论文部分内容阅读
为筛选得到罗非鱼白细胞免疫相关基因,促进鱼类免疫防治进展,本研究应用抑制性差减杂交(SSH)技术成功构建了罗非鱼链球菌疫苗免疫前后正、反两个白细胞cDNA差减文库,富集了免疫过程中表达谱发生变化的白细胞基因。随机挑选阳性克隆PCR检测,显示插入片段在300～750bp。正反文库各随机挑选300个阳性克隆(共600个克隆)进行测序及EST初步分析。正库筛选得到18个免疫期间表达丰度上调基因,其中2个与罗非鱼已知基因具有相似性,6个与其他鱼类已知基因相似,其他6个与其他物种已知基因具有相似性,4个为未知新基因。负库筛选得到22个免疫期间表达丰度下调的基因,其中4个与罗非鱼已知基因具有相似性,5个与其他鱼类已知基因相似,6个与其他物种已知基因具有相似性,7个为未知新基因。随着序列的进一步分析及利用RACE等技术得到罗非鱼抗菌或免疫相关全基因,深入开展鱼类功能基因研究奠定良好的基础。
In order to screen the leukocyte immune-related genes of tilapia and promote the progress of immunization of fish, the positive and negative leukocyte cDNA subtractions library of Streptococcus tilapia before and after immunization with Suppression subtractive hybridization (SSH) , Enriched leukocyte genes that change in the expression profile during immunization. Randomly selected positive clones PCR test showed that the inserted fragment at 300 ~ 750bp. The positive and negative libraries were randomly selected 300 positive clones (a total of 600 clones) sequencing and EST preliminary analysis. In the positive database, 18 genes were up-regulated during the immunization, two of them were similar to the known genes of tilapia, 6 were similar to other known genes of fish and the others were similar to the known genes of other species Sex, 4 are unknown new genes. Negative library screening resulted in 22 down-regulated genes during immunization, of which 4 were similar to the known genes of tilapia, 5 were similar to other known genes and 6 were similar to other known genes Sex, 7 unknown new genes. With the further analysis of the sequence and the use of RACE technology to obtain tilapia antibacterial or immune related genes, in-depth study of fish functional genes laid a good foundation.