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1、___________________________________________________________________________http://www.paper.edu.cn日本血吸蟲(chóng)DNA多價(jià)疫苗pVIVO2SjFABP－23的構(gòu)建及其保護(hù)性免疫11※11222李春艷余龍江劉智朱路朱曉華胡媛石佑恩摘要：目的：構(gòu)建日本血吸蟲(chóng)多價(jià)DNA疫苗pVIVO2SjFABP－23，并觀察其在小鼠抗血吸蟲(chóng)感染中的免疫保護(hù)作用。方法：根據(jù)SjFABP和Sj23的基因序列合成兩對(duì)特異性引物,利用重組
2、PCR技術(shù)將日本血吸蟲(chóng)SjFABP和Sj23的基因片段進(jìn)行拼接,得到融合基因SjFABP-23,再將該片段重組于pGEM-T載體中,進(jìn)行PCR擴(kuò)增及DNA序列測(cè)定。然后經(jīng)雙酶切將目的片段SjFABP-23克隆到pVIVO2的一個(gè)多克隆位點(diǎn)上轉(zhuǎn)化E.coliGT110獲得重組質(zhì)粒pVIVO2SjFABP－23，免疫小鼠進(jìn)行免疫保護(hù)性實(shí)驗(yàn)。結(jié)果：PCR擴(kuò)增出一條大小約為1.1Kb的目的片段。免疫小鼠獲得52.14%減蟲(chóng)率(P<0.05)和60.93%的減卵率（P<0.01）。結(jié)論：PCR成功擴(kuò)增SjFABP
3、－23的基因片段，血吸蟲(chóng)DNA多價(jià)疫苗pVIVO2SjFABP－23構(gòu)建成功，該疫苗在小鼠抗血吸蟲(chóng)感染中具有一定的免疫保護(hù)作用。關(guān)鍵詞：日本血吸蟲(chóng)；23KDa膜蛋白；脂肪酸結(jié)合蛋白；多價(jià)DNA疫苗；免疫保護(hù)CONSTRUCTIONOFTHEMULTIVALENTDNAVACCINEpVIVO2SjFABP－23OFSCHISTOSOMAJAPONICUMANDSTUDIESONITSIMMUNOPROTECTIONINMICE11※1LIChun-YanYULong-JiangLIUZhietal(Co
4、llegeofLifeScience&Technology,HuazhongUniversityofScience&Technology,Wuhan430074,China；TongjiMedicalCollege,HuazhongUniversityofScienceandTechnology,Wuhan430030,China)Abstract:ObjectiveToconstructandpreparethemultivalentDNAvaccinepVIVO2SjFABP－23andobserv
5、eitsimmunoprotectionagainstSchistosomajaponicuminmice.MethodsTwocouplesofspecificprimersweredesignedaccordingtogenesequencesofSjFABP(fattyacidbindingprotein)andSj23（23KDamembraneprotein）,twofragmentswereobtainedandsplicedthroughalinkersequencebyrecombina
6、tionPCR.FusionfragmentformedwasinsertedintopGEM-TEasyvector,andidentifiedbyrestrictionanalysisandsequence,digestedwithBamHIandEcoRI,clonedintoexpressedplasmidpVIVO2/MCSandtransformedinE.coliGT110,recombinationplasmidpVIVO2SjFABP－23wasextracted,purifiedan
7、dinoculatedintoBALB/cmicebyintramuscularinjection.MicewerechallengedbySchistosomajaponicumcercariaeandthenkilled.Adultwormandeggwerecounted,respectively.ResultsElectrophoresisofSjFABP-23on0.7%agarosegelshowedanobviousbandof1.1Kb,whileonesofSjFABPandSj23w
8、ere440bpand670bp,respectively.RecombinationplasmidpVIVO2SjFABP－23wasthenconstructed。Protectiveexperimentsappearedthatwormreductionratewas52.14%(P<0.05),andeggsreductionratewas60.93%(P<0.01)whencomparedwiththecontrolgroupin
