X连锁迟发性脊椎骨骺发育不良家系的早期产前基因诊断

Early prenatal diagnosis for a family affected with X-linked spondyloepiphyseal dysplasia tarda family

目的联合应用性别鉴定、连锁分析和致病基因检测三种方法对X连锁迟发性脊椎骨骺发育不良（spondyloepiphyseal dysplasia tarda，SEDL）家系进行产前基因诊断。方法收集1个SEDL家系的7位成员（先证者、3名携带者及3名健康成员）外周血2mL，抽取2位胎儿的羊水标本20mL。用聚合酶链反应扩增2位胎儿的SRY基因和AMEL基因进行性别鉴定；应用PCR扩增产物双向直接测序方法检测SEDL基因突变；利用与SEDL基因毗邻的微卫星遗传标记DXS16进行连锁分析。结果经SRY和AMEL基因性别鉴定2位胎儿均为男性，1位胎儿SEDL基因存在IVS2—2A→C突变，且具有与致病基因连锁的DXS16等位基因；另1名胎儿SEDL基因检测未发现上述突变，DXS16等位基因与呈野生型SEDL基因连锁。经随访，新生儿和引产胎儿的基因诊断结果与产前诊断一致。结论SEDL基因突变检测结合微卫星DXS16多态性连锁分析和性别鉴定可以准确有效地进行x连锁迟发性脊椎骨骺发育不良家系的产前诊断。

Objective X-linked spondyloepiphyseal dysplasia tarda （ SEDL ） is osteochondrodysplasia caused by mutations of SEDL gene, which usually onset in late childhood without systemic complications. In this study, we have provided prenatal diagnosis for an affected family with a combined strategy including direct sequencing, fetal-sex identification and microsatetlite linkage analysis. Methods Two amniotic fluid samples from carrier gravida and 7 blood samples from individuals in this SEDL pedigree were obtained. Genomic DNA was extracted from the samples using standard phenol- chloroform method. SRY and AMEL genes were employed to assess fetal sex. Microsatellite DXS16 was genotyped for linkage analysis. A pathogenic mutation of the SEDL gene was identified by bi-directionally direct sequencing of the third exon as well as its exon/intron boundaries. Results Two male fetuses were confirmed by fetal-sex assessment. The mutation of the SEDL gene was identified as a nucleotide substitution of the splice-aceeptor site in intron 2, IVS2--2A→C. DNA sequencing indicated that one fetus is hemizygote carrying the mutation, whilst another is not a carrier. Linkage analysis was identical with the sequencing results. Follow-up also confirmed the result of prenatal diagnosis. Conclusion Fetal-sex assessment combined with microsatellite linkage analysis and bi-directionally direct sequencing is a more accurate and ready strategy for prenatal diagnosis of families affected with SEDL.