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【目的】获得石榴果实PgANS、PgCHS和PgACT基因,并对这三个基因进行序列分析。【方法】以喀什甜石榴为试材,通过酚-氯仿法提取籽粒中的总RNA,用TIANScript cDNA试剂盒反转录合成第一链cDNA,并用特异引物对其RT-PCR扩增,将得到的cDNA片段连接到PMD19-T载体上,然后将该载体转到DH5α,并对阳性克隆进行序列分析。【结果】PgANS基因片段长度为474bp,Genbank登录号为KT779433,与NCBI数据库中KF841619、GU376749两种石榴核苷酸的同源性均为99%,与甘薯核苷酸(GU59821)、白桑核苷酸(JF499385)、草莓核苷酸(AY695817)的同源性分别为85%、79%、76%;PgCHS和PgACT基因片段长度分别为140bp和148bp,分别与NCBI数据库中JF747148、GU376750两种石榴核苷酸同源性为99%和93%。【结论】获得PgANS、PgCHS和PgACT基因片段,为进一步研究贮藏条件对石榴籽粒花色苷的影响奠定了基础。
【Objective】 The PgANS, PgCHS and PgACT genes of pomegranate fruit were obtained, and the sequence analysis of these three genes was carried out. 【Method】 Using Kashi sweet pomegranate as test material, the total RNA was extracted from the grains by phenol-chloroform method. The first strand cDNA was reverse transcribed with TIANScript cDNA kit and amplified by RT-PCR with specific primers. CDNA fragment was ligated to the PMD19-T vector, which was then transferred to DH5α and the positive clones were subjected to sequence analysis. 【Result】 The PgANS gene fragment was 474 bp in length and GenBank accession number KT779433 was 99% homologous to KF841619 and GU376749 in the NCBI database. The homology of PgANS gene to sweetpotato nucleotide (GU59821), white mulberry nucleus The homologies of nucleotide (JF499385) and strawberry (AY695817) were 85%, 79% and 76%, respectively. The lengths of PgCHS and PgACT genes were 140bp and 148bp, respectively, which were respectively linked with JF747148 and GU376750 in NCBI database Pomegranate nucleotide homology was 99% and 93%. 【Conclusion】 The PgANS, PgCHS and PgACT gene fragments were obtained, which laid the foundation for further study on the effect of storage conditions on anthocyanin in pomegranate grain.