慢病毒载体介导的转人ahsp基因的β^(654)地中海贫血小鼠的制备

Generation of Human ahsp Transgenic β^(654)-Thalassemia Mice Mediated by Lentiviral Vector

目的:经慢病毒载体介导,制备转人α血红蛋白稳定蛋白基因(ahsp)的β654地中海贫血小鼠。方法:用巢式PCR从人血DNA中获得人ahsp基因,构建含有人ahsp基因的慢病毒载体,制备假病毒,通过卵周隙显微注射手段将其导入β654地贫小鼠的受精卵,经移植至假孕母鼠输卵管,最终孕育出转人ahsp基因的β654地贫小鼠;分析小鼠体内外源ahsp基因的表达情况及其遗传稳定性。结果:共获得了8只人ahsp阳性小鼠,转基因阳性率为32%(8/25),其中3只同时具有β654突变基因;人ahsp基因在小鼠体内的表达水平维持在小鼠自身ahsp表达量的1%左右,且可稳定遗传至子代。结论:制备了转人ahsp基因小鼠,并可遗传至子代,为在个体水平上研究α血红蛋白稳定蛋白与β地贫之间的关系提供了工具。

Objective： To obtain human α-hemoglobin stabilizing protein（ahsp） transgenic β654-thalassemia mice. Methods： Human ahsp gene with its promoter was amplified by nested-PCR from human blood geoomic DNA, then it was cloned into a lentiviral vector to construct its pseudo virus. The human ahsp transgenic mice were subsequently generated by infecting the fertilized oocytes of β654-thalassemia mice with pseudo virus. The transcription of the human ahsp gene and its genetic satiability were investigated. Results： We obtained 8 human ahsp transgenic mice, among which, 3 carried β654 gene. mRNA quantity of human ahsp accounted for approximately 1% of that of mouse ahsp. The transgene could be in- herited to the next generation. Conclusion： Human ahsp transgenic mice can be obtained by the method described herein, and they can be useful for studying the relationship between α-hemoglobin stabilizing protein and β654-thalassemia.