of specific primers can directly amplify the full-length sequence of pear SFBB gene.At present,this technology still has the defect of poor primer versatility and needs to be further improved.Finding a method comparable to amplifying the full-length sequence of the pear S-RNase gene is the key to the study of the pear SFBB gene.The existing SFBB gene full-length sequence family that can correspond one-to-one with the pear S-RNase gene has 4 members,and the sequences between each member are relatively conservative.All are smaller than S-RNase.Existing data show that pear SFBB-beta gene may participate in self-incompatibility alone,and pear SFBB-alpha and SFBB-gamma genes may not participate in self-incompatibility alone.From a theoretical point of view,whether all the four SFBB gene subtypes are involved in self-incompatibility or whether SFBB-beta alone is involved in self-incompatibility needs more research to verify.