XJ 160 virus was isolated in 1990 from Xinjiang, China. The biological characteristics of XJ 160 virus suggested that it might be a Togavirus. The serological tests showed that XJ 160 virus is a Sindbis like virus. Th...XJ 160 virus was isolated in 1990 from Xinjiang, China. The biological characteristics of XJ 160 virus suggested that it might be a Togavirus. The serological tests showed that XJ 160 virus is a Sindbis like virus. The complete 11626 base nucleotide sequence of XJ 160 has been determined recently. It is necessary to establish a reverse genetic system for rescue of XJ 160 virus from its cDNA clone. We describe here the constructon of full length cDNA clone of XJ 160 virus. The total RNA were extracted from BHK cells infected with XJ 160 virus. Specific primers were used to amplify five fragments covering the whole genome of XJ 160 virus. All of these five fragments have the restriction enzyme sites at both termini to be assembled into the full length cDNA clone. The five PCR fragments were cloned into pGEM T vector. The clones with SP6 inserted direction were selected for subsequent manipulation. After a series of plasmid manipulation, the full length cDNA clone of XJ 160 virus was assembled into pBluescript vector. The restriction enzyme assay and sequencing analysis demonstrated that the whole genome of XJ 160 virus was correctly assembled into pBluescript vector, with added SP6 promoter in 5′ terminus and poly A in its 3′ terminus.展开更多
经过对其他物种CCR mRNA序列的对比分析后,设计保守区兼并引物,首先用RT-PCR方法得到229 bp CCR mRNA部分序列,之后通过RACE(Rapid Amplification of cDNA Ends)方法,成功克隆得到包括5'UTR和3'UTR在内的CCR全部mRNA序列,并进...经过对其他物种CCR mRNA序列的对比分析后,设计保守区兼并引物,首先用RT-PCR方法得到229 bp CCR mRNA部分序列,之后通过RACE(Rapid Amplification of cDNA Ends)方法,成功克隆得到包括5'UTR和3'UTR在内的CCR全部mRNA序列,并进行了分析,所得CCRmRNA全序列共1 243个碱基,CDS共999个碱基(103-1 101),编码氨基酸332,和其他多个物种的CCR序列比对结果显示相似度均在80%以上。此序列成功克隆之前,尚未见到白花泡桐木质素代谢关键酶基因的报道,这对丰富白花泡桐基因资源、有针对性的进行白花泡桐品质或材质改良有巨大的意义。展开更多
文摘XJ 160 virus was isolated in 1990 from Xinjiang, China. The biological characteristics of XJ 160 virus suggested that it might be a Togavirus. The serological tests showed that XJ 160 virus is a Sindbis like virus. The complete 11626 base nucleotide sequence of XJ 160 has been determined recently. It is necessary to establish a reverse genetic system for rescue of XJ 160 virus from its cDNA clone. We describe here the constructon of full length cDNA clone of XJ 160 virus. The total RNA were extracted from BHK cells infected with XJ 160 virus. Specific primers were used to amplify five fragments covering the whole genome of XJ 160 virus. All of these five fragments have the restriction enzyme sites at both termini to be assembled into the full length cDNA clone. The five PCR fragments were cloned into pGEM T vector. The clones with SP6 inserted direction were selected for subsequent manipulation. After a series of plasmid manipulation, the full length cDNA clone of XJ 160 virus was assembled into pBluescript vector. The restriction enzyme assay and sequencing analysis demonstrated that the whole genome of XJ 160 virus was correctly assembled into pBluescript vector, with added SP6 promoter in 5′ terminus and poly A in its 3′ terminus.
