A preliminary study on prenatal diagnosis of congenital malformations using chromosome-specific variable number of tandem repeats (VNTR).

以pYNH24 （VNTR）为探针，采用Southern blot杂交技术对30例先天性畸形的DNA样本进行分析。2份样品检出异常条带。由于pYNH24位点定位在人类2号染色体上，我们进一步研究了先天性畸形与2q33-36位点γ -crystal Pst - i多态性的关系。多态性分布在2.1和1.7 kb的条带上。这些波段的频率如下：正常对照组为2.1/1.7（53%）和1.7/1.7(27%)，重度先天性畸形为2.1/1.7（7%）和1.7/1.7（73%）。这一结果提示可能存在一个与先天性畸形密切相关的基因座。从基因定位数据中，我们发现人类同源盒4基因簇存在于γ -结晶基因附近。因此，我们的研究重点是Hox - 4基因。同源盒基因是高度保守的基因，被认为在胚胎发生过程中编码了身体计划的程序。作为同源盒基因突变研究的第一步，我们分离了Hox 4A基因组DNA克隆并确定了约6000 bp的核苷酸序列。根据序列数据，合成PCR引物，对23例重度先天性畸形的dna样本进行PCR- sscp分析。我们在PCR-SSCP中未发现异常条带，现尝试在人类先天性畸形中检测Hox 4B基因异常。

DNA samples from 30 cases of congenital malformations were analyzed by Southern blot hybridization by using pYNH24 (VNTR) as a probe. Abnormal bands were detected in 2 samples. Since the locus of pYNH24 was mapped in human chromosome 2, we further investigated the association of congenital malformations and Pst I-polymorphism of gamma-crystalline which was mapped on 2q33-36. The polymorphism was found in 2.1 and 1.7 kb bands. The frequency of these bands was as follows; 2.1/1.7 (53%) and 1.7/1.7 (27%) in normal controls, while 2.1/1.7 (7%) and 1.7/1.7 (73%) in severe congenital malformations. This result suggests that there may be a locus which is closely associated with congenital malformations. From the gene mapping data, we found that human homeobox 4 gene cluster is present in the proximity of gamma-cristalline gene. Therefore, we focused our investigation on Hox 4 genes.Homeobox gene are highly conserved genes and considered to encode programs for body plan during embryogenesis. As a first step to investigate mutations of homeobox genes, we isolated Hox 4A genomic DNA clone and determined the nucleotide sequence of about 6000 bp. Based on the sequence data, PCR primers were synthesized and PCR-SSCP analysis was carried out in 23DNA samples from severe congenital malformations. We failed to find abnormal bands in PCR-SSCP and now trying to detect Hox 4B gene abnormalities in human congenital malformations.