五声调式基因论

文中引入了当今世界所广泛使用的“文化基因”的研究成果,首次提出了五声调式基因(pentatonic - meme )的新概念。并由此从理论到实践进一步论证:在中国传统“均、宫、调”的核心三层次中,“调”以下实际存在着一个更低的第四层次。

中国“创始”神话中隐喻的“造物”原型

从创始神话开始,华夏开启了造物与信仰象征的大"设计"活动,在至今流传的神话中都有着对应的造物(设计)隐喻。创始神话以"物"所展开的智慧,成为中国设计的"原型"基因。中国"造物"寻求"精神与物质"的双性融合,以人类的创造为语义内涵。创始神话展示了"宇宙循环""物质符号"和"文字文本"的互通转译,更具有解决人类生命共同结构的当代价值。

叶酸及其代谢相关的还原型叶酸载体、甲硫氨酸合成酶还原酶基因多态性与子宫颈癌关系的研究

目的 探讨血清叶酸、叶酸代谢相关基因还原型叶酸载体（RFC-1）A80G和甲硫氨酸合成酶还原酶基因（MTRR）A66G多态性与子宫颈癌易感性间的关系。方法 采用以医院为基础的病例对照研究，选择经病理学确诊的子宫颈鳞状细胞癌新发病例107例作为病例组，同期就诊子宫肌瘤患者107例作为对照组。用放射免疫分析法测定血清叶酸水平，PCR-限制性片段长度多态性（RFLP）法检测基因RFC-1 A80G和MTRR A66G多态性。结果 病例组血清叶酸水平[（1.86±0.60）ng／ml]低于对照组[（2.30±1.14）ng／ml]（Z＝2.08，P＝0.04），随着血清叶酸水平的降低，发生子宫颈癌的风险增加（χ2趋势＝12.57，P＝0.001）。多因素分析结果显示，RFC-1 80位点GG基因携带者患子宫颈癌的危险性是AA型的2.42倍（95 % CI 1.01～5.81），是A等位基因（AA＋AG）携带者的1.65倍（95 % CI 0.77～3.53）；MTRR 66位点GG携带者患子宫颈癌的危险性是AA型个体的1.35倍（95 % CI 0.40～4.56），是A等位基因（AA＋AG）携带者的1.26倍（95 % CI 0.38～4.16）。结论 血清叶酸缺乏可增加子宫颈癌发生的风险，RFC-1 A80G可能是子宫颈癌发生的易感因素，携带GG基因型的个体患子宫颈癌的风险增加。尚不能认为MTRR基因多态性与子宫颈癌发生的易感性有关。

还原型叶酸载体基因多态性与急性白血病易感性的相关性

目的研究还原型叶酸载体（reduced folate carrier，RFC）80G／A单核苷酸多态性（single nucleotide polymorphism，SNP）与急性自血病（acute leukemia，AL）之间的关联性。方法采用逆转录-聚合酶链反应-变性梯度凝胶电泳（reverse transcriptase-polymerase chain reaction—denaturing gradient gel electrophoresis，RT-PCR—DGGE）结合DNA直接测序，对98例AL患儿和135名正常儿童进行RFC第80位点SNP筛查。结果在AL组和正常儿童组中发现RFC80GA基因型的例数分别为53和，52，AA基因型例数分别为24和30，两组A等位基因频率分别为0．515和0．415。应用χ^2分析发现RFC80G／A与AL存在交互作用。结论确定了中国儿童RFC80G／A的等位基因频率，并初步认为RFC80G／A与AL发生存在一定的相关性。

Relationship of human leukocyte antigen class II genes with the susceptibility to hepatitis B virus infection and the response to interferon in HBV-infected patients

