中文摘要：

[目的]研究大腹园蛛组织蛋白酶B相关基因Avg 1（基因登录号：AY 302573）编码蛋白的生物学特性．[方法]设计合成引物（AvgS，AvgX），扩增Avg 1，去除信号肽部分基因Avg，将其与原核表达载体pET-28a（＋），pET-20b连接，转化到E．coli BL 21（DE3）中．[结果]鉴定为阳性克隆后，用IPTG诱导表达，SDS-PAGE电泳显示Avg-28a-DE3在近35．7ku处出现明显的表达带，其大小与预计分子质量（35．2ku）相当；经薄层扫描分析显示，表达蛋白量占菌体蛋白总量的39．6％，且表达形式主要为包涵体，而Avg-20 b-DE3表达产物未见表达带，但RT—PCR方法检测到了Avg mRNA的转录；将表达的包涵体蛋白进行变性及复性处理，与诱导的Avg-20 b-DE 3表达菌经超声波碎菌处理上清液-同进行蛋白酶活性测定，呈复性的包涵体蛋白不具有酶活性，而Avg-20 b-DE 3的碎菌上清液显示明显的蛋白酶活性．表达的包涵体蛋白经电洗脱纯化后，常规方法免疫吉绒獭兔，抗血清经ELISA法检测，呈明显阳性反应，经Western blot检测在35．7ku处出现特异结合带．[结论]Avg 1原核表达蛋白具有良好的免疫原性．

英文摘要：

OBJECTIVE To study the biological specialties of protein coded by Araneus Ventricousus cathepsin B-like gene Avg 1（genbank AY 302573）. METHODS The free signle peptide Avg was amplified by PCR with the primers AvgS and AvgX, which designed according to Avg 1 cDNA. Two recombinant expression plasimid of Avg-28 a and Avg-20 b were constructed by T 4 ligase and transformed into host strain E. coli BL 21 （DE 3）. RESULTSAvg-28 a-DE3 was induced by IPTG and the special protein （about 35.7 ku） was detected by SDS-PAGE. Thin-layer scan result showed that the special protein amounted to 39.6 % of the total protein of E. coli BL21 （DE3）. The pattern of expression was mainly inclusion body. SDS-PAGE of inducing Avg-20 b-DE 3 by IPTG did not show the special protein but the Avg mRNA transcription was detected by RT-PCR. The inclusion body protein treated by denaturing and renaturing was not shown the activity of cathepsin, but the supernatant of Avg-20 b-DE 3 by supersonic treatment showed the high activity. The rabbits were immunized by the inclusion body protein after being purified by SDS-PAGE and electrophoresis eluted. CONCLUSION The inclusion body protein has an excellent antigenicity.