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栉孔扇贝foxl2基因在卵子发生中的必要性
刘晓玲, 刘 军, 王振东, 王振华, 李彦伸, 崔龙波
烟台大学 生命科学学院
摘要:
采用RNAi技术研究了栉孔扇贝(Chlamys farreri)foxl2基因在卵子发生和卵巢功能维持中的作用。使用体外合成的栉孔扇贝foxl2双链RNA(dsRNA), 以每只50 μg/次的剂量注射进闭壳肌中, 连续注射2次(第一次注射后7天, 再进行第二次注射); 以注射相同体积PBS(相同注射方法)的扇贝作为阴性对照组, 以不注射扇贝作为空白对照组。qRT-PCR检测干扰组扇贝卵巢中的foxl2 mRNA表达量较空白对照组下降了62%, Western blotting检测该蛋白在卵巢中的含量也明显下降, 显示靶基因的表达水平被有效地敲降。组织学观察发现, 栉孔扇贝foxl2干扰后的卵巢中卵母细胞形态异常、细胞核固缩, 卵子发生明显受阻。表明该基因在栉孔扇贝卵子发生中起重要作用。
关键词:  栉孔扇贝  卵巢  卵子发生  foxl2  RNAi
DOI:10.11759/hykx20180323001
分类号:
基金项目:山东省高等学校科技计划项目(J15LE05); 山东省现代农业产业技术体系建设专项资金(SDAIT-14)
Necessity of foxl2 in the oogenesis of the scallop Chlamys farreri
LIU Xiao-ling, LIU Jun, WANG Zhen-dong, WANG Zhen-hua, LI Yan-shen, CUI Long-bo
Abstract:
This study elucidates the foxl2 function in the oogenesis and ovarian maintenance of the scallop Chlamys farreri. We synthesized the C. farreri foxl2 dsRNA in vitro and then injected into the adductor muscle with 50 μg dsRNA for each scallop. The injection was administered two times; the second injection was on the 7th day after the first injection. The scallops injected with the PBS solution using the same volume and method were considered the negative control, and the scallops without any injection were the blank control. qRT-PCR revealed a 62% decline in the foxl2 mRNA level in the ovaries of RNAi scallops and an apparent reduction in the FOXL2 protein content in the RNAi ovaries, suggesting the effective knockdown in the foxl2 mRNA level. Furthermore, histologically, the abnormal oocyte morphology, enhanced eosinophilia of the cytoplasm, and nucleus pulsation were observed in the ovaries after RNAi, suggesting that oogenesis was apparently blocked. This study establishes that foxl2 plays a vital role in C. farreri oogenesis.
Key words:  scallop  Chlamys farreri  ovary  oogenesis  foxl2  RNAi
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