枇杷属植物3个新S基因鉴定及大渡河枇杷分类地位的新证据  

Identification of three novel S-RNase alleles in Eriobotrya Lindl. and new evidence for the taxonomic status of E. prinoides var. dadunensis

作　　者：付燕[1] 杨芩 彭舒 蒋瑶 刘伦沛 FU Yan;YANG Qin;PENG Shu;JIANG Yao;LIU Lunpei(Qiandongnan Vocational and Technical College,Kaili 556000,Guizhou,China;College of Life and Health Science,Kaili University,Kaili 556000,Guizhou,China)

机构地区：[1]黔东南民族职业技术学院,贵州凯里556000 [2]凯里学院大健康学院,贵州凯里556000

出　　处：《果树学报》2018年第11期1316-1323,共8页Journal of Fruit Science

基　　金：国家自然科学基金地区项目(31460510)

摘　　要：【目的】鉴定枇杷属普通枇杷野生种、栎叶枇杷和大渡河枇杷3个种的S-RNase基因型,为利用其优良性状开展种质创新,以及大渡河枇杷分类地位的探讨提供科学依据。【方法】以苹果S-RNase基因高度保守区设计兼并引物对3个种的基因组DNA进行PCR扩增,片段回收、克隆及测序,分别采用BLASTn、BLASTx、DNAMAN和CLUSTALW软件进行同源性检索、序列比对和结构分析。【结果】从参试的3个种中共分离了4个S等位基因,分别为S2-RNase、S26-RNase、S32-RNase和S34-RNase,其中S26-RNase、S32-RNase和S34-RNase为新分离的枇杷S-RNase基因,GenBank登录号分别为:MG765271、MG846012和MG812504。所克隆获得的4个枇杷S-RNase基因与苹果S-RNase基因的氨基酸序列结构相同,具有5个保守区(C1、C2、C3、RC4和C5)和1个高变区(HV)。此外,所获得的4个枇杷S-RNase基因电泳图谱及同源性和进化分析结果表明,大渡河枇杷可能为普通枇杷和栎叶枇杷的杂交后代。【结论】确定了普通枇杷野生种、栎叶枇杷和大渡河枇杷的S-RNase基因型分别为S2S26、S32S34和S26S32。大渡河枇杷S-RNase基因型及S-RNase的同源性和系统进化分析结果支持其可能为普通枇杷和栎叶枇杷杂交后代的结论。【Objective】Loquat（Eriobotrya japonica） demonstrates gametophytic self-incompatibility that is controlled by the S-locus encoding a polymorphic stylar ribonuclease（S-RNase）. The S-genotype is therefore an important consideration in breeding strategies. However, there has been no previous study dealing with assessement of the S-alleles in wild Eriobotrya species. Therefore, the objective of this study is to determine the S-genotypes of three loquat species, including E. japonica, E. prinoides var. dadunensis, and E. prinoides, in order to provide the scientific basis for the germplasm innovation and taxonomic reseach of E. prinoides var. dadunensis.【Methods】Using two pairs of primers designed according to conserved nucleotide sequences of known S-RNase alleles in Malus（apple）. These primers included a forward primer localized in the C1 region（‘FTQQYQ＇： 5＇-TTTACGCAGCAATATCAG-3＇）, and two reverse primers localized in between the hypervariable region（HV） and the C3（‘IIWPNV＇： 5＇-ACRTTCGGCCAAATMATT-3＇） and C5（‘FI（D/N）CP（H/R）＇： 5＇-GYGGGGGCARTYTATGAA-3＇） conserved regions. PCR amplification was carried out of genomic DNAs of three loquat species. Amplified products were separated by electrophoresis on 2.0% agarose gels, stained with ethidium-bromide（0.5%）, photographed using the UVP-gel documentation system（EC3 System; UVPCo., Upland, CA, USA）. A 100-bp DNA ladder（Tiangen Biotech Co. Ltd., Beijing, PR China） was used for estimating the molecular sizes of the amplicons. Reproducible amplified target fragments were purified using a DP210 DNA gel extraction kit（Tiangen Biotech Beijing, China）, were cloned into the pGEM-T Easy Vector（Promega Corporation, Australia）, were transformed into Escherichia coli, identified by colony PCR, and then were bidirectionally sequenced by BGI Life Tech Co., Ltd.（Beijing）. The obtained nucleotide and putative amino acid sequences were searched against NCBI using BLASTn and BLASTx to identify homo

关 键 词：大渡河枇杷 S基因 S基因型 分类地位 

分 类 号：S667.3[农业科学—果树学]