联合运用四种技术进行血友病A的基因诊断

Gene diagnosis for hemophilia a using A combination of 4 techniques

目的 最大限度地提高血友病A(HA)患者及家系成员的基因诊断、携带者检出及产前诊断的可诊断率。方法 对于 2 6例HA患者和家系的女性家属首先采用长距离DNA扩增 (LD PCR)技术 ,直接检测是否为FⅧ基因倒位及其携带者 ;对于非倒位的HA家系依次采用BclⅠPCR/RFLP分析技术、基因内含子 13(CA) n、内含子 2 2 (GT) n(AG) n 二核苷酸重复序列多态性分析技术以及与FⅧ基因紧密连锁的可变串联重复序列多态性分析技术 (St14VNTR/PCR)进行间接诊断。结果 在 2 6个HA家系的 16个重型家系中查出 7个基因倒位 ,占重型HA的 43 8%。 19个非倒位HA家系 ,用上述三种间接诊断技术分别有 16、13及 17个HA家系可以作出诊断 ,可诊断率分别为 84 2 %、6 8 4%和89 5 % ,联合上述四种技术 ,对 2 6个HA家系全部作出了诊断。结论 联合采用四种基因诊断技术 。

Objective To improve the gene diagnosis,carrier detecting and prenatal diagnosis for hemophilia A families to a maximum. Methods Long distance PCR (LD PCR) was used to detect FⅧ gene inversion among 26 patients with severe HA and their female offspring to identify those with FⅧ gene inversion and its carriers. Three gene linkage analysis based upon PCR, BclⅠPCR/RFLP, St14 VNTR/PCR analysis and two simple dinucleotides repeats polymorphism analysis were used among 19 non inversion HA families for indirect diagnosis. Results Sixteen families with severe HA were detected by LD PCR among the 26 hemophilia A families. Seven were confirmed as with FⅧ gene inversion (43 8%). The diagnostic rates by the above mentioned three indirect diagnostic techniques among nineteen families without gene inversion were 84 2%, 68 4% and 89 5% respectively. Confirmed diagnosis was successfully made among all of the HA families by the four techniques. Conclusion Gene diagnosis and carrier detecting can be confirmed among almost all of the HA families can be diagnosed by the combined use of the four direct and indirect gene diagnostic methods.