MUTANT HEMOGLOBINS THAT ALLOW HBS TO SICKLE

允许 HBS 镰刀化的突变血红蛋白

• 批准号: 3362021
• 负责人: RAYMOND A POPP
• 金额: $ 15.76万
• 依托单位: UNIVERSITY OF TENNESSEE KNOXVILLE
• 依托单位国家: 美国
• 财政年份: 1989
• 资助国家: 美国
• 起止时间: 1989-07-01 至 1994-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3362021
• 关键词: chemical association; disease /disorder model; electrofocusing; genetically modified animals; hemoglobin Ss; laboratory mouse; microinjections; model design /development; mutant; respiratory oxygenation; sickle cell anemia; southern blotting

A colony of mice will be produced that has hemoglobin with oxygen association-dissociation properties that are similar to human sickle cell hemoglobin (HbS). This will be done by breeding a stock of mice that carry two mutations at the hemoglobin loci. Mice that are homozygous for the Hbag2 alpha-globin haplotype will be mated with mice that are homozygous for the Hbbs2 beta-globin haplotype. F2 progeny will be screened to identify mice that are doubly homozygous for the alpha- and beta-globin gene mutations. The hemoglobin of Hbag2/Hbag2;Hbbs2/Hbbs2 mice will have oxygen association-dissociation properties similar to that of Hbs. The mice will be called MHOAH. MHOAH mice will be made transgenic for the human alpha- and sickle cell or sickle cell Antilles beta- globin genes to produce a transgenic mouse model for sickle cell disease. The transgenes will be introduced into MHOAH mice directly by breeding with stocks of mice that carry the transgenes or directly by microinjecting minilocus constructs of DNA into the male pronucleus of fertilized eggs of MHOAH mice. Mice that carry the transgenes will be identified by Southern blotting. Transgenic mice that express high levels of the transgenes will be identified by electrophoresis of blood hemolysates. The oxygen association- dissociation and gelation properties of hemoglobins in MHOAH mice that express high levels of HbS or HbS Antilles will be studied. Transgenic mice that express high levels of HbS Antilles are very likely to be an animal model for sickle cell disease. HbS Antilles has a lower solubility and a lower oxygen affinity than HbS. About 60 percent of the HbS Antilles will be deoxygenated at 40 mm of Hg oxygen tension; only 25 percent of the high oxygen affinity hemoglobin of MHOAH mouse would be deoxygenated at the same oxygen tension. These conditions produce sickle cell disease in HbA/S Antilles heterozygotes and are likely to produce gelation of HbS Antilles and sickling of erythrocytes in transgenic MHOAH mice. The mouse model for sickle cell anemia would facilitate research on the pathophysiology of the disease and on the development and testing of anti-sickling drugs.

将产生一群有血红蛋白和氧气的小鼠 与人类相似的缔合-解离特性 镰状细胞血红蛋白（HbS）。 这将通过繁殖一只 在血红蛋白基因座上携带两个突变的小鼠。 对于Hbag 2 α-珠蛋白单倍型是纯合的小鼠将 与Hbs 2 β-珠蛋白纯合子小鼠交配 单倍型。 将筛选F2后代以鉴定 α-和β-珠蛋白基因突变的双重纯合子。 Hbag 2/Hbag 2; Hbbs 2/Hbbs 2小鼠的血红蛋白将有氧 缔合-解离性质与Hb相似。 的 小鼠将被称为MHOAH。 MHOAH小鼠将被转基因 人α-和镰状细胞或安的列斯β-镰状细胞 珠蛋白基因构建镰状细胞转基因小鼠模型 疾病 将转基因导入MHOAH小鼠 直接通过与携带转基因的小鼠进行繁殖 或直接通过将DNA的微轨迹构建体显微注射到 MHOAH小鼠受精卵的雄性原核。 携带病毒的老鼠 通过Southern印迹鉴定转基因。 转基因 将鉴定出表达高水平转基因的小鼠 通过血液溶血产物的电泳。 氧气协会- 小鼠血红蛋白的解离和凝胶化特性 表达高水平HbS或HbS的安的列斯群岛。 转基因小鼠表达高水平的HbS安的列斯群岛是非常 很可能是镰状细胞病的动物模型。 安的列斯群岛HbS 具有比HbS更低的溶解度和更低的氧亲和力。 关于 安的列斯群岛60%的HbS将在40毫米汞柱时脱氧 氧张力;只有25%的高氧亲和力 MHOAH小鼠的血红蛋白在相同的氧浓度下会发生脱氧 张力 这些情况下产生镰状细胞病的HbA/S 安的列斯杂合子和可能产生HbS凝胶化 转基因MHOAH小鼠红细胞的安的列斯群岛和镰状变。 镰状细胞贫血的小鼠模型将有助于研究 疾病的病理生理学和发展和 抗镰刀菌药物的测试。