【摘要】：對家蠅GNBP3基因進行克隆及生物信息學分析,并對該基因在感染白色念珠菌Candida albicans后的表達情況進行研究。從構(gòu)建的家蠅Musca domestica幼蟲c DNA質(zhì)粒文庫中篩選到GNBP3基因,克隆并運用生物信息學方法對該基因及其編碼蛋白進行預(yù)測和分析。白色念珠菌注射感染家蠅幼蟲并收集標本,逆轉(zhuǎn)錄,通過熒光定量PCR檢測感染樣本中GNBP3基因的表達情況,結(jié)果進行統(tǒng)計學分析。研究結(jié)果表明,GNBP3基因ORF全長1473 bp,編碼490個氨基酸,理論分子量為55.8 k Da,等電點為6.85;感染后不同時間點,感染組與對照組比較,在3 h、12 h、36 h、48 h GNBP3 mRNA的表達明顯增高,兩組比較有顯著性差異(P0.05),而24 h表達差異性最小;感染后不同組織中感染組與對照組比較,3 h和12 h時,在體壁、血淋巴、脂肪體中的表達量升高,具有顯著性差異(P0.05);24 h時,在血淋巴中的表達量較對照組高,而在唾液腺和脂肪體中的表達量呈下降趨勢,均具有顯著性差異(P0.05);48 h,在血淋巴和脂肪體中表達量較對照組高,在中腸的表達量較對照組低,均具有顯著性差異(P0.01)。成功克隆GNBP3基因且在家蠅感染白色念珠菌后GNBP3的表達水平隨時間的推移呈現(xiàn)先升高后降低再升高的趨勢,在各組織中以在免疫組織(血淋巴、脂肪體)中的表達顯著增高,推測其在真菌識別過程中發(fā)揮潛在作用。
[Abstract]:The GNBP3 gene of Musca domestica was cloned and analyzed by bioinformatics, and the expression of the gene was studied after infection with Candida albicans of Candida albicans. The GNBP3 gene was screened from the constructed plasmid library of Musca domestica larva c DNA of housefly, and the gene and its encoded protein were predicted and analyzed by bioinformatics. Candida albicans were injected into housefly larvae and samples were collected. The expression of GNBP3 gene in infected samples was detected by fluorescence quantitative PCR. The results were statistically analyzed. The results showed that the ORF of GNBP3 gene encoded 490amino acids with a theoretical molecular weight of 55.8k Da, isoelectric point of 6.85.The expression of GNBP3 mRNA in the infected group was significantly higher than that in the control group at 3 h, 12 h, 36 h and 48 h after infection. There was significant difference between the two groups (P0.05), but the expression difference at 24 h was the least, and the expression level in the body wall, hemolymph and fat body increased at 3 h and 12 h after infection (P0.05), and there was significant difference at 24 h after infection (P0.05). The expression level in hemolymph was higher than that in control group, but the expression level in salivary gland and fat body showed a decreasing trend (P0.05) for 48 h. The expression level in hemolymph and fat body was higher than that in control group, and the expression level in midgut was lower than that in control group. There was significant difference between the two groups (P0.01). GNBP3 gene was cloned successfully and the expression level of GNBP3 increased first and then decreased and then increased after infection with Candida albicans in housefly, and increased significantly in immune tissues (hemolymph, fat body). It is speculated that it plays a potential role in the process of fungi recognition.
【作者單位】： 貴州醫(yī)科大學基礎(chǔ)醫(yī)學院;畢節(jié)市第一人民醫(yī)院;貴州省疾病預(yù)防控制中心;貴州博康生物工程有限公司;
【基金】：國家自然科學基金(81360254) 貴州省科技廳省校合作項目(黔科合LH字[2014]7076號) 貴州省衛(wèi)生計生委科學技術(shù)項目(gzwjkj2014-2-100) 貴州省科學技術(shù)基金(黔科合J字[2013]2042號)
【分類號】：Q78;Q963

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