Establishment of apoE Gene Knockout Model on SD Rat Using Talen Method

doi:10.3969/j.issn.1674-5817.2015.04.003

Abstract

Abstract: Objective To establish the apoE gene knockout-rat model using Talen method, and provide a basis for further study on the function of the apoE gene, the pathogenesis and treatment of atherosclerotic disease. Methods The Talen sequences were designed and synthesized for the rat apoE gene, and then the linearized and purified fragments were injected into fertilized eggs of SD rat through microsopic injection. Positive rats were detected by PCR-sequencing and TA clone-sequencing. The homozygous type offspring were obtained by siblings mating, and the genotype was identified. Western blot was used to detect the expression level of ApoE protein in some tissues of apoE^-/- rat. The plasma levels of total cholesterol and triglyceride levels have also been detected. The expression of ATP binding cassette transporter A1 (ABCA1) which was related to the lipid metabolism was detected by reverse transcriptase polymerase chain reaction (RT-PCR). Results Sequencing analysis indicated that there had a deletion of 1 bp in the second exon, resulting in open reading frame shift mutation in the rat, and obtained the apoE^-/- rats. Western blot test showed that no expression of ApoE protein in the heart, liver, kidney, brain and ovary of apoE^-/- rats, and the apoE gene was successfully knocked out. The total cholesterol and triglyceride levels of apoE^-/- rats were higher than that of wild type SD rats, and the total cholesterol levels have the significantly different (P<0.05). The expression of ABCA1 in the liver of the apoE^-/- rats was decreased, which may play a role of AS promoting. Conclusions apoE^-/- rats were obtained by Talen method. The apoE gene has been knocked out, and the rats can be considered as the model rats showed hyperlipemia and atherosclerosis.

Key words: Talen, Apolipoprotein E (ApoE), Gene knockout, Rat

CLC Number:

Q95-33

XU Ying-qi, LI Xiao-jing, YANG Wei, ZHOU Sheng-lai, YU Yang, WANG Wei, ZHANG E-nuo, ZHAO Wen, ZHANG Mei-ying, GUO Da-yong, WANG Hong-yu, ZHENG Zhi-hong. Establishment of apoE Gene Knockout Model on SD Rat Using Talen Method[J]. Laboratory Animal and Comparative Medicine, 2015, 35(4): 277-282.

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