| Infertility is a disease which affects human existence and reproduction. According to the statistics by relative department, there are 10% of infertility in the breeding aged couples in our country. Among which 50% are aroused by male infertility. In the research on the pathogeny of infertility, 30%plus of patients have no clear reason, part of them may be the problems on fertilization. The binding and fusion between sperm membrane and egg are the integral part of fertilization and sperm and egg membrane proteins are the key material basis and core part to hold this key. In the research of sperm membrane proteins genes, we found the genes of ADAM family and CRISP1 family are the most probable related gene for the fertilization. Among which, the ADAM family gene is with biggest possibility.ADAM family is a group of newly identifed transmembrane glycoproteins which anchored in the cell membrane. This family has distinct structural characteristics. There are some conservative potential functional areas, which contain signal peptide, prodomain, metalloprotease domain, disintegrin domain, cysteine-rich region, repeated epidermal growth factor domain, transmembrane region and cytoplasmic tail. It has many functions, including the combination of sperm and egg, cell fusion and differentiation, tissue or organ occurrence and regeneration, the activate and release of bioactivation factors, and signal conduction. From them, the studies on function of combination of sperm and egg are receiving more and more attention. Among which, more are on three types of genes: ADAM1, ADAM2, ADAM3, the research on new gene ADAM32 is still vacant both home and abroad, but ADAM32 and ADAM2 are recognized the most functional gene based on structure, so the research on ADAM32 is very important.This research applies recycling medical confirmed thinking, first accumulated the clinical materials, and had analysis on the possibility between condition of semen and being pregnant with SPSS analytical software.. According to WHO diagnosis flow chart, we made statistics on infertility pathogeny, the relativity between index of semen and being pregnant, and concurrently set up data model, which is used to have some forecast on pregnant possibility of infertile patients, and thus provide guidance on ART choices. The research result shows that the pathogeny of infertility is unknown reason at most, then is varicocele, which indicates the research on infertility needs more working. Statistics results on relativity for index of semen show that age is inverse correlation, the semen amount, sperm density and sperm vitality are positive correlation, among which, the A type vitality of sperm percentage has the largest correlation. Whereas tradional therapy, anti-sperm antibody has no correlation with pregnant. This shows that there is no efficiency method to treat infertility now. so ART will be the development trend for treating infertility. After setting up data modal, we had some forecast on being pregnant possibility in infertile patients. We found the average correct rate to be 78.9% and among which, the correct rate for forecast will-not-happen is even higher.From clinical material statistics, we found unknown reason for infertility had the biggest percentage, and sperm is the key for pregnant. So we apply RT-PCR method to check the ADAM family and CRISP 1 family gene expression in the human mature ejaculation sperm. Experimental results show that among the chosen ADAM family'genes (ADAMI, ADAM2, ADAM3, ADAM7, ADAM15, ADAM32), only AD AMI 5 has no expression and the others all have expression on ejaculation sperm; the ARP gene, which is considered as CRISP 1 family gene, has no expression either; and the PRM-2 gene, which is considered as sperm mature symbol gene, has expression on the all ejaculation sperm samples. And thus explain!? mature sperm has mRNA expression; only one part of ADAM genes exists on sperm, and is related with the fertility; ARP gene has no expressiom, whether it is related with fertility or not needs further discussion; PRM-2 gene is also considered as sperm mature symbol.To explore on unknown reason infertile patients whether there exists lack of ADAMgene or not, we apply RT-PCR method to check from normal group semen, infertile group semen, and aspermia group semen in order to know the ADAM gene expression ( all the chosen ADAM genes are expression on normal mature sperm). The checking result indicates that although on statistics, there are no remarkable difference on gene expression between infertile group and normal group, there do exists ADAM gene lacking, especially two or three kinds of ADAM genes lacking at the same time, which indicates ADAM genes lacking is probably the reason for infertility. In the experiment, we found ADAM 10 exists in semen of aspermia group, which indicates this gene not only exists on sperm, but also has expression on the cells; PRM-2 has no single expression in aspermia group, which indicates again that PRM-2 can be used to as symbol gene of sperm.To further investigate on time and location of ADAM gene happening, we apply again RT-PCR method on checking the ADAM gene, ARP gene and PRM-2 gene expression on testicle and epididymis in 4 groups including foetus, children, grown-ups and old men. The experiment result shows that the ADAM genes, ARP and PRM-2 gene all had no expression on testicle and epididymis in foetus group. Expression only found in infants after birth, among which the ADAM1, ADAM 10 and ADAM32 are the first to have expression, which indicates these genes have relation of the formation of testicle and spermatogonia which already exist on original times of sperm; ADAM 3 is the first one have expression on epididymis of children; ADAM7 and ARP only have expression on epididymis of grown-ups, this indicates these genes have epididymis specificity, and have relation with epididymis maturity. With the age increasing, some genes may be lost, like ADAM 10 on testicle, ADAM7 and ARP on epididymis. ADAM 15 gene has no expression on sperm, testicle, epididymis, which indicates this gene may have no close relation with fertility.Based on the above experiment results, we found ADAM32 is a kind of gene having close relation with reproduction. To further investigate and prove the function characteristic of this newly-found gene, we cloned out ADAM32 gene from the human mature ejaculation sperm. We apply RT-PCR method for dividing segment to get parts ofgene, then subcloned into pMD18-T, transfected, chosen of clones, gotten from vector, PCR, cut from gel, and gotten the target segment genes, subsequently, connected gene by TP-PCR, synthesized fusion gene, inserted into retrovirus vector pGEZ-Term, cotransfected into the package cell 293T, infected L929 cells, at last we get the stabilized expression of ADAM32 transfected cells. Research result shows that the method of segment gene cloned and TP-PCR gene connected is a good and efficient way for cloning bigger gene( above 2000bp plus). For checking of new gene without mono-cloning antibody, we apply the specially-marked intermediate plasmid pDsRed-Nl, by digesting with the restriction endonucleases, and synthesized fusion gene, then found indirect expression on transfect gene cell. The result is clear and reliable. This provides a reliable method to symbolize new no mono-cloning antibody cells. Stably expressed the transfected gene cells can provide the material condition for further immuring to BALB/c mouse, getting anti-ADAM 32 mono-cloning antibody, and researching the biology function of ADAM32.Summing up the above, we apply recycling medical confirmed thinking, from clinical materials, do scientific and objective analysis on data, check gene expression of ADAM family, CRISP 1 family and PRM-2 , which are related with reproduction, based on the different times of matured sperm, testicle, epididymis. We get the expression of those genes and results of relationship with infertility. These results have some creativity, especially the new gene ADAM32 successful structuring for cloning and transfected cells from human matured ejaculation sperm have never been published home and abroad, so it has very creative and advanced, which will also set up a good material basis for research on the mystery of reproduction.
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