石榴果皮POD基因克隆与序列分析.docVIP

石榴果皮POD基因克隆与序列分析 　　摘要：本研究以‘泰山?t’石榴果实为试材，利用RT-PCR和RACE技术克隆果皮中POD基因cDNA片段，利用BLAST和ORF Finder分析其核苷酸序列，并利用ProtParam、SPOMA、DANMAN等软件分析其氨基酸序列。结果表明，克隆得到1 092 bp的cDNA片段，该基因含828 bp的CDS序列，编码276个氨基酸。该基因核苷酸序列与烟草、茜草、花生二倍体杂生种和潘那利番茄等植物的相似性分别为77%、77%、76%和76%；氨基酸序列与荔枝、克莱门柚、苹果和白梨的相似性分别为89%、87%、84%和84%；GenBank登录号为KY129694。预测蛋白质理论分子量为30 582.59 D，等电点（pI）为8.83，具有过氧化物酶完整的保守结构域，含有Cd00693和 Cl00196两个保守功能域，为亲水性蛋白；蛋白质二级结构主要由无规则卷曲（38.41%）、α-螺旋（37.68%）、延伸链（13.77%）和β-转角（10.14%）组成。该研究结果为揭示石榴果实褐变机理奠定了理论基础。 　　关键词??石榴；POD基因；RT-PCR；克隆；序列分析 　　中图分类号：S665.4 文献标识号：A文章编号：-4942（2017）03-0016-05 　　AbstractThe cDNA sequence of POD was cloned from Taishanhong pomegranate peel by RT-PCR and RACE technique. The nucleotide sequence of POD was analyzed by BLAST and ORF Finder. The amino acid sequence of POD was analyzed by ProtParam， SPOMA and DANMAN softwares. The results indicated that 1 092 bp cDNA sequence of POD was obtained. It contained 828 bp CDS sequence and encoded 276 amino acids. The similarity of POD nucleotide sequence in pomegranate with Nicotiana tabacum， Rubia cordifolia， Arachis ipaensis and Solanum pennellii was 77%， 77%， 76% and 76%， respectively. Based on the deduced amino acid sequences of POD genes， the similarity of pomegranate with Litchi chinensis， Citrus clementina， Malus domestica and Pyrus bretschneideri was 89%， 87%， 84% and 84%， respectively. The GenBank accession number was KY129694. The theoretical molecular mass of POD protein was 30 582.59 D， and the isoelectric point was 8.83. The protein of POD gene was hydrophilic protein， which had the complete conservative structure domain of peroxidase， and contained two conservative functional domains， Cd00693 and Cl00196. The secondary structure of POD protein was composed by random coil （38.41%）， α-helix （37.68%）， extended strand （13.77%） and β-turn （10.14%）. The results established a theoretical foundation for revealing the mechanism of peel browning in pomegranate fruit. 　　KeywordsPomegranate； POD gene； RT-PCR； Cloning； Sequence a