AIM： To study the relationship of human leukocyte antigen （HLA）-DRB1 and -DQB1 alleles with the genetic susceptibility to HBV infection and the response to interferon （IFN） in HBV-infected patients. METHODS： Low-resolution DNA typing kit was used to determine HLA-DR-1 and -DQB1 genes in 72 patients with chronic hepatitis B （CHB） and HLA-DRB1 in 200 healthy people ready to donate their bone marrow in Shanghai. Among CHB patients, 35 were treated with IFNα-1b for 24 wk. RESULTS： The frequencies of HLA-DRBI＊06, DRBI＊08 and DRB1＊16 alleles in 72 patients were higher than in 200 healthy people （2.08% vs0%, OR = 3.837, P= 0.018; 11.11% vs5.50%, OR = 2.148, P= 0.034; and 6.94% vs 3.00%, OR = 0.625, P = 0.049, respectively）; whereas that of DRBI＊07 allele was lower （2.78% vs 7.75%, OR = 0.340, P= 0.046）. The frequency of HLA-DRBI＊ 14 allele was higher in 11 responders to IFN compared with 24 non-responders （18.18% vs2.08%, OR = 10.444, P = 0.031）, whereas that of DQBI＊07 allele was inverse （9.09% vs37.50%, OR = 0.167, P= 0.021）. CONCLUSION： The polymorphism of HLA class II may influence the susceptibility to HBV infection and the response to IFN in studied CHB patients. Compared with other HLA-DRB1 alleles, HLA-DRBI＊06, DRBI＊08, and DRB1＊16 may be associated with chronicity of HBV infection, HLA-DRBI＊07 with protection against HBV infection, and HLA-DRB1＊14 allele may be associated with a high rate of the response of CHB patients to IFN treatment. Compared with other HLA-DQB1 alleles, HLA-DQBI＊07 may be associated with low response rate to IFN. 2005 The WJG Press and Elsevier Inc. All rights reserved

Are polymorphisms of N-acetyltransferase genes susceptible to primary liver cancer in Luoyang, China?

AIM: To identify whether the polymorphisms of the Nacetyltransferase (NAT) genes are susceptible to primary liver cancer (PLC) in Luoyang, a PLC low-incidence area of China.METHODS: The NAT1 and NAT2 genotypes of 96 PLC cases and 173 controls were determined by PCR-RFLP.Both interaction between NAT1 or NAT2 and environmental risk factors were analyzed based on case control study.RESULTS: Compared to the control group, the frequencies of alleles NAT1*3, NAT1*4, NAT1*10, NAT1*14B and alleles NAT2*4, NAT2*6, NAT2*7 in PLC group showed no statistically significant difference (x2 = 2.61 and 4.16,respectively, both P＞0.05). The frequencies of NAT1 genotypes NAT1*3/*3, NAT1*3/*4, NAT1*3/*10,NAT1*3/*14B, NAT1*4/*4, NAT1*4/*10, NAT1*4/*14B,NAT1*10/*10, NAT1*10/*14B, and NAT2 genotypes NAT2*4/*4, NAT2*4/*6, NAT2*4/*7, NAT2*6/*6,NAT2*6/*7 and NAT2*7/*7 also had no statistically significant difference between the two groups (x2 = 11.86 and 2.94respectively both, P＞0.05). Neither the frequencies of rapid and slow NAT1 acetylators nor the frequencies of rapid and slow NAT2 acetylators were significantly different between the two groups (x2 = 0.598 and 0.44,respectively, both P＞0.05). The interaction betweenNAT1*10 and occupational exposures was found significant with an odds ratio of 3.40 (x2 = 8.42, P = 0.004,OR 95%CI:1.03-11.22). But no interaction was found between NAT2 and any environmental risk factors.CONCLUSION: The polymorphisms of NAT1 and NAT2are not susceptible to PLC in Luoyang. Allele NAT1*10interacts with occupational exposures.

