日本鳗鲡肝脏表达抗菌肽2基因的克隆与表达

发布时间：2018-02-21 17:01

本文关键词： 抗菌肽 LEAP- 日本鳗鲡迟缓爱德华氏菌　出处：《水生生物学报》2016年02期 　论文类型：期刊论文

【摘要】：为探讨鱼类抗菌肽基因的生物学功能,研究应用RACE方法克隆获得了日本鳗鲡(Anguilla japonica)肝脏表达抗菌肽2基因(Liver-Expressed Antimicrobial Peptide 2,LEAP-2),即AJLEAP-2的cDNA序列,全长为450 bp,开放阅读框编码89个氨基酸。其成熟肽含有LEAP-2保守基序"C-X5-C-X4-C-X4-C"。AJLEAP-2基因组结构与其他脊椎动物LEAP-2相同,都包含有三个外显子。利用荧光定量PCR检测了AJLEAP-2在日本鳗鲡不同组织/器官中的表达,发现其转录子在肝脏中表达量最高,是内参基因(β-acHn)的6倍;其次是肠道,但其表达量仅为肝脏的1/130。此外,还检测了AJLEAP-2在日本鳗鲡玻璃鳗(Glass eel)阶段的转录表达水平,结果显示,玻璃鳗中AJLEAP-2的转录表达量仅低于黑仔期的肝脏,为黑仔鳗肠道表达量的2倍。LPS和迟缓爱德华菌(Edwardsiella tarda)刺激能显著上调鳗鲡血液中AJLEAP-2的转录表达,刺激16h后上调倍数最高,分别为对照组的86倍和12倍。此外,LPS刺激72h和E tarda刺激8h后,肠道中AJLEAP-2显著上调表达(P0.05),为对照组的8倍。Poly I:C刺激24h后,血液中AJLEAP-2转录表达显著下调。结果表明,AJLEAP-2在日本鳗鲡抗细菌感染过程中起重要的作用。
[Abstract]:In order to study the biological function of antimicrobial peptide gene in fish, RACE method was used to clone Anguilla japonica (Anguilla japonica) liver expressing antimicrobial peptide 2 gene, the cDNA sequence of AJLEAP-2 was obtained. The mature peptide contains LEAP-2 conserved motif "C-X5-C-X4-C-X4-C" .AJLEAP-2 has the same genomic structure as other vertebrate LEAP-2. Using fluorescent quantitative PCR to detect the expression of AJLEAP-2 in different tissues / organs of Japanese eel (Anguilla japonica), it was found that the expression of AJLEAP-2 was the highest in the liver, six times that of 尾 -acHnn. the second was intestinal tract. But its expression was only 1 / 130 of that in the liver. In addition, the transcriptional expression of AJLEAP-2 in the glass eel of Japanese eel (Anguilla japonica) was detected. The results showed that the transcriptional expression of AJLEAP-2 in the glass eel was only lower than that in the black eel. LPs and Edwardsiella tarda stimulated eels significantly up-regulated the transcription of AJLEAP-2 in the blood of Anguilla japonica, which was the highest after 16 hours of stimulation. In addition, after 72 h and 8 h of E tarda stimulation, the expression of AJLEAP-2 was significantly up-regulated in the intestine, which was 8 times as much as that of the control group after 24 hours of stimulation with 路Poly I: C. The results showed that AJLEAP-2 played an important role in the process of anti-bacterial infection of Japanese eel (Anguilla japonica).
【作者单位】：集美大学水产学院;鳗鲡现代产业技术教育部工程研究中心;中国科学院水生生物研究所;福建省海洋生物资源开发利用协同创新中心;
【基金】：国家自然科学基金(31172438和U1205123) 福建省自然科学基金(2012706008,201311180002和2014705042) 教育部留学回国人员科研启动金资助~~
【分类号】：S917.4

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