CAJ | 학술논문

采用同源克隆和半定量RT-PCR方法,对虾夷马粪海胆Strongylocentrotus intermedius NLR家族基因片段进行了克隆和组织表达特征分析。将紫球海胆S. purpuratus的9组NLR家族聚类的编码区序列进行比对后,根据保守区域设计引物扩增虾夷马粪海胆中相应的基因,通过测序获得了9组不同NLR聚类基因片段,每组各10个测序结果,通过基因序列及氨基酸序列比对分析,发现存在长度及序列结果上的差异,这说明每组NLR家族聚类都存在多个家族成员。同时选取虾夷马粪海胆的不同组织提取RNA,反转录后使用同样的引物进行半定量RT-PCR分析,结果表明, NLR基因在虾夷马粪海胆的管足、肠、围口膜、体腔细胞、性腺中存在高效表达,且在围口膜及性腺中表达量最高,表明NLR基因在虾夷马粪海胆中普遍表达,并存在表达差异。研究表明, NLR基因在海胆免疫过程中起着重要作用。
채용동원극륭화반정량RT-PCR방법,대하이마분해담Strongylocentrotus intermedius NLR가족기인편단진행료극륭화조직표체특정분석。장자구해담S. purpuratus적9조NLR가족취류적편마구서렬진행비대후,근거보수구역설계인물확증하이마분해담중상응적기인,통과측서획득료9조불동NLR취류기인편단,매조각10개측서결과,통과기인서렬급안기산서렬비대분석,발현존재장도급서렬결과상적차이,저설명매조NLR가족취류도존재다개가족성원。동시선취하이마분해담적불동조직제취RNA,반전록후사용동양적인물진행반정량RT-PCR분석,결과표명, NLR기인재하이마분해담적관족、장、위구막、체강세포、성선중존재고효표체,차재위구막급성선중표체량최고,표명NLR기인재하이마분해담중보편표체,병존재표체차이。연구표명, NLR기인재해담면역과정중기착중요작용。
In this study, homologous cloning and semi-quantitative RT-PCR technologies were used to clone the fragments of NLR genes and to detect their expression patterns in tissues of sea urchin Strongylocentrotus intermedi-us. The primers were designed according to the homology alignment of NLR genes from purple sea urchin S. purpu-ratus, and 9 fragment groups of NLR genes were obtained from the sea urchin, by sequencing of 10 clones for each group. Then bioinformatic analysis of these genes was conducted by the method of Blast, finding the diversity of cDNA sequences and amino acid sequences of these NLR genes. Also, the RNA was extracted from different tissues in the sea urchin, and the expression of NLR genes was analyzed by semi-quantitative PCR in different tissues of the sea urchin after reverse transcription. The results showed that 9 fragment groups of NLR genes were all ex-pressed in coelomocytes ( CL ) , membrana peristomialis ( WK ) , intestines ( GT ) , tube-feet ( GZ ) , and gonad ( GD) at a high level, the maximum in membrana peristomialis and gonad, indicating that NLR gene family plays an important role in the congenital immune system of the sea urchin, with different expression. The findings will provide the foundation for further study of NLR genes in the sea urchin.