Human leukocyte antigen class II DQB1*0301, DRB1*1101 alleles and spontaneous clearance of hepatitis C virus infection: A meta-analysis

AIM： To assess the associations of human leukocyte antigen （HI_A） class Ⅱ DQB1＊0301 and/or DRB1＊1101 allele with spontaneous hepatitis C virus （HCV） clearance by meta-analysis of individual dataset from all studies published till date. METHODS： To clarify the impact of HLA class Ⅱ polymorphisms on viral clearance, we performed a metaanalysis of the published data from 11 studies comparing the frequencies of DQB1＊0301 and DRB1＊1101 alleles in individuals with spontaneous resolution to those with persistent infection. As we identified the heterogeneity between studies, summary statistical data were calculated based on a random-effect model. RESULTS： Meta-analyses yielded summary estimatesodds ratio （OR） of 2.36 [95%CI （1.62, 3.43）, P〈0.00001] and 2.02 [95%CI （1.56, 2.62）, P〈0.00001] for the effects of DQB1＊0301 and DRB1＊1101 alleles on spontaneous clearance of HCV, respectively. CONCLUSION： These results support the hypothesis that specific HLA class Ⅱ alleles might influence the susceptibility or resistance to persistent HCV infection. Both DQB1＊0301 and DRB1＊1101 are protective alleles and present HCV epitopes more effectively to CD4^＋T lymphocytes than others, and subjects with these two alleles are at a lower risk of developing chronic HCV infection. Large, multi-ethnic confirmatory and welldesigned studies are needed to determine the host genetic determinants of HCV infection.

Prokaryotic Expression of P1 Gene of Type Asia1 Foot and Mouth Disease Virus(FMDV)and the Preparation of Its Antiserum

[Objective] The aim was to study the prokaryotic expression of P1 gene of foot-and-mouth disease virus（FMDV）type Asia 1and the preparation of its antiserum.[Method]The P1 gene of FMDV type Asia 1 was obtained by gene cloning techniques,and then cloned into pET-32a（＋）plasmid;subsequently the recombinant plasmid was transformed into E.coli BL21（DE3）;after the IPTG induction and protein purification,SDS-PAGE analysis was carried out;the ultrasonic wave was use to lyse the cultivated recombinant strain,and after the isolation and purification,this fusion protein was utilized to immunize New Zealand rabbits so as to prepare P1 protein antiserum.[Result]The positive clones were obtained;SDS-PAGE result showed that the target band was appeared at 105 kD;Western blot analysis showed that the antisera could bind to the expressed P1 fusion protein specifically;the ELISA titer of the rabbit anti-FMDV-P1 sera was approximately 1∶5 120.[Conclusion]This study had provided foundations for FMDV serological diagnostic methods and genetically engineered vaccine.

Quantitation of HBsAg predicts response to entecavir therapy in HBV genotype C patients

AIM： To analysis the factors that predict the response to entecavir therapy in chronic hepatitis patients with hepatitis B virus （HBV） genotype C. METHODS： Fifty patients [hepatitis B e antigen （HBeAg）- negative：HBeAg-positive = 26：24] with HBV genotype C, who received nalve entecavir therapy for 〉 2 years, were analyzed. Patients who showed HBV DNA levels ≥ 3.0 log viral copies/mL after 2 years of entecavir ther- apy were designated as slow-responders, while those that showed 〈 3.0 log copies/mL were termed rapid- responders. Quantitative hepatitis B surface antigen （HBsAg） levels （qHBsAg） were determined by the Archi- tect HBsAg QT immunoassay. Hepatitis B core-related antigen was detected by enzyme immunoassay. Pre-C and Core promoter mutations were determined using by polymerase chain reaction （PCR）. Drug-resistance muta- tions were detected by the PCR-Invader method. RESULTS： At year 2, HBV DNA levels in all patients in the HBeAg-negative group were 〈 3.0 log copies/mL. In contrast, in the HBeAg-positive group, 41.7% were slow-responders, while 58.3% were rapid-responders. No entecavir-resistant mutants were detected in the slow-responders. When the pretreatment factors were compared between the slow- and rapid-responders; the median qHBsAg in the slow-responders was 4.57 log IU/mL, compared with 3.63 log IU/mL in the rapid- responders （P 〈 0.01）. When the pretreatment factors predictive of HBV DNA-negative status at year 2 in all 50 patients were analyzed, HBeAg-negative status, low HBV DNA levels, and low qHBsAg levels were signifi- cant （P 〈 0.01）. Multivariate analysis revealed that the low qHBsAg level was the most significant predictive factor （P = 0.03）. CONCLUSION： Quantitation of HBsAg could be a use- ful indicator to predict response to entecavir therapy.

