本文采用RT-PCR和RACE法分离、克隆了翘嘴红鲌G6Pase催化亚基基因全长cDNA,共1900 bp[不含poly（A）],包括49 bp 5′非翻译区,1068 bp阅读框以及含Poly（A）信号AATAAA的778 bp 3′非翻译区[不包括Poly（A）]。阅读框共编码355个氨基酸...
详细信息
本文采用RT-PCR和RACE法分离、克隆了翘嘴红鲌G6Pase催化亚基基因全长cDNA,共1900 bp[不含poly（A）],包括49 bp 5′非翻译区,1068 bp阅读框以及含Poly（A）信号AATAAA的778 bp 3′非翻译区[不包括Poly（A）]。阅读框共编码355个氨基酸,分子量为39.89 ku。序列比对分析表明翘嘴红鲌G6Pase与斑马鱼G6Pase的相似性高达95%,与鼠、狗、人G6Pase的相似性为63%,和光滑爪蟾、金头鲷、河豚等G6Pase的相似性分别为69%、55%、76%,并具有G6Pase特有的3个保守基序。为了研究摄食以及饲料中碳水化合物对G6Pase的影响,使用实时定量RT-PCR分别测定了饲喂等能但不含碳水化合物和含23.98%碳水化合物的饲料的翘嘴红鲌肝脏G6Pase基因的表达水平,在使用上述饲料饲喂8周后,禁食48 h,然后测定禁食和摄食后3、6、12、24 h G6Pase mRNA的表达量,结果显示摄食后12 h,两组G6Pase的表达明显增加,说明摄食影响G6Pase基因的表达,禁食和摄食后312 h,含糖组G6Pase基因的表达量为无糖组的24倍,这表明碳水化合物可以诱导G6PasemRNA的表达。
Sex determination is composed of somatic and germ-line sex differentiation hierarchies whose interaction is poorly understood. A single gene known to control somatic sex determination, the DM-domain containing （Doubl...
详细信息
Sex determination is composed of somatic and germ-line sex differentiation hierarchies whose interaction is poorly understood. A single gene known to control somatic sex determination, the DM-domain containing （Doublesex/Mab-3 DNA-binding motif） gene, is highly conserved across species. Vertebrate DMRT1 （DM-related transcription factor 1） expression occurs predominantly in the testis. Here, however, isolated two distinct DM-domain cDNA from Oreochromis aurea ovary and testis have been named DMRT4 （DM-related transcription factor 4） and DMRT1 by BLAST, respectively. Despite high homology in the DM-domain there is little similarity outside the *** better understand the structure, function, and possible roles of DMRT4 and DMRT1 as potential candidates for sex differentiation and sex determination, the intact regions encoding DMRT4 and DMRT1 obtained by PCR were sub-cloned into the vector pMAL-c2x and introduced into the Escherichia coli TB1 cell for efficient fusion expression. After purification and cleavage, DMRT4 and DMRT1 proteins were used to immunize adult rabbits following standard protocols. Consequently, it was found by using Western blot analysis that polyclonal antibodies against DMRT4 and DMRT1 had high specificity. The relative expression levels of DMRT4 and DMRT1 mRNA were determined by fluorescent Real-time RT-PCR in female and male Oreochromis aurea with 13-actin as the internal standard. DMRT1 was expressed only in testis, whereas DMRT4 was over expressed in the ovary, but in both female and male, a slight expression in the brain was also detected. Statistical analysis showed that in the brain, mean DMRT4 mRNA levels in female were significantly higher than in male. Meanwhile, the expression of DMRT4 and DMRT1 protein was also analyzed using the purified antibodies through Western blot and immunohistochemistry. It was found that DMRT4 was exclusively expressed in the ovary and DMRT1 in the testis. Study on DMRT4 and DMRT1 expression facilitate
暂无评论