Clinical and gene mutation analyses of three patients with ornithine carbamoyltransferase deficiency / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To analyze the clinical and genetic characteristics of three children with ornithine carbamoyltransferase deficiency(OTCD), and to provide a practical method for gene diagnosis and genetic counseling of the disease.</p><p><b>METHODS</b>All exons of the ornithine carbamoyltransferase (OTC) gene were screened by polymerase chain reaction-DNA direct sequencing in the three OTCD patients.</p><p><b>RESULTS</b>One patient firstly presented as vomiting at 6 month of age. A missense mutation of T262I was detected. His mother had the same mutation without any clinical symptoms. The second patient presented as restlessness, and had a missense mutation of R277W. Gene analysis of his parents was not available. The third patient presented as neonatal lethargy, harbored a missense mutation of I172M. His mother had the same mutation without any clinical symptoms.</p><p><b>CONCLUSION</b>Gene mutation analysis is a feasible way for diagnosing OTCD. Patients with I172M mutation present symptom early, while those with T262I and R277W mutations manifest symptoms later. Gene mutation analysis will be important for asymptomatic and prenatal diagnosis and genetic counseling.</p>

Gene polymorphism of vascular endothelial growth factor in children with Henoch-Schonlein purpura nephritis / 中国当代儿科杂志

<p><b>OBJECTIVE</b>To study the relationship of -634G/C gene polymorphism of vascular endothelial growth factor (VEGF) with Henoch-Schonlein purpura nephritis (HSPN) in children.</p><p><b>METHODS</b>One hundred ethnic Han children with HSP, including 50 children with concurrent nephritis (HSPN group) and 50 children without nephritis (HSP without nephritis group), were enrolled. Fifty age-, sex-and ethnics-matched healthy children were used as the control group. VEGF-634G/C genotypes were determined by PCR-RFLP. Plasma VEGF levels were measured using ELISA.</p><p><b>RESULTS</b>CC genotype distribution (32%) and C allele frequency (56%) in the HSPN group were significantly higher than those in the control group (10% and 35% respectively) and the HSP without nephritis group (10% and 33% respectively) (P<0.01). The incidence of nephritis in HSP patients with CC genotype increased significantly when compared with those with GG genotype (76% vs 31%; P<0.01). Plasma VEGF levels in patients with CC genotype (180.5+/- 40.7 pg/mL) were significantly higher than those in patients with CG (145.2+/- 48.3 pg/mL) and GG (101.5+/- 26.5 pg/mL) genotypes (P<0.05).</p><p><b>CONCLUSIONS</b>VEGF-634G/C gene polymorphism may be associated with the development of HSPN. C allele may a susceptible gene of HSPN.</p>

Construction of a trans-splicing ribozyme for restoring EGFP truncation mutation / 生物工程学报

Special designed group I intron ribozymes can specifically splice objective RNA, repair the mutant gene in RNA level. The specificity of ribozyme is determined by nucleotides specific internal guide sequence (IGS) introduced to the enzyme. In this study, fragment sequence containing Tetrahymena thermophilia intron I of 26S rRNA gene was cloned and cis-splicing activity of this ribozyme was confirmed by in vitro transcription. For evaluating the trans-splicing activity of this ribozyme, a truncated mutant Green Fluorescence Protein (GFP) vector, XYQ5/XYQ10- pEGFP-C2, was constructed. This vector deleted the 3' end 564bp fragment of EGFP coding sequence, led to the lost the activity of emitting green fluorescence. Trans-splicing ribozyme plasmids ptrans-rib-CMV2 for remedy of the truncated mutant EGFP was constructed by PCR and molecular cloning techniques. This vector utilizing cloned 26S rRNA intron 1 as core enzyme; selecting T-G site at 194bp of EGFP coding sequence as splicing receptor, designed an IGS which is inversely complement to the 188-193nt of EGFP mRNA; the 195-890bp fragment of EGFP coding sequence was ligated to the 3'-end of ribozyme core. The fragment containing these components was inserted to a eukayotic expression vector pRC-CMV2. Using linearized XYQ5/XYQ10- pEGFP-C2 and ptrans-rib-CMV2 as templates, truncated EGFP mRNA and the constructed ribozyme vector were transcribed and mixed to evaluate the trans-splicing activity. Analysis of in vitro transcription products mix by RT-PCR verified the existence of wild type EGFP mRNA molecule. Co-transfection of XYQ5/XYQ10- pEGFP-C2 with ptrans-rib-CMV2 to Hela cells proved this ribozyme restored green fluorescence within cell, but the efficiency was low.