Significance of hepatitis B virus surface antigen, hepatitis C virus expression in hepatocellular carcinoma and pericarcinomatous tissues

AIM： To investigate the correlation between hepatitis B virus surface antigen （HBsAg）, hepatitis C virus （HCV） expression in hepatocellular carcinoma （HCC）, the HAI score of the noncancerous region of the liver and the serum Alpha fetoprotein （AFP） level. METHODS： The patterns of HBsAg and HCV in 100 cases of HCC and their surrounding liver tissues were studied on paraffin-embedded sections with immunohistochemistry, the histological status was determined by one pathologist and one surgeon simultaneously using the hepatitis activity index （HAIl score, and AFP was detected by radioimmunity. The study included 100 consecutive patients who underwent curative resection for HCC. Based on HBsAg and HCV expression, the patients were classified into 4 groups： patients positive for HBsAg （HBsAg group）, patients positive for HCV （HCV group）, patients negative for both HCV and HBsAg （NBNC group） and patients positive for both HBsAg and HCV （BC group）. RESULTS： The BC group had significantly higher HAI scores than the other three groups. （BC 〉 HCV 〉 HBsAg 〉 NBNC）. HBV and HCV virus infection was positively correlated with HAI （rs = 0.39, P = 0.00011. The positive rate of AFP （85.7%） and the value of AFP （541.2 ng/mL） in the group with HBV and HCV co-infection were the highest among the four groups. The positive rate （53.3%） of AFP and the value of AFP （ 53.3 ng/mL） in the group with none-infection of HBV and HCV were the lowest. HBV and HCV virus infection was positively correlated with AFP（rs = 0.38, P = 0.0001）. CONCLUSION： The AFP increase in patients with liver cancer was positively correlated with the infection of HBV and HCV. The-serum AFP elevation by the infection of HBV and HCV is one of mechanisms which lead to hepatocarcinogenesis, and the antivirus intervening treatment of hepatitis is significant for the prognosis of liver cancer. From our Spearman＇s rank correlation analysis, we can conclude that the severity of virally induced inflammation is correlated with HBsAg and HCV expression in HCC tissues and noncancerous tissues. Prior co-infection of HBV in HCV patients may be an adverse risk factor for intrahepatic inflammation.

Cytotoxic T-lymphocyte antigen 4 gene polymorphisms and susceptibility to chronic hepatitis B

AIM： To assess the three polymorphJsm regions within cytotoxic T-lymphocyte antigen 4 （CTLA-4） gene, a C/T base exchange in the promoter region-318 （CTLA-4 -318C/T）, an A/G substitution in the exon 1 position 49 （CTLA-4 49A/G）, a T/C substitution in 1172 （CTLA-4 -1172T/C） in patients with chronic hepatitis B. METHODS： Fifty-one patients with chronic hepatitis B virus infection and 150 healthy subjects were recruited sequentially as they presented to the hepatic clinic. Classification of chronic hepatitis B virus （HBV）-infected patients was as asymptomatic carrier state （26 patients） and chronic hepatitis B （25 patients）. Genomic DNA was isolated from anti-coagulated peripheral blood Bully coat using Miller＇s salting-out method. The presence of the CTLA-4 gene polymorphisms was determined using polymerase chain reaction amplification refractory mutation system （ARMS）. RESULTS： We observed a significant association between -318 genotypes frequency （T＋C-, T＋C＋, T-C＋） and susceptibility to chronic hepatitis B （P=0.012, OR=0.49, 95%CI： 0.206-1.162）. However, we did not observe a significant association for ＋49 genotype frequency （T＋C＋, T＋C- T-C＋） and -1172 genotype frequency （C＋T＋, T＋C- C＋T-） and state of disease. CONCLUSION： Our results suggest that CTLA-4 gene polymorphisms may partially be involved in the susceptibility to chronic hepatitis B.

