Association of coagulation factor Ⅻ gene C46T polymorphism and coronary artery disease in patients documented angiography

AIM: To study the distribution of C46T polymorphism of factor Ⅻ(FⅫ) in Chinese Han population and the association of the polymorphism with coronary artery disease(CAD) and acute coronary syndrome(ACS). METHODS: Selected coronary angiography was performed in 168 CAD patients and 210 controls. Genetype of FⅫ was typed by mutagenically separated polymerase chain reaction assay(MSPCR). RESULTS: FⅫ allelic frequencies of C and T were 29.8%, 70.2% and 31.4%, 68.6% in CAD and controls, respectively(P>0.05). Genetype distribution was in accordance with Hardy-Weinberg equilibrium. The frequency of CC, CT, TT in CAD and control was 8.7%, 40.5%, 50.0% and 5.2%, 52.6%, 42.2%. The association between FⅫ genetype and CAD(2=6.393, P<0.05) was observed. As compared with the CC group, the CT genetype was a protective factor for CAD(OR 0.43, 95% CI 0.19-0.97). When compared to stable coronary artery disease, the frequency of TT genetype is significant less in ACS group(45.0% vs 62.5%, 2=4.200, P<0.05). The distribution of genetype in C46T was no significant difference among the numbers of stenosed coronary artery. CONCLUSION: The C46T polymorphism of FⅫ is association with CAD in Chinese Han population. The C→T mutation may be a protective factor against CAD and ACS.     

Genetic polymorphisms of RTN4 gene in Chinese Guangxi population

AIM: To investigate the distribution characteristics of rs2920891A/C and rs17046647A/G polymorphisms of RTN4 gene in Guangxi population, and to compare the differences among different populations. METHODS: The genotypes of RTN4 gene at rs2920891A/C and rs17046647A/G in 323 healthy persons of Guangxi were performed by the technique of SNaPshot and DNA sequencing. The results were compared with the alleles and genotypes of other populations (HapMap-CEU, HapMap-HCB, HapMap-JPT and HapMap-YRI in HapMap). RESULTS: In Guangxi population, 3 genotypes, AA, AC and CC, and 2 alleles, A and C, were found in rs2920891A/C. The allele frequencies between male and female showed significant differences (P<0.05). The genotype and allele frequencies compared with HapMap-JPT, HapMap-CEU and HapMap-YRI had differences with statistical significance (P<0.05). Three genotypes, AA, AG and GG, and 2 alleles, A and G, were found in rs17046647A/G. The genotype and allele frequencies between male and female showed no significant differences (P>0.05), but there were significant differences of the genotype and allele frequencies as compared with HapMap-JPT, HapMap-CEU and HapMap-YRI (P<0.01).CONCLUSION: The rs2920891A/C and rs17046647A/G polymorphisms of RTN4 gene in Chinese Guangxi population are different from those in other races.     

Distribution of α1 immunoglobulin gene VNTR polymorphisms in Chinese

AIM: To investigate the polymorphisms of the variable number tandem repeat (VNTR) in the α1 immunoglobulin gene in the Chinese population and to compare them with the Caucasians. METHODS: 3 VNTR loci (VNTR1:α1 gene hs1,2 enhancer VNTR; VNTR2: 6 kb forward (change to upstream or down stream) the VNTR1 and E5VNTR: located in the fifth exon) and their lengths in the α1 gene have been retrieved from the genetic databases and literature. Genomic DNA was extracted from 201 healthy Chinese Han subjects. The sizes of the 3 VNTRs were determined by polymerase chain reaction (PCR) and gel electrophoresis, and confirmed by sequencing randomly selected samples. RESULTS: The VNTR1 locus revealed three different lengthed alleles, designated as α1A ,α1B and α1C for one, two and three repeat sequences, respectively, with the frequencies of 30.3%, 59.7% and 10.0%, respectively. 6 genotypes were formed from the alleles with the frequencies of 12% for the AA; 32.3% for the AB; 37.8% for the BB; 4.5% for the AC; 11.4% for the BC; and 2.0％ for the CC, respectively. Compared with the reported Caucasian population the frequencies of the C allele and the BC、CC、AC genotypes were significantly higher (P＜0.01), the A allele and the AB genotype frequencies were significantly lower (P＜0.01). All the examined subjects showed the uniformed lengths of 136 bp for the VNTR2 and 535 bp for E5VNTR alleles. CONCLUSIONS: The repeat numbers of the VNTR1 of the α1 gene in the Chinese Han population are significantly different from the Caucasians with a higher C allele frequency and BC、 AC genotypes, and lower A allele frequency and AB genotype. We could not find evidence of polymorphism in the VNTR2 and E5VNTR loci in the examined subjects. The results represent the first data from the Chinese population regarding the VNTR polymorphisms in the I alpha 1 gene, and provide a useful tool for the gene and the gene related studies.     

