摘要
目的 :制备人β2 m转基因小鼠 ,研究HLA B2 70 4的表达。方法 :应用显微注射将人 β2 m基因注入C5 7BL/6×昆明鼠和昆明鼠×昆明鼠F1代受精卵 ,出生动物及其后代经PCR筛选 ,采用斑点杂交和Southern杂交对阳性鼠基因组DNA标本行进一步鉴定和测定整合拷贝数 ,利用RT PCR检测阳性鼠中人 β2 m转基因的表达。结果 :6只原代仔鼠及 7只它们的下一代 (F1)带有人 β2 m基因。由微注射基因后移卵出生的 86只小鼠中 ,C5 7BL/6×昆明鼠杂交仔鼠 35只 ,其中 4只阳性 (11.4% ) ,昆明鼠×昆明鼠杂交仔鼠 5 1只 ,其中 2只阳性 (3.9% ) ,含有人 β2 m基因的原代鼠×昆明鼠杂交仔鼠 2 0只 ,其中 7只阳性。整合的转基因均为单拷贝。Southern杂交证实上述阳性鼠确有转基因整合。阳性鼠的皮肤、结肠、睾丸和脾脏组织中均有人 β2 m转基因mRNA的表达。结论 :在转基因动物制备中 ,C5 7BL/6×昆明鼠F1代明显优于昆明鼠×昆明鼠F1代。与人HLA B2 70 4基因相比 ,人 β2 m基因不易整合 ,其整合率与整合拷贝数均较低。得到的人 β2 m转基因小鼠能够将人 β2 m基因传给下一代并可与人HLA B2 70 4转基因鼠交配。
Objective: To produce human β 2 microglobulin transgenic mice for the study of human HLA B2704 gene related to arthrospondylosis in Chinese. Methods: Human β 2m gene was introduced into fertiliged eggs of C57/6×Kunming and Kunming ×Kunming F 1 mice by microinjection. By PCR, the founder mice and their progeny were screened for integration of the human β 2m transgene into the mouse genomes which were further confirmed by Southern blotting. The copy number of transgenes was determined by dot blotting. The expression of human β 2m transgene in mice at the mRNA level was analysed by RT PCR. Results: Six founder mice and seven founder F 1 contained human β 2m genes. Four of 35 (11.4%) hybrid mice from C57BL/6×Kunming and two of 51 (3.9%)hybrid mice from Kunming×Kunming background carried human β 2m genes. Statistical analysis showed that there was significant difference of the integration rate between the two groups ( P < 0.05 ), and seven of 20 hybrid mice from founder mice containing human β 2m gene ×Kunming background carried human β 2m gene. The transgene was single copy in all the founder mice and founder F 1. The human β 2m gene was expressed at the mRNA level in skin, colon, testicle and spleen tissues of the transgenic mice. Conclusion: Human β 2m transgenic mice have been obtained. For the production of transgenic mice, C57BL/6×Kunming F 1 is better than Kunming×Kunming F 1. Compared with the HLA B2704 gene, the integration of human β 2m gene into mouse genome is more difficult
出处
《北京大学学报（医学版）》
CAS
CSCD
北大核心
2001年第4期366-369,共4页
Journal of Peking University:Health Sciences
基金
国家自然科学基金 (394786 94)资助&&
