β1,4-半乳糖基转移酶-I表达质粒在许旺细胞中的转染与鉴定

Construction and transfection of expression plasmids of β1,4-galactosyltransferase I in Schwann cells

目的 构建 β1,4 半乳糖基转移酶 I(β1,4 galactosyltransferaseI,β 1,4 GalT I)表达质粒 ,并将其转染到许旺细胞中 ,为探讨周围神经许旺细胞表达 β 1,4 GalT I的生物学意义提供研究基础。 方法 构建 β 1,4 GalT I表达质粒 ,并将其转染到分离培养的许旺细胞中 ;用特异识别 β1,4 半乳糖苷键的蓖麻凝集素 I(RicinusCommunisAgglutinin I,RCA I)免疫组织化学方法和NorthernBlot鉴定转染情况。结果 通过酶切鉴定和测序分析 ,实验成功构建 β 1,4 GalT I表达质粒。将构建的表达质粒转染到许旺细胞中 ,经鉴定表明 ,转染的表达质粒在许旺细胞中能表达 β 1,4 GalT I,并随转染浓度增高而表达增加。 结论 β 1,4 GalT I在许旺细胞中的转染和表达 ,对分析许旺细胞表达 β 1,4 GalT I的意义如对其自身增殖的影响和对神经元生长作用 。

Objective In order to study the biological effects of β1,4 galactosyltransferase I(β 1,4 GalT I)expressed in Schwann cells. The expression plasmids of β 1,4 GalT I were constructed and transfected into Schwann cells in this study.Methods The whole gene sequence of β 1,4 GalT I was obtained by RT PCR from total RNA of mouse brains, cloned into the pGEM T vector and subcloned into the expression plasmid pcDNA3.1. The constructed expression plasmids were transfected into purified Schwann cells and certificated by lectin histochemistry with Ricinus Communis Agglutinin I (RCA I) and Northern Blot analysis. Results It was found that the expression plasmids of β 1,4 GalT I were constructed and transfected into the Schwann cells successfully.Conclusion The Schwann cells transfected with the expression plasmids of β 1,4 GalT I will help to study the effects of β 1,4 GalT I on such as the proliferation of Schwann cells and the growth of neuron.