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以 ｇｔ１１为载体，构建家兔睾丸文库。平板扩增６×１０～５ｐｆｕｓ，提取 ＤＮＡ作为ＰＣＲ模板。根据ＧｅｎＢａｎｋ已发表ｓｐ１０ｃＤＮＡ的同源序列，选择高度保守区域设计５＇端引物（ＳＰ５＇），另加与构建文库载体λｇｔ１１序列互补的正向（ＥＰ）或反向引物（ＲＰ）共同配对组成ＰＣＲ循环引物。利用ＳＰ５＇＋ＲＰ或ＳＰ５＇＋ＦＰ引物对扩增出ｓｐ１０ｃＤＮＡ的３＇端．根据３＇端的测序结果，设计ｓｐ１０的３＇端引物（ＳＰ３＇），利用ＳＰ３＇＋ＲＰ或ＳＰ３＇＋ＦＰ引物对扩增出ｓｐ１０ｃＤＮＡ的５＇端．应用该方法成功地克隆了家兔精子膜蛋白ｓｐ１０ｃＤＮＡ（ｒｓｐ１０），说明该方法是一种快捷可行的ｃＤＮＡ克隆方法。
Gt11 as a vector to construct a rabbit testis library. The plate was amplified 6 × 10 ~ 5pfus, DNA was extracted as PCR template. According to the homologous sequences of published sp10 cDNA in GenBank, 5 ’primer (SP5’) was designed by highly conserved regions and co-paired with forward primer (EP) or reverse primer (RP) complementary to the sequence of λgt11 Loop primers. The 3 ’end of the sp10 cDNA was amplified using SP5’ + RP or SP5 ’+ FP primer pairs. According to the sequencing results of the 3 ’end, the 3’ end primer (SP3 ’) of sp10 was designed and the 5’ end of the sp10 cDNA was amplified using the SP3 ’+ RP or SP3’ + FP primer pairs. This method was successfully used to clone rabbit sperm membrane protein sp10 cDNA (rsp10), indicating that this method is a fast and feasible cDNA cloning method.