为了比较不同地理株的捕食线虫性真菌-少孢节丛孢菌丝氨酸蛋白酶基因的同源性,首先利用Biospin真菌基因组DNA提取试剂盒,提取少孢节丛孢菌内蒙古株(ARthRobotRys oligospoRa CHIM)的DNA,然后根据GenBank中相应丝氨酸蛋白酶基因,分别设计上下游引物;再经PCR扩增后,纯化并回收扩增产物,然后进行序列测定并分析。结果表明:少孢节丛孢菌内蒙古株丝氨酸蛋白酶基因序列与少孢节丛孢菌瑞典株(AY444597)和少孢节丛孢菌云南株(AF516146)同源性分别为80.3%和84.1%;虽然这3株菌具有较高的亲缘性,但其差异也非常明显,这一结果说明少孢节丛孢菌在不同国家或地区间有差异。
To asceRtain the homology of the seRine pRotease gene of ARthRobotRys oligospoRa in diffeRent geogRaphical isolates． DNA of A． oligospoRa CHIM was extRacted and puRified by Biospin fungi genomic DNA extRacted kit． Then the paRtial seRine pRoteinase gene of the A． oligospoRa was amplified with specific pRimeRs，and PCR pRoduct was Recycled． LateR， the nucleic acid sequence was deteRmined and compaRed with the Related gene sequences of A． oligospoRa fRom GenBank． The Results showed that the homology of seRine pRotease genes of A． oligospoRa fRom InneR Mongolia compaRed with the stRains fRom Sweden( AY444597) oR Yunnan( AF516146) was 80． 3% and 84． 1% Respectively． All the Results indicate although the thRee stRains weRe consanguinity， the diffeRence was obvious，and A． oligospoRa weRe diffeRent in stRains fRom diffeRent nations and Regions．
[1] 杨晓野，杨莲茹，刘珍莲，等． 捕食线虫性真菌的分离培 养及分布规律[J]． 中国兽医学报， 2003， 23( 4) : 344 - 347．
[2] 杨晓野，杨莲茹，刘珍莲，等． 捕食线虫性真菌———少 孢节丛孢菌体外杀寄生性线虫幼虫的研究[J]． 畜牧 兽医学报， 2004， 35( 3) : 334 - 337．
[3] 杨晓野，吴彩艳，杨莲茹，等． 口服少孢节丛孢菌孢子 对家畜粪便中线虫幼虫的杀灭研究[J]． 畜牧兽医学 报， 2005( 9) ， 927 - 930．
[4] 杨晓野，杨莲茹，刘珍莲，等． 捕食线虫性真菌———少 孢节丛孢菌CHIM 株捕食特性研究[J]． 中国兽医寄 生虫病， 1999，7 ( 2) : 14 - 17．
[5] 杨莲茹，杨晓野，刘珍莲，等． 捕食线虫性真菌生长特 性的研究[J]． 中国兽医科技， 2003， 33( 2) : 3 - 7．
[6] 邓长胜，王瑞，杨晓野，等． 氨基酸对少孢节丛孢菌 ( ARthRobotRys oligospoRa) 捕食器产生的影响[J]． 畜牧 兽医学报， 2009， 40( 4) : 561 - 553．
[7] 王瑞，杨晓野，杨莲茹，等． 不同碳氮源及其比例和 培养基初始pH 值对少孢节丛孢菌菌丝生长的影响
[J]． 黑龙江畜牧兽医， 2008( 9) : 11 - 14．
[8] 王瑞，杨晓野，韩海宾，等． 不同氨基酸对少孢节丛 孢菌( ARthRobotRyso ligospoRa) 捕食器诱导效果的研究
[J]． 黑龙江畜牧兽医， 2008( 7) : 7 - 9．
[9] 韩海宾，杨晓野，杨莲茹，等． 少孢节丛孢菌粗蛋白和 氨基酸含量的测定[J]． 中国兽医科学，2008，38( 7) : 612 - 615．
[10] 王瑞，杨莲茹，杨晓野，等． 少孢节丛孢菌捕食器表 面聚合物的诱导与提取及生化性质研究[J]． 畜牧与 兽医， 2010， 42( 8) : 10 - 14．
[11] 王瑞，杨莲茹，杨晓野，等． 少孢节丛孢菌( ARthRobotRys oligospoRa) 表面聚合物的提取及生化性质研究
[J]． 畜牧兽医学报， 2010， 41( 10) : 1301 - 1305．
[12] 王瑞，杨莲茹，杨晓野，等． 少孢节丛孢菌胞外蛋白 酶的诱导与生化性质[J]． 中国兽医科学，2010，40 ( 5) : 446 - 451．
[13] 王军，王瑞，杨晓野，等． 少孢节丛孢菌捕食器表 面聚合物在捕食线虫过程中的作用研究[J]． 畜牧医 学报， 2010， 41( 7) : 859 - 865．
[14] Li J，Yu L，Yang J， et al． New insights into the evolution of subtilisin-like seRine pRotease genes in Pezizomycotina
[J]． BMC Evol Biol， 2010，9 ( 10) : 68．
