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目的:探讨DAZ基因突变对男性生殖的影响及表型特征。方法:筛选无明显诱因的男性非梗阻性无精及少精患者,首先排除染色体核型异常病例,进一步在AZFa、AZFb、AZFc和AZFd区域均匀选择缺失高发位点排除缺失,设130例正常男性与10例女性做对照,对DAZ1～4基因进行突变性研究。包括DAZ1～4共缺失性研究和基因内点突变研究。选择同时存在于DAZ1～4基因内部的STS位点sY587,结合其PCR扩增产物中DraI酶切位点的差异判断DAZ1～4共缺失情况;选择DAZ1～4高度同源保守的1～6外显子、接头以及上下游调控区域,采用PCR-SSCP、限制性酶切(RFLP)、PCR产物测序以及特异性基因型nest相结合方法对外显子进行突变检测;设sY14(SRY)基因作为阳性内参照。结果:经筛选得到80例先天非梗阻性无精与20例少精病例,所有患者及男性对照sY14(SRY)检测均为阳性,女性对照均为阴性;共发现16例DAZ共缺失患者,发生频率为20%;无精患者11例,DAZ1～4共缺失及DAZ1/DAZ2共缺失分别为1例和10例;少精患者5例,DAZ1～4共缺失及DAZ1/DAZ2共缺失分别为2例和3例;分别在3个不同患者的DAZ1的第2、4内含子发现3个变异位点,DAZ基因的调控序列、外显子及接头处未发现突变,而在130例正常对照中未见缺失及变异发生。结论:DAZ基因缺失是引起男性生殖异常的主要原因;共缺失的部位及长度与表型的严重程度未见相关性;基因组比较确定DAZ1第2内含子第64碱基处发杂合变异G→A为新的变异,是否是新的SNP位点、是否能够造成无精或少精需要进一步研究证实;DAZ基因突变发生情况也有待进一步探讨。
Objective: To investigate the effect of DAZ mutation on male reproductive performance and phenotypic characteristics. Methods: To screen non-obstructive azoospermia and oligospermia in male patients with no obvious predisposition, first to exclude the cases of chromosomal abnormalities in karyotype, and to further eliminate the deletions in AZFa, AZFb, AZFc and AZFd regions, In comparison with 10 women, DAZ1 ~ 4 genes were mutated. Including DAZ1 ~ 4 co-deletion studies and intragenic mutations. The STZ locus sY587, which is also located in DAZ1 ~ 4 gene, was selected to determine the common deletions of DAZ1 ~ 4 in combination with the differences of DraI restriction sites in the PCR products. The highly conserved DAZ1 ~ 4 Exon, linker and upstream and downstream regulatory regions were detected by polymerase chain reaction-restriction fragment length polymorphism (PCR-SSCP), restriction endonucleases (RFLP), sequencing of PCR products and specific genotype nest. The sY14 (SRY) Internal reference. RESULTS: Eighty cases of congenital non-obstructive azoospermia and 20 cases of oligozoospermia were screened. All patients and male controls were positive for sY14 (SRY) test and negative for female controls. A total of 16 patients with DAZ co-deletion were found Frequency of 20%; 11 cases of azoospermia, DAZ1 ~ 4 total loss and DAZ1 / DAZ2 co-deletion were 1 and 10 cases respectively; 5 cases of oligozoospermia, DAZ1 ~ 4 total loss and DAZ1 / DAZ2 co-deletion were 2 Three cases were found in the first and second intron of DAZ1 in three different patients. No mutation was found in the regulatory sequence, exon and linker of DAZ gene. In 130 normal controls No missing and mutation occurred. Conclusion: The deletion of DAZ gene is the main cause of male genital abnormalities. The total missing part and length have no correlation with the severity of phenotype. The genome-wide comparison confirmed the mutation of the 64th base of DAZ1 intron 2 → A is a new mutation, whether it is a new SNP site, whether it can cause azoospermia or oligospermia need further study confirmed; DAZ gene mutations also need to be further explored.