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根据烟草花叶病毒组（Ｔｏｂａｍｏｖｉｒｕｓｅｓ）３’端非编码区保守区域，人工合成下游引物Ｐ１，Ｐ２，Ｐ３，分别以这些引物引导在反转录酶作用下合成双链 ｃＤＮＡ，并克隆到 ｐＢＳ载体上。Ｈｉｎｄ Ⅲ＋ ＰｓｔⅠ双酶切分析重组稿子后，得到 １５个阳性重组子，其中重组子 ｐＢＦ３含有 ２ ４３２ ｂｐ外源 ＤＮＡ。序列分析结果表明：该 ２ ４３２ｂｐ的序列含 ２个开放读框，即鸡蛋花花叶病毒移动蛋白基因序列和外壳蛋白基因序列。移动蛋白基因序列起始于８０６位ＡＴＧ，中止于 １５７６位ＴＡＧ。推导的氨基酸序列共２５６个氨基酸，分子量为２８．５ ｋＤａ。外壳蛋白基因序列起始于 １６３５位ＡＴＧ，中止于 ２ １５８位ＴＡＡ。推导的氨基酸序列共 １７４个氨基酸，分子量为１９．４ ｋＤａ．此外该片段还包含部分１８０ ｋＤａ蛋白和全部３’端非编码区核苷酸序列。
According to the conserved region of 3 ’untranslated region in tobacco mosaic virus (Tobamoviruses), the downstream primers P1, P2 and P3 were synthesized and double-stranded cDNA was synthesized under the action of reverse transcriptase using these primers respectively. on. After digestion with Hind Ⅲ + PstⅠ, 15 positive recombinant plasmids were obtained, in which recombinant pBF3 contained 2 432 bp foreign DNA. Sequence analysis showed that the 2 432 bp sequence contained two open reading frames (ORFs) and the coat protein gene sequence. The mobile protein gene sequence starts at 806 ATG and ends at 1576 TAGs. The deduced amino acid sequence has a total of 256 amino acids with a molecular weight of 28.5 kDa. The coat protein gene sequence begins at ATG at position 1635 and ends at the TAA at position 2,158. The deduced amino acid sequence was 174 amino acids with a molecular weight of 19.4 kDa. In addition this fragment also contains part of the 180 kDa protein and all 3 ’non-coding region nucleotide sequences.