梨Ty1-copia类逆转座子逆转录酶序列的克隆及分析

摘要

Primers of reverse transcriptase of tyl — copia-like retrotransposon were designed based on the conserved domains; fragments was amplified by PCR from 10 Pyrus species (cultivars). The amplicons were cloned into pMD18-T vectors after purification; positive clones were selected and identified by colony PCR, then sequenced and analyzed. 14 sequences of reverse transcriptase of tyl— copia—like retrotransposon with length varied from 250 bp to 267 bp were obtained from P. Communis cv. Red Cornice and clustered into 4 families. Sequence alignment revealed high heterogeneity mainly characterized by deletion mutation. Analysis on the amino acid sequences showed that stop codon mutation occurred in 2 sequences. Amino acid sequence alignment with reported corresponding sequences of other species showed high consistency.%根据Ty1-copia类逆转座子逆转录酶的保守区设计简并引物,从10份梨属(Prrus)材料中扩增该逆转座子片段.扩增产物经纯化后克隆于pMD 18-T质粒载体,选择阳性克隆,再经菌落PCR鉴定、测序及序列分析,获得了14条梨Ty1-copia类逆转座子逆转录酶序列.这些核苷酸序列长度为250～267 bp,具有较高的异质性,主要表现为缺失突变,通过核苷酸聚类可将14条序列分为4个家族.对这些片段的氨基酸序列进行分析,结果显示有2个序列发生了终止密码子突变.该研究获得的梨属植物Ty1-copia类逆转座子逆转录酶氨基酸序列与已报道的其他生物的相应序列有较高的一致性.