Mutation screening and prenatal diagnosis of tuberous sclerosis complex / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To screen mutations of tuberous sclerosis complex (TSC) patients to confirm a clinical diagnosis of TSC, and to perform prenatal diagnosis for families with mutations.</p><p><b>METHODS</b>In this study, PCR-denaturing high-performance liquid chromatography(DHPLC), supplemented with sequencing when necessary, was used to screen TSC1 and TSC2 mutations in 21 patients from 19 pedigrees visited author's hospital in the last five years. For novel mutations, one hundred unrelated healthy individuals were screened to exclude the possibility of polymorphism.</p><p><b>RESULTS</b>Seventeen different mutations were found in 21 patients of 19 pedigrees with 13 being novel mutations, including c. 2672delA, c. 2672insA of TSC1 gene and c.4918insCGCC, c.1143delG, Intron27+1 G>A, c.1957-1958delAG, Intron5+1 G>A, c.910insCT, c.2753 C>G, c.4078dupAGCAAGTCCAGCTCCTC, Intron 11 -1 G>A, Intron 14+1 G>A, c.684 C>A of TSC2 gene, indicating a high frequency of de novo mutations in TSC. Three of these mutations were in the TSC1 gene (N762S, c.2672insA and c. 2672delA), while all remaining 14 were in the TSC2 gene. Prenatal diagnosis for TSC was performed for 7 fetuses from these pedigrees. The six fetuses that tested negative for TSC mutations were carried to term and, to date, none of these children has shown symptoms of TSC.</p><p><b>CONCLUSION</b>Author's data showed that a mutation detection rate of tuberous sclerosis was 89.5%(17/19) among patients in author's hospital. The ratio of TSC2 and TSC1 mutations was about 1:1 in the familial cases, but TSC2 mutation was more common than TSC1 mutation in sporadic cases. Author's data demonstrated that birth of TSC children for those with familial history of TSC could be prevented through prenatal diagnosis.</p>

Mutation detection of PKD1 gene in patients with autosomal dominant polycystic kidney diseases / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To detect gene mutation in the patients with autosomal dominant polycystic kidney disease (PKD).</p><p><b>METHODS</b>Polymerase chain reaction (PCR)-denaturing high-performance liquid chromatography (DHPLC) analyses were performed in 3o single copy region of PKD 1 gene (PKD1). DNA sequencing were carried out on PCR products with abnormal peak shape afterwards.</p><p><b>RESULTS</b>A new nonsense mutation (C11901A in exon 42 of PKD1 was identified to cause serine in position 3897 turning to a stop codon. A missense mutation, C10737T, was detected in exon 35 which caused threonine in position 3509 turn to methionine. Two kinds of samesense mutation, G11824A and C11860T in exon 42, were found in normal control.</p><p><b>CONCLUSION</b>PKD1 mutation were detected successfully by PCR-DHPLC. A new nonsense mutation, a missense mutation and two polymorphisms are identified in this study.</p>

Diagnosis of hemophilia A by a combination of St14(DXS52) VNTR polymorphism and (CA)n repeat polymorphism within FVIII gene / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To improve the accuracy and the diagnostic rate of gene diagnosis and prenatal gene diagnosis for hemophilia A (HA) families.</p><p><b>METHODS</b>Linkage analysis was performed by using St14(DXS52) VNTR polymorphism and intron 13 (CA)n repeat polymorphism of the factor VIII gene among HA families for indirect diagnosis.</p><p><b>RESULTS</b>The diagnostic rates using linkage analysis based upon one of the above mentioned two polymorphic loci among 9 HA families were 66.7% and 66.7%, respectively. The diagnostic rate rose to 88.9% by using a combination of the two polymorphic loci. Prenatal gene diagnoses were performed for 4 HA families. A wrong prenatal diagnosis which may happen when linkage analysis was performed by using only St14 VNTR was monitored.</p><p><b>CONCLUSION</b>The rapid and accurate gene diagnosis and prenatal gene diagnosis could be performed by a combination of the two polymorphic loci for about 90% HA families.</p>

46,XY female sex reversal patient with a novel point mutation in the coding sequence of the SRY gene / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To investigate the molecular mechanism of a Chinese patient with 46, XY sex reversal.</p><p><b>METHODS</b>DNA fragments of the SRY gene from the typical XY female sex reversal patient and her father were amplified by polymerase chain reaction (PCR). The amplified PCR fragments were cloned into the pUCm-T vector, and direct sequencing was carried out on an ABI 377-3 automated DNA sequencer to detect the mutation. PCR-restriction enzyme digestion was applied to detect the results of DNA sequencing.</p><p><b>RESULTS</b>A novel mutation of the SRY gene was identified in the XY sex reversal patient of this study. A T is replaced by an A in codon 129 at position +387, resulting in the replacement of the polar amino acid tyrosine (TAT) by the stop code (TAA) in the HMG-box, whereas her father was proved to have the wild-type sequence. Because the mutation introduced an enzyme site of MaeIII, the PCR-restrict enzyme digestion showed that there were three bands (131 bp,231 bp and 247 bp) in the patient, whereas two bands (131 bp and 478 bp) in normal man. It verified the results of sequencing analysis. The results after searching the Human Gene Mutation Database showed that this mutation was not described before and should be a new mutation.</p><p><b>CONCLUSION</b>The novel mutation in SRY gene has provided valuable information for the understanding of molecular mechanism of the patient with 46,XY female sex reversal.</p>

Diagnosing achondroplasia by single cell nested-PCR / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To research on the reliability of diagnosing achondroplasia (ACH) on single cell level and to provide a basis for preimplantation genetic diagnosis(PGD).</p><p><b>METHODS</b>The high-frequency mutation region G380R of fibroblast growth factor receptor 3(FGFR3) gene was amplified by nested-PCR with single lymphocyte and single blastomere. The products of PCR were digested by restriction enzyme Bfm I, then the digested products were detected by 10% polyacrylamida gel electrophoresis(PAGE).</p><p><b>RESULTS</b>The amplification success rate, allele dropout rate and correct diagnosis rate of single lymphocyte's PCR were 90.4%, 8.2% and 91.8%,respectively. The amplification success rate of single blastomere was 75.4%.</p><p><b>CONCLUSION</b>The diagnosis of ACH by single cell nested-PCR is comparatively stable and reliable.</p>