P210T315I-BCR/ABL转基因小鼠模型的建立与鉴定
Generation and identification of P210T315I-BCR/ABL transgenic mice
目的 建立P210T315I-BCR/ABL转基因小鼠模型,探讨慢性髓性白血病(CML)的发病机制和筛选抗T315I型CML的有效治疗药物.方法 构建APN/CD13启动子与增强子调控P210T315I-BCR/ABL基因和eGFP基因同时表达的转基因载体,通过显微注射法将构建好的载体注射到C57小鼠的单细胞受精卵中,用PCR方法鉴定转基因鼠首建鼠及其子代基因型,用荧光显微镜和RT-PCR方法检测转基因小鼠外周血中eGFP和BCR-ABL基因表达,经血常规、外周血和骨髓涂片、组织病理学检测转基因小鼠CML表现.结果 共得到3个同时表达eGFP和BCR-ABL基因的转基因小鼠品系,转基因小鼠2月龄时WBC、PLT等较对照组明显升高,以后逐月升高,6月龄时,WBC最高达23.9×109/L,中性粒细胞百分比最高达74.6%,PLT最高达4 136×109/L,外周血和骨髓涂片中可见少量原始粒细胞,动物死亡后病理解剖发现,脾脏肿大,脾脏中可见少量白血病细胞浸润.结论 成功建立了P210T315I-BCR/ABL转基因小鼠模型,为研究CML的发病机制和筛选抗T315I型CML的有效治疗药物奠定基础.
更多Objective To construct the P210T315I-BCR/ABL transgenic mice model.Methods The transgenic vector in which the P210T315I-BCR/ABL gene and eGFP gene was derived by APN/CD 13 promoter was constructed and microinjected into the single-cell fertilized eggs of C57 mice.Transgene integration was conformed by PCR genotyping and P210T315I-BCR/ABL expression levels was evaluated by RT-PCR.The CML phenotype was confirmed by blood routine examination,Wright' s staining for peripheral blood and bone marrow smears,HE staining for organs oftransgenic mice.Results Three transgenic mice lines with high expression of P210T315I-BCR/ABL gene and eGFP gene was selected.Compared with the wild type mice,the levels of WBC,platelet and neutrophil granulocyte of transgenic mice began to increase gradually at 2 months,and increase to 23.9× 109/L,4 136× 109/L,and 74.6% respectively at 6 months.The remarkable hyperplasia of granulocytes was seen in the peripheral blood and bone marrow smears with splenomegaly infiltrated by leukemic cells.Conclusion The P210T315I-BCR/ABL transgenic mice was constructed and provided a model to explore the mechanism of T315I CML and screen out the drug for T315 CML patient.
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