抑制消减杂交(SSH)及其在鱼类基因克隆中的应用

Suppression subtractive hybridization and its application to fish gene cloning

  • 摘要: 和其它脊椎动物一样,在鱼类的生命进程中,某种特定类型的细胞中只有比例约为10%~15%的基因表达.由于这些基因的特异性决定了从受精卵到机体死亡的整个生命过程,所以这些基因的时空表达是受到严格调控的.在不同发育阶段,不同生理状态和不同类型的细胞中,表达的基因各不相同.因此,比较两种不同类型细胞中基因表达的差异,可以探究表型的遗传原因,了解生命过程的基本信息,从而找到与发育、疾病等相关的基因.

     

    Abstract: There are 10 percent to 15 percent genes expression in certain cells during the life time of f ishes like other vertebrates. The genes were different at different development stage, under different physiological conditions, and in different kinds of cells. So comparing the differences of gene expression in different cells can help us understand the genetic nature of phenotypic dif ferences, and understand the basic information of life period, and f ind the genes in relation to development and diseases, and f inally benefit mankind. Several methods were developed to clone differential expression gene in recent years. They are subtractive hybridizat ion ( SH) , differential display ( DD) , representional dif ference analysis (RDA) , and so on. These methods all have postive influences on cloning special genes, but they all have some defects, such as higher false..positive, lower replication, lower sensitive and diff iculty to manipulate. Suppression subtract ive hybridization ( SSH) was developed by Diatchenko et al in 1996. SSH was based on suppression PCR and combines normalization and subtraction in a single procedure. It is a more effective and convenient method than all others mentioned above. The principle and the rules of manipulation of SSH in detail was illuminated and the novel genes cloned by SSH was listed. They are immune related genes and reproduction and development related genes. The reproduction and development related genes are as follows: ZP3, Cyclin A2, CB102, YA2, FSTRAP. The immune related genes are as follows: NKEF( natural killer enhancing factor ) , CC chemokine, CXCR1, CXCR2, CXCR4, AIF..1( allograft inf lammatory factor..1) , IL..1..( inteleukin..1) , Fc..RI..( .. submit of high aff inity Fc receptor for IgE) , SSA( serum amyloid A) , LECT2( leucocyte cell..derived hemotaxin 2) , GMF..( glia maturation factor..) , CD45, Lysozyme C, PBEF ( Pre..B cell enhancing factor) , C..type lectin, PTX( Pentraxin) , IL..1RII, IL..8..like CXC chemokine, TF( tissue factor) , trout chemokine 2, TNF decoy receptor, M17. Some subtracted cDNA libraries were also built by SSH method.

     

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