动物医学进展
動物醫學進展
동물의학진전
PROGRESS IN VETERINARY MEDICINE
2015年
2期
39-42
,共4页
王秀祯%罗永芳%夏文君%罗雪刚%彭少静%周念%李继祥
王秀禎%囉永芳%夏文君%囉雪剛%彭少靜%週唸%李繼祥
왕수정%라영방%하문군%라설강%팽소정%주념%리계상
鸭疫里默杆菌%mazp基因%免疫原性%保护指数
鴨疫裏默桿菌%mazp基因%免疫原性%保護指數
압역리묵간균%mazp기인%면역원성%보호지수
Riemerella anatipesti f er%mazp gene%immunogenicity%protective index
为探索鸭疫里默杆菌不同血清型菌株的共同保护性抗原,克隆表达m az p基因,并采用动物试验测定表达蛋白的免疫原性。根据RA‐GD株mazp基因核苷酸序列设计特异性引物进行PCR扩增,克隆测序获得RA‐A F株细菌1326 bp的核苷酸片段,比较分析发现其保守性高。以克隆的mazp基因构建重组表达质粒pET‐32a‐mazp并转化 E .coli BL21(DE3),IPTG诱导获得重组蛋白。重组蛋白为免疫抗原免疫小鸭15 d后,RA‐BSY(血清1型)和RA‐AF(血清2型)攻毒的保护指数分别为77.8和62.4;抗血清能延迟发病和死亡时间。结果表明,鸭疫里默杆菌mazp蛋白具有免疫原性,免疫鸭对血清1型和2型RA感染具有抵抗力。
為探索鴨疫裏默桿菌不同血清型菌株的共同保護性抗原,剋隆錶達m az p基因,併採用動物試驗測定錶達蛋白的免疫原性。根據RA‐GD株mazp基因覈苷痠序列設計特異性引物進行PCR擴增,剋隆測序穫得RA‐A F株細菌1326 bp的覈苷痠片段,比較分析髮現其保守性高。以剋隆的mazp基因構建重組錶達質粒pET‐32a‐mazp併轉化 E .coli BL21(DE3),IPTG誘導穫得重組蛋白。重組蛋白為免疫抗原免疫小鴨15 d後,RA‐BSY(血清1型)和RA‐AF(血清2型)攻毒的保護指數分彆為77.8和62.4;抗血清能延遲髮病和死亡時間。結果錶明,鴨疫裏默桿菌mazp蛋白具有免疫原性,免疫鴨對血清1型和2型RA感染具有牴抗力。
위탐색압역리묵간균불동혈청형균주적공동보호성항원,극륭표체m az p기인,병채용동물시험측정표체단백적면역원성。근거RA‐GD주mazp기인핵감산서렬설계특이성인물진행PCR확증,극륭측서획득RA‐A F주세균1326 bp적핵감산편단,비교분석발현기보수성고。이극륭적mazp기인구건중조표체질립pET‐32a‐mazp병전화 E .coli BL21(DE3),IPTG유도획득중조단백。중조단백위면역항원면역소압15 d후,RA‐BSY(혈청1형)화RA‐AF(혈청2형)공독적보호지수분별위77.8화62.4;항혈청능연지발병화사망시간。결과표명,압역리묵간균mazp단백구유면역원성,면역압대혈청1형화2형RA감염구유저항력。
To study the mutual protective antigen of Riemerella anatipesti f er (RA ) strains ,the reserch cloned and expressed membrane‐associated zinc metalloprotease (mazp) gene of RA ,and studied its ex‐pression immunogenicity using animal test .The specific primers were designed based on the nucleotide se‐quence of mazp gene of RA‐GD strain and the mazp gene of RA‐AF strain was cloned and sequenced .Com‐pared with other strains through BLAST analysis ,the 1 326 bp nucleotide sequence of mazp gene of RA -AF strain was highly conserved .Then the mazp gene fragment was cloned into pET‐32a (+ ) to construct expression plasmid pET‐32a‐mazp .It expressed recombination protein in E .coli BL21 (DE3) when in‐duced by isopropyl thiogalactoside (IPTG) .Subcutaneous inoculation of the recombination protein in 5‐day‐old ducklings gave adequate protective index (PI=62 .4) against challenge with RA‐AF strain (sero‐type 2) 15 days after vaccination ,while the protection (PI=77 .8) was obtained challenged with RA‐BSY strain (serotype 1) .The antiserum of recombination protein was used as antibody to immunize ducklings , delaying morbility and death was obtained .The result indicated the recombination protein mazp has immu‐nogenicity ,and may play a role in protecting duck from infection of Riemerella anatipesti f er (sreotype 1 , 2) .