Quantification of HBsAg:Basic virology for clinical practice

Hepatitis B surface antigen (HBsAg) is produced and secreted through a complex mechanism that is still not fully understood. In clinical fields, HBsAg has long served as a qualitative diagnostic marker for hepatitis B virus infection. Notably, advances have been made in the development of quantitative HBsAg assays, which have allowed viral replication monitoring, and there is an opportunity to make maximal use of quantitative HBsAg to elucidate its role in clinical fields. Yet, it needs to be underscored that a further understanding of HBsAg, not only from clinical point of view but also from a virologic point of view, would enable us to deepen our insights, so that we could more widely expand and apply its utility. It is also important to be familiar with HBsAg variants and their clinical consequences in terms of immune escape mutants, issues resulting from overlap with corresponding mutation in the P gene, and detection problems for the HBsAg variants. In this article, we review current concepts and issues on the quantification of HBsAg titers with respect to their biologic nature, method principles, and clinically relevant topics.

Isolation and Characterization of Exosomes Derived from Tumor Cells Genetically Expressing Model Antigen

Tumor cell-derived exosomes have been proposed as non-cellular nanomeric vaccine which could induce potent anti- tumor immune response in mice. In order to develop the protocols to prepare tumor cell-derived exosomes for basic research and clinical trail, we isolated exosomes from ovalbumin (OVA)-expressing thymoma cells EG.7-OVA by various preparation methods. We demonstrate the non-sedimentation method is simple, rapid, efficient with higher yield and purity of exosomes. EG.7-OVA-derived exosomes are 40-100 nm in diameter sequestered by lipid bi-layer, and contain rich heat shock protein (HSP) and OVA. The result of the size distribution determination is consistent with the calculation by the visual microscopic inspection, with 90.4% particles at the range of 50-90 nm. Moreover, as a model antigen of the EG.7 cells, OVA concentra- tion in EG.7-derived exosomes can be regarded as a good quality control parameter. Therefore, we have established a platform to efficiently prepare exosomes for tumor immunotherapy.

DNA TYPING SYSTEM FOR HLA-A2 ALLELES BY POLYMERASE CHAIN REACTION WITH SEQUENCE- SPECIFIC PRIMERS

Objective. To establish a PCR- SSP method for discriminating as many HLA- A* 02 alleles, which could easily be introduced into a routine laboratory. Methods. In this study we typed HLA- A* 02 polymorphisms by a sequence- specific primer (SSP) method, which involved round 1 and round 2 PCR reactions to detect 17 HLA- A* 02 alleles (they are HLA- A* 0201- 0217 alleles) covering exon 2 and exon 3. Results. We have found that DNA sample concentration and purity were the most important variables in determining the quality of the results. For identifying correct band size, the size marker used was important. We noticed that different PCR machines performed differently. By this method, we detected 20 HLA- A* 02 positive genomic DNA samples and found 4 kinds of HLA- A* 02 alleles. They were HLA- A* 0201, 0203, 0206 and 0210. Conclusion. The HLA- A* 02 PCR- SSP method was proven to be a reliable and easily applicable typing method. Our results suggest that the SSP described here provides an optimal HLA- A* 02 typing technique that may be useful in selecting donor- recipient pairs in bone marrow transplantation between unrelated individuals.

Cloning of Omp1 Gene from Chlamydia trachomatis F Genotype

Objective: To directionally clone the ompl gene fromChlamydia trachomatis (Ct) F genotype onto a plasmid vectorfor constructing a rudimentary DNA vaccine. Methods: The complete ompl gene from genomic DNA of CtF genotype wild species was amplified with primers designedby computer. The recombinant gene was obtained byrestriction enzyme cutting, linking the gene with the plasmidvector in vitro, transforming the recombinant gene intobacteria, and extracting the DNA from the bacteria. Results: DNA extracted from the bacteria was composed ofthe ompl gene and plasmid, which is identified by threemethods of singular restrictive enzyme cutting, doublerestrictive enzyme cutting and PCR. Conclusion: Cloning of the ompl gene from the Ct Fgenotype means that a rudimentary DNA vaccine wassuccessfully constructed.

