建立一个带有JAK2-V617F、MPLW515L或CALR-Type I基因突变的骨髓增殖性肿瘤（MPN）移植小鼠模型，并建立一个系统的评估体系以验证模型构建的成功性。通过以下步骤获得小鼠骨髓c-kit+细胞：用颈椎脱位的方法杀死小鼠，分离股骨、胫骨和髂骨，收集骨髓细胞。与CD117磁珠孵育后将c-kit+细胞分选出来。构建小鼠原代突变细胞的方法如下：使用逆转录系统构建带有GFP标签的基因突变载体，并将逆转录病毒载体包装进Platinum-E细胞中，以获得病毒上清液，然后用其感染小鼠的c-kit+细胞。MPN小鼠模型的构建如下：在感染后收集含有突变基因的小鼠原代c-kit+细胞，然后通过尾静脉移植到同种的经致命剂量γ射线（8.0Gy）照射的雌性受体小鼠体内。MPN小鼠模型的评估如下：移植后，定期从尾静脉采集小鼠的外周血进行完全血细胞计数检测，评估脾脏大小和骨髓纤维化程度。从骨髓成功获得带有突变基因的小鼠c-kit+细胞。成功构建MPN小鼠模型：MPN移植小鼠的外周血细胞携带异源植入的GFP阳性细胞，白细胞（WBC）、血小板（PLT）和红细胞计数（HCT）均增加；移植小鼠体重减轻，饮水和摄食减少；进一步病理分析显示移植小鼠出现脾肿大和骨髓纤维化。这些结果表明MPN小鼠模型构建成功。根据三种MPN小鼠模型的共同和不同特点，总结了初步的评估体系以判断MPN小鼠模型构建的成功与否，主要包括以下指标，例如，小鼠外周血中GFP阳性细胞的比例；WBC、PLT和HCT；脾脏肿大程度和骨髓纤维化程度。利用逆转录系统成功建立了带有JAK2-V617F、MPLW515L或CALR-Type I基因突变的MPN小鼠模型，这为MPN发病机制和药物靶向治疗的研究提供了重要的实验动物模型。

To construct a myeloproliferative neoplasms (MPN) transplanted mouse model with JAK2-V617F, MPLW515L or CALR-Type I gene mutation, and establish a systematic evaluation system to verify the success of model construction.The bone marrow c-kit+ cells of the mice were obtained by the following steps: The mice were killed by cervical dislocation, the femur, tibia and ilium were separated, and the bone marrow cells were collected. The c-kit+ cells were sorted after incubation with CD117 magnetic beads. The method of constructing mouse primary mutant cells is as follows: A gene mutation vector with a GFP tag was constructed by the retroviral system, and the retroviral vector was packaged into the Platinum-E cells to obtain the virus supernatant, and then used it to infect the c-kit+ cells of mice. The MPN mouse model was constructed as follows: the mouse primary c-kit+ cells containing the mutant genes were collected after infection, and then transplanted them via the tail vein into the female recipient mice of the same species which were irradiated with a lethal dose of gamma rays (8.0 Gy). The MPN mouse model was evaluated as follows: After transplantation, the peripheral blood of the mice was regularly collected from the tail vein to perform the complete blood count test, and the size of spleen and the degree of bone marrow fibrosis were estimated.The mouse c-kit+ cells with the mutant genes were successfully obtained from the bone marrow. MPN mouse model was successfully constructed: The peripheral blood cells of the MPN-transplanted mice carried exogenous implanted GFP-positive cells, and the white blood cells (WBC), platelet (PLT) and hematocrit (HCT) were all increased; the body weight loss, and the water and food intake were reduced in the transplanted mice; further pathological analysis showed that the transplanted mice displayed splenomegaly and bone marrow fibrosis. These results suggested that the MPN mouse model was successfully constructed. According to the common and different characteristics of the three MPN mouse model, a preliminary evaluation system for judging the success of MPN mouse model construction was summarized, which mainly included the following indicators, for example, the proportion of GFP-positive cells in the peripheral blood of mice; WBC, PLT and HCT; the degree of spleen enlargement and the bone marrow fibrosis.The MPN mouse model with JAK2-V617F, MPLW515L or CALR-Type I gene mutation is successfully established by retroviral system, which can provide an important experimental animal model for the research of MPN pathogenesis and drug-targeted therapy.