猪精子和睾丸组织RNA的初步分析

Analysis of RNA in swine spermatozoa and testis

试验采用Trizol一次法提取猪精子和睾丸组织的总RNA,然后通过RT-PCR扩增的方法,检测精子和睾丸组织中Protamine-1(PRM-1)、Protamine-2(PRM-2)、Clusterin、Heat shock protein 90(HSP90)、C-myc基因的mR-NA是否存在;通过不同精子数量的使用,检测在本试验条件下扩增出清晰PRM-1带所需要的适宜精子数量;通过琼脂糖电泳检测精子与睾丸组织RNA的电泳特征。结果,猪睾丸组织中有PRM-1、PRM-2、Clusterin、HSP90、C-myc的mRNA,而射出的猪精子(上游处理)本试验只检测出PRM-1的mRNA;在使用RNA沉淀剂时,3.0×107～6.5×107精子可得到清晰的PRM-1条带;本试验首次得到猪精子总RNA的普通琼脂糖胶电泳图。初步研究的结果表明:射出猪精子中存在的RNA种类与睾丸组织中存在的RNA种类差异大;猪精子RNA的18S和28S片段与睾丸组织无明显差异。猪精子的RNA需要更多的研究。

In the present study,total RNA in swine spermatozoa and testis was extracted by Trizol. Transcript of Protamine-1（PRM-1）, Protamine-2 （PRM-2）, Clusterin, Heat shock protein 90 （HSP90）, C-myc in total RNA were subsequently detected and analysed by RT-PCR and PCR. Total RNA recovered was evaluated by agarose gel electrophoresis. The results revealed that transcript of PRM-1, PRM-2 ,Clusterin, HSP90,C-myc was detected in testis, while only transcript of PRM-1 were detected in swine spermatozoa. The best sperm number required for RNA extraction and detection of PRM-1 was 3.0 × 10^7-6. 5 × 10^7 under the use of RNA carrier. The results indicates that there was no significant difference between the bands of total RNAs from spermatozoa and testis. It is concluded that there are difference between kinds of RNA transcripts from swine spermatozoa and testis. No significant difference between bands of total RNA eleetrophoresis from swine spermatozoa and testis was observed in this study.