鸡传染性法氏囊病病毒内蒙古毒株VP2基因的克隆和序列分析

The Cloning and Sequencing of cDNA Gene Coding for VP 2 of Chinese IBDV NM Strain

以鸡传染性法氏囊病病毒内蒙古毒株（ＩＢＤＶＮＭ）ｄｓＲＮＡ为模板，用ＲＴＰＣＲ法扩增其主要寄主保护抗原ＶＰ２基因的全长ｃＤＮＡ，克隆于ｐＵＣ１９的ＸｂａＩ／ＫｐｎＩ位点，进行了全序列分析。序列比较发现ＩＢＤＶＮＭ毒株ＶＰ２基因与已报道的其它７个ＩＢＤＶＣＪ８０１ｂｋｆ、Ｃｕ１、ＰＢＧ９８、５２／７０、００２—７３、ＳＴＣ及ＶａｒｉａｎｔＥ毒株之间高度同源，其核苷酸序列的同源率为９１６％～９６．２％，推测的氨基酸序列的同源率为９６２％～９８．６％。ＩＢＤＶＮＭ毒株ＶＰ２高变异区的第一个亲水区氨基酸序列与ＣＪ８０１ｂｋｆ、Ｃｕ１、ＰＢＧ９８、５２／７０、ＳＴＣ、００２—７３比较，有一个氨基酸差异，第二个亲水区氨基酸序列与上述６个毒株完全相同。而与ＶａｒｉａｎｔＥ比较，两个亲水区内各有两个氨基酸差异。此外，ＩＢＤＶＮＭ毒株ＶＰ２具有强毒株所特有的７肽保守区：ＳＷＳＡＳＧＳ。这些结果表明，ＩＢＤＶＮＭ毒株为标准血清Ⅰ型ＩＢＤＶ强毒株。

The cDNA of major host protective antigen VP 2 gene from Chinese IBDV NM strain isolated from Inner Mongolia in North China was synthesized by reverse transcription followed by polymerase chain reaction (PCR) using a pair of specific synthetic primers.The amplified cDNA was cloned at XbaI and KpnI sites in pUC19.The nucleotide sequence of VP 2 gene cDNA was determined and the amino acid sequence was deduced.Both the nucleotide sequence and amino acid sequence were compared with seven published sequences of VP 2 gene from IBDV strains CJ 801bkf (Chinese isolate),Cu1 (German strain),PBG98 (British attenuated vaccine strain),52/70 (a virulent British isolate),002-73(Australian strain),STC (the United States standard challenge strain) and Variant E (Delaware variant strain E).All eight strains are closely related.The sequence homology between IBDV NM strain and the seven other strains was 91 6%～96 2% at the nucleotide level and 96 2%～98 6% at the amino acid level.In the first hydrophilic region of VP 2 only one amino acid is different between IBDV NM and CJ 801bkf,Cu1,PBG98,52/70,STC,and 002 73 strains.But in the second hydrophilic region there were not any amino acid changes between IBDV NM and these six strains.Compared with VP 2 gene of Variant E two different amino acids in each of both hydrophilic regions were found.The VP 2 of IBDV NM strain has a typical heptapeptide motif:SWSASGS found in all highly virulent strains,suggesting that the Chinese IBDV NM strain is a highly virulent strain of standard serotype I of IBDV.