A novel animal model for in vivo study of liver cancer metastasis

AIM:To establish an animal model with human hepatocyte-repopulated liver for the study of liver cancer metastasis.METHODS:Cell transplantation into mouse livers was conducted using alpha-fetoprotein(AFP)-producing hu-man gastric cancer cells(h-GCCs) and h-hepatocytes as donor cells in a transgenic mouse line expressing urokinase-type plasminogen activator(uPA) driven by the albumin enhancer/promoter crossed with a severe combined immunodeficient(SCID) mouse line(uPA/SCID mice).Host mice were divided into two groups(A and B).Group A mice were transplanted with h-GCCs alone,and group B mice were transplanted with h-GCCs and h-hepatocytes together.The replacement index(RI),which is the ratio of transplanted h-GCCs and h-hepatocytes that occupy the examined area of a histological section,was estimated by measuring h-AFP and h-albumin concentrations in sera,respectively,as well as by immunohistochemical analyses of h-AFP and human cytokeratin 18 in histological sections.RESULTS:The h-GCCs successfully engrafted,repopulated,and colonized the livers of mice in group A(RI = 22.0% ± 2.6%).These mice had moderately differentiated adenocarcinomatous lesions with disrupted glandular structures,which is a characteristics feature of gastric cancers.The serum h-AFP level reached 211.0 ± 142.2 g/mL(range,7.1-324.2 g/mL).In group B mice,the h-GCCs and h-hepatocytes independently engrafted,repopulated the host liver,and developed colonies(RI = 12.0% ± 6.8% and 66.0% ± 12.3%,respectively).h-GCC colonies also showed typical adenocarcinomatous glandular structures around the h-hepatocyte-colonies.These mice survived for the full 56 day-study and did not exhibit any metastasis of h-GCCs in the extrahepatic regions during the observational period.The mice with an h-hepatocyte-repopulated liver possessed metastasized h-GCCs and therefore could be a useful humanized liver animal model for studying liver cancer metastasis in vivo.CONCLUSION:A novel animal model of human liver cancer metastasis was established using the uPA/SCID mouse line.This model could be useful for in vivo testing of anti-cancer drugs and for studying the mechanisms of human liver cancer metastasis.

Establishment of an orthotopic transplantation tumor model of hepatocellular carcinoma in mice

AIM:To improve the outcome of orthotopic transplantation in a mouse model,we used an absorbable gelatin sponge(AGS) in nude mice to establish an orthotopic implantation tumor model.METHODS:MHCC-97L hepatocellular carcinoma(HCC)cells stably expressing the luciferase gene were injected into the subcutaneous region of nude mice.One week later,the ectopic tumors were harvested and transplanted into the left liver lobe of nude mice.The AGS was used to establish the nude mouse orthotopic implantation tumor model.The tumor suppressor gene,paired box gene 5(PAX5),which is a tumor suppressor in HCC,was transfected into HCC cells to validate the model.Tumor growth was measured by bioluminescence imaging technology.Semi-quantitative reverse transcription polymerase chain reaction(RT-PCR) and histopathology were used to confirm the tumorigenicity of the implanted tumor from the MHCC-97L cell line.RESULTS:We successfully developed an orthotopic transplantation tumor model in nude mice with the use of an AGS.The success rate of tumor transplantation was improved from 60% in the control group to 100% in the experimental group using AGS.The detection of fluorescent signals showed that tumors grew in all live nude mice.The mice were divided into 3 groups:AGS-,AGS+/PAX5-and AGS+/PAX5 +.Tumor size was significantly smaller in PAX5 transfected nude mice compared to control mice(P < 0.0001).These fluorescent signal results were consistent with observations made during surgery.Pathologic examination further confirmed that the tissues from the ectopic tumor were HCC.Results from RT-PCR proved that the HCC originated from MHCC-97L cells.CONCLUSION:Using an AGS is a convenient and efficient way of establishing an indirect orthotopic liver transplantation tumor model with a high success rate.