[15] Tunlid A， Jansson S． PRoteases and theiR involvement in the infection and immobilization of nematodes by the nematophagous fungus ARthRobotRys oligospoRa[J]． Appl EnviRon MicRobiol， 1991， 57( 10) : 2868 - 2872．
[16] Tunlid A，Rosen S，Ek B， et al． PuRification and chaRacteRization of an extRacellulaR seRine pRotease fRom the nematode-tRapping fungus ARthRobotRys oligospoRa[J]． MicRobiology， 1994， 140( Pt 7) : 1687 - 95．
[17] Wang M，Yang J，Zhang K Q． ChaRacteRization of an extRacellulaR pRotease and its cDNA fRom the nematodetRapping fungus MonacRospoRium micRoscaphoides[J]． Can J MicRobiol， 2006， 52( 2) : 130 - 9．
[18] Ahman J，Ek B，Rask L，et al． Sequence analysis and Regulation of a gene encoding a cuticle-degRading seRine pRotease fRom the nematophagous fungus ARthRobotRys oligospoRa
[J]． MicRobiology， 1996， 142( 7) ; 1605 - 1616．19] Nagee A，AchaRya A，Shete A， et al． MoleculaR chaRacteRization of an expRessed sequence tag RepResenting the cuticle - degRading seRine pRotease gene ( PII) fRom the nematophagous fungus ARthRobotRys ovifoRmis by diffeRential display technology[J]． Genet Mol Res， 2008，7 ( 4) : 1200 - 1208．
[20] Minglian Z，Minghe M，Keqin Z． ChaRacteRization of a neutRal seRine pRotease and its full-length cDNA fRom the nematode-tRapping fungus ARthRobotRys oligospoRa
[J]． Mycologia， 2004， 96( 1) : 16 - 22．
[21] Xu J，Mo M H，Huang X W， et al． ImpRovement on genetic tRansfoRmation in the nematode-tRapping fungus ARthRobotRys oligospoRa and its quantification on dung samples
[J]． Mycopathologia， 2005， 159( 4) : 533 - 538．
[22] Yang J，Liang L，Zhang Y， et al． PuRification and cloning of a novel seRine pRotease fRom the nematode-tRapping fungus Dactylellina vaRietas and its potential Roles in infection against nematodes[J]． Appl MicRobiol Biotechnol， 2007， 75( 3) : 557 - 565．
[23] Yang J，Li J，Liang L， et al． Cloning and chaRacteRization of an extRacellulaR seRine pRotease fRom the nematodetRapping fungus ARthRobotRys conoides[J]． ARch MicRobiol， 2007， 188( 2) : 167 - 174．
[24] Ahman J，Johansson T，Olsson M，et al． ImpRoving the pathogenicity of a nematode-tRapping fungus by genetic engineeRing of a subtilisin with nematotoxic activity[J]． Appl EnviRon MicRobiol， 2002， 68( 7) : 3408 - 3415．
[25] Fekete C，TholandeR M，RajashekaR B， et al． PaRalysis of nematodes: shifts in the tRanscRiptome of the nematodetRapping fungus MonacRospoRium haptotylum duRing infection of CaenoRhabditis elegans[J]． EnviRon MicRobiol，2008， 10(2)：364 - 375．
