褐黄血蜱脂肪酸结合蛋白对褐黄血蜱卵及其原生质蛋白影响的研究  

Effect of FABP on spawning and protoplasmic proteins of Haemaphysalis flava

作　　者：王兰兰 刘金宝 程天印[1] WANG Lan-lan;LIU Jin-bao;CHENG Tian-yin(College of Veterinary Medicine,Hunan Agricultural University,Changsha 410128,China;Hunan Zhengda Livestock and Poultry Co.,LTD.,Ningxiang 410600,China)

机构地区：[1]湖南农业大学动物医学院,湖南长沙410128 [2]湖南正大畜禽有限公司,湖南宁乡410600

出　　处：《中国预防兽医学报》2024年第4期342-349,共8页Chinese Journal of Preventive Veterinary Medicine

基　　金：湖南省自然科学基金面上项目(2023JJ30287)。

摘　　要：为了解蜱类脂肪酸结合蛋白(FABP)及其基因的结构和功能,本研究基于褐黄血蜱转录组文库中的Contig 879序列设计引物,采用cDNA末端快速扩增技术(RACE)扩增其两翼缺失部分,拼接后得到编码褐黄血蜱FABP (HfFABP)的全长cDNA序列;采用qPCR法检测该基因在褐黄血蜱的不同发育阶段、不同组织器官中的转录水平;采用平行反应监测(PRM)法分析HfFABP dsRNA干扰对褐黄血蜱产卵和孵化,以及卵原生质蛋白的影响。结果显示,HfFABP的cDNA全长1 443 bp,ORF长396 bp,编码132个氨基酸的多肽。HfFABP基因在雌蜱中的转录水平高于幼蜱、若蜱和雄蜱;在蜱的卵巢、中肠和体壳中的转录水平高于其在唾液腺中的转录水平;吸血使HfFABP基因的转录水平升高。HfFABP dsRNA干扰使蜱产卵减少,其中约30%的卵大小不均、蜡质增多、塌瘪易裂;表观正常的卵孵化率降低,原生质中的FABP、卵黄蛋白原(Vitellogenin)、弹性蛋白酶抑制剂(Neutrophil elastase inhibitor)、天冬氨酸蛋白酶(Aspartic protease)和热休克蛋白83 (Heat shock protein 83)等12种蛋白的丰度大幅下降(P<0.01、P<0.05)。上述结果首次表明HfFABP在蜱卵发育过程中发挥重要作用,本研究为HfFABP在抑制雌蜱生殖中的应用提供科学依据。In order to understand the structure and function of fatty acid binding proteins(FABP)and the genes in ticks,primers were designed based on the sequence of Contig879 in the transcriptome library of Haemaphysalis flava,and the missing sequences of the two ends were amplified by the rapid-amplification of cDNA ends(RACE)method.Real-time PCR(qPCR)was used to determine the transcription level of the gene in different developmental stages and organs of ticks.The effect of H.flava FABP(HfFABP)dsRNA interference on the protoplasmic proteins of the eggs was analyzed by the parallel reaction monitoring(PRM)method.The results showed that the full-length HfFABP cDNA was 1443bp,and the ORF was 396bp,and it encoded a polypeptide of 132aa.The transcription level of this gene in female ticks was higher than that in larvae,nymphs and male ticks;the transcription level in the ovary,midgut,and integument was higher than that in the salivary gland.Blood sucking increased the transcription level of HfFABP.The interference of HfFABP dsRNA reduced spawning,and about 30%of the eggs were discrepant in size,waxy,fragile and easy to crack.The hatching rate of apparently normal eggs was reduced.The abundance of many proteins in eggs,such as FABP,vitellogenin,neutrophil elastase inhibitor,aspartic protease,and heat shock protein 83,decreased significantly in ticks with HfFABP dsRNA interference(P<0.01,P<0.05).These results indicate that HfFABP plays an important role in the development of tick eggs.This study provides a scientific basis for the application of HfFABP in the anti-female tick reproduction.

关 键 词：褐黄血蜱 脂肪酸结合蛋白(FABP) cDNA末端快速扩增技术(RACE) 平行反应监测(PRM) 

分 类 号：S855.9[农业科学—临床兽医学]