A versatile cloning vector facilitates target geneexpression in prokaryotic and eukaryotic cells

Objective： To facilitate manipulation of gene expression in different host cells, we used pEGFP-N1 as backbone to construct a versatile vector that can drive foreign gene expression in prokaryotic and eukaryotic cells. Methods： A cloning and expression vector, pEGFP-NI-lac, was constructed by inserting the prokaryotic lac promoter of pUC 19 into the eukaryotic expression vector, pEGFP-N1, between the eukaryotic PCMV promoter and enhanced green fluorescent protein （EGFP） open reading frames. To assess the function of pEGFP-NI-lac, the nucleotide sequence encoding the hepatitis C virus （HCV） core protein was cloned into the multiple cloning sites. Western blotting analysis was used to detect the expression of the HCV core protein in Escherichia coli DH5a and HepG2 cells. Results： Restriction enzyme digestion and sequence analysis indicated that pEGFP-NI-lac was successfully constructed and the HCV core gene was cloned into this vector. The Western blotting results showed that pEGFP-NI-lac promoted expression of HCV core gene in prokaryotic E. coli DH5a and eukaryotic HepG2 cells. Conclusion： The pEGFP-NI-lac vector has been successfully constructed and functions in both prokaryotic and eukaryotic cells. The EGFP reporter can be used as an insert-inactivation marker for clone selection or as an expression tag. This vector can be used for cloning and expression of genes in both prokaryotic and eukaryotic cells, making gene cloning, expression and functional studies convenient as well as time- and labor-efficient

CTLA-4 gene A/G polymorphism associated with diabetes mellitus in Han Chinese

OBJECTIVE: To investigate the association of cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) gene A/G polymorphism with susceptibility to diabetes mellitus in Han Chinese. METHODS: An A/G transition at position 49 of exon 1 was analyzed in 31 patients with type 1 diabetes, 31 patients with type 2 diabetes, and 36 controls were analyzed by polymerase chain reaction-restriction fragment length polymorphism analysis. RESULTS: A highly significant increase in the frequency of the G allele was seen in patients with type 1 diabetes compared with controls (66.1 % vs. 34.7%, respectively; P

Genotyping and preparation of the recombinant nucleocapsid protein antigen of hantavirus

Objective To identify new recombinant antigens with potential for diagnosis of hemorrhagic fever with renal syndrom (HFRS) and establish reverse transcription-polymerase chain reaction-restriction fragments length polymorphism (RT-PCR-RFLP) for genotyping of hantavirus.Methods One group of primers was used to clone the full-length S genome segment and the partial S genorme segment of the N-terminal. The two cloned genes were both fusionally expressed and nonfusionally expressed in the T7 system. The other group of primers was used to establish a RT-PCR method to detect RNAs in 37 virus isolates, 2 positive standard virus strains of hantavirus and 5 negative controls.The method for typing RFLP was set up by digesting the PCR products of 20 virus isolates with Ras Ⅰ and Hind Ⅲ.Results The non-fusionally expressed products with a working concentration of 1:10 000 by chapping enzyme-linked immunosorbent assay (cELISA), presented good biological activity though yields were lower than that of the fusionally expressed products.The specific component of the hantavirus genome (299 bp or 577 bp) wes seen in all viral samples. The genotyping of hantavirus showed that 9 out of the total were hantann (HTN) viruses, 8 were seol (SEO) viruses and 3 were not determined.Conclusions The good working titrer of expressed recombinant antigen showed that it has the potential to replace the natural antigen for detecting hantavirus antibodies. On comparison with cELISA, the detection rates for these two methods were 100% and 84.6%, and the coincidence rate was 84.6%. The former had a 15.4% higher sensitivity than the latter. The typing efficiency of RT-PCR-RFLP and sero-typing method was 85% (17/20) and 55% (11/20), respectively, showing that the former was 30% higher than the latter, while their results were highly consistant.