几种PCR方法在克隆鸭肠炎病毒未知基因中的应用

Application of Several PCR Methods in Cloning Unknown Genes of Duck Enteritis Virus

摘要:

以DEV基因组DNA为模板, 用简并PCR、改良Targeted gene walking PCR、改良的热不对称交错PCR和Long-PCR, 获得了5350 bp、11083 bp和2905 bp三段DEV未知基因片段, DNA序列分析发现包含9个开放阅读框, 将这些序列提交GenBank分别获得的登录号为: EF554396~EF554403。结果表明, 多种PCR方法联合使用可以高效的实现对鸭肠炎病毒未知基因的克隆。

Abstract:

Degenerate PCR, modified targeted gene walking PCR, modified thermal asymmetric interlaced PCR and Long-PCR were performed with genomic DNA of duck enteritis virus as a template and three fragments (5350 bp, 11083 bp, 2905 bp) of DEV genome were obtained. DNA sequence analysis revealed nine open reading frames encoding UL3, UL4, UL5, UL25, UL26, UL27, UL28, and UL30 gene of DEV, respectively. These sequences have been submitted to GenBank with accession numbers from EF554396 to EF554402. Joint application of several PCR techniques is efficient in cloning unknown genes of duck enteritis virus.