7型腺病毒疫苗株DNA左侧17.5mu片段克隆及4.8mu片段核苷酸序列分析

Cloning of 017.5 mu and sequencing of 04.8 mu of the lefe end DNA of the Ad7 vaccine strain left end

目的获得７型腺病毒疫苗株ＤＮＡ左侧０～１７５ｍｕ片段克隆，分析０～４８ｍｕ片段（末端倒置重复序列、包装信号位点和Ｅｌａ区）核苷酸序列。方法从Ａｄ７疫苗株感染的Ａ５４９细胞提取Ａｄ７ＤＮＡ，将其０３～１７５ｍｕ片段克隆到质粒ｐＡｄ７Ｔ，并用自动和银染方法测序。结果获得了Ａｄ７疫苗株左侧１７５ｍｕ片段克隆，测定了左侧１７３７ｂｐ核苷酸序列，其中Ｅｌａ区所编码的６３００、２４０００、２８０００等３个蛋白的ＤＮＡ序列，与Ｇｏｍｅｎ株的同源性分别为９８９％、９７３％和９７５％，推导编码氨基酸的同源性分别为９６６％、９６５％和９６９％。这３个蛋白与Ｇｒｉｄｅｒ株的同源性分别为１００％、９９７％、和９９７％；推导编码氨基酸的同源性分别为１００％、９９１％和９９２％。结论Ａｄ７疫苗株左侧１７３８ｂｐ核苷酸与Ａｄ７Ｇｏｍｅｎ株和Ｇｒｉｄｅｒ株相应片段，具有高度的同源性。

Objective Cloning of 017.5 mu DNA fragment of adenovirus 7 vaccine strain and sequencing of 04.8 mu fragment(1 737 bp). Methods Isolating and purifying Ad7 vaccine genome from the A549 cultured cells, putting 0.317.5 mu fragment into pAd7T plasmid, then sequencing the 04.8 mu fragment including inverted terminal repeats (ITR), packaging sequence and Ela region. Results We obtained 017.5 mu fragment of Ad7 vaccine strain genome and sequenced its left terminal 1 737 bp. Sequence analysis showed that the Ad7 vaccine strain Ela region encodes 6 300, 24 000 and 28 000 proteins. Compared with equivalent region of Ad7 Gomen strain, they share the homology of the nucleotide sequence 98.9%, 97.3%, 97.5% and the homology of the deduced amino acid sequence 96.6%, 96.5%, and 96.9% respectively. When compared with Ad7 Grider strain, they share the homology of the nucleotide sequence100%, 99.7%, 99.7% and the homology of the deduced amino acid sequence 100%, 99.1%, and 99.2% respectively. Conclusion The Ad7 vaccine strain left terminal 1 737 bp nucleotide sequence showed a high homology with corresponding region of Ad7 Gomen strain and Grider strain.