Relation between severe coronary stenosis lesion distribution and risk factors in young and middle-aged adults

AIM: To evaluate the classical and novel risk factors in young and middle-aged patients with coronary artery disease (CAD), and to analyze the relation between coronary risk factors and coronary lesion distribution. METHODS: A group of one hundred and eighty-nine patients, aged not more than 60 years with severe coronary stenosis on coronary angiography, and the other group of age-matched one hundred and sixty-one patients having normal or non-severe stenosis on coronary angiography were comprised in the study. Conventional factors (such as smoking, diabetes, hypertension, lipid spectrum and fasting plasma glucose) and novel risk factor (homocysteine) were compared between the groups. Moreover, the relation between the risk factors, and coronary lesion distribution including left main artery (LMA) or proximal or mid-left anterior descending (LAD) artery and remaining coronary lesions were investigated. Logistic regression analysis was used to define confounding factors for predicting severe CAD and coronary lesion distribution. RESULTS: Male and smoking were more prevalent in the group with severe coronary stenosis compared to the other group. The levels of homocysteine, triglyceride and apolipoprotein B were also higher in the group with severe coronary stenosis than those in the other group. From the perspective of gender analysis, homocysteine and apolipoprotein B significantly increased regardless of gender in severe coronary stenosis group. In men, the prevalence of diabetes rates and fasting plasma glucose were higher in severe coronary stenosis group. In women, the prevalence of triglyceride is obviously increased in severe coronary stenosis group. Male, smoking, homocysteine and apolipoprotein B were independent predictors of severe CAD in young and middle-aged patients according to logistic regression analysis with odds ratios of 2.798 (95% CI: 1.520~5.152; P<0.01), 3.570 (95% CI: 2.125~5.996; P<0.01), 1.079 (95% CI: 1.028~1.133; P<0.01), and 2.883 (95% CI: 1.427~5.825; P<0.01). Group analysis in the severe coronary stenosis patients revealed that only homocysteine was an independent predictor of LMA or proximal or mid-LAD lesion presence with an odds ratio of 0.911 (95% CI: 0.831~0.998; P<0.05). CONCLUSION: In our study, young and middle-aged patients with severe CAD have different risk profiles with higher frequency of male, smoking and increased levels of apolipoprotein B and homocysteine. Only homocysteine predicts coronary lesion distribution in LMA and proximal or mid-LAD.     

Distribution characteristics of polymorphisms of interleukin-22 gene in Guangxi population and their association with serum lipid levels

AIM: To investigate the distribution characteristics of interleukin-22 (IL-22) gene rs2227485C/T and rs2227491A/G polymorphisms in Guangxi people and the distribution differences with other ethnic groups, and to explore the difference levels of common lipid indexes in different genotypes. METHODS: SNaPshot technique and DNA sequencing were used in 280 Guangxi persons to examine IL-22 genotypes and to analyzed the distribution frequencies of allele and genotype in these sites. The distribution frequencies in different sexes, and the differences between groups and diffe-rence levels of common lipid indexes in different genotypes were analyzed statistically. RESULTS: Three genotypes of CC, CT and TT were found in rs2227485C/T with the frequency distribution of 17.1%, 49.3% and 33.6%, respectively. No significant difference between different sexes of each genotype and allele frequency in the Guangxi population was observed (P>0.05). Compared with the distribution frequencies of genotype and allele in HapMap-TSI, HapMap-HCB, HapMap-JPT and HapMap-MEX, those in Guangxi population showed statistically significant differences (P<0.05). Three genotypes of AA, AG and GG were found in rs2227491A/G with the frequency distribution of 16.1%, 52.8% and 31.1%, respectively. There was no significant difference between different sexes of each genotype and allele frequency in the Guangxi population (P>0.05). The significant differences of genotype frequencies among Guangxi population, HapMap-TSI, HapMap-JPT and HapMap-MEX were detected (P<0.05). Compared with the other 4 populations, allele frequencies in Guangxi population had significant difference (P <0.05). There were significant differences in the levels of HDL-C and LDL-C among the 3 genotypes of rs2227491A/G. The level of HDL-C had difference between AG/AA genotype and GG genotype. In addition, the level of LDL-C had difference between AG/GG genotype and AA genotype (P<0.05). CONCLUSION: rs2227485C/T and rs2227491A/G polymorphisms of IL-22 gene have differences in different populations. The rs2227491A/G polymorphism may be associated with serum lipid levels.     

Study on the association of the polymorphism at the position -418A/G and -384C/T in the Apo(a) promoter

AIM: To investigate two single nucleotide polymorphisms (SNP) in the apolipoprotein(a) promoter at positions -418 and -384 and to compare distributing difference of genotype frequencies of single nucleotide among different races and to explore the influencies of them on serum lipid level and their association with coronary heart disease (CHD). METHODS: Using PCR-RFLP (BsgI,BfaI) method, we determined genotypes of these two SNPs in 156 unrelated healthy controls of HanZu Chinese and 56 unrelated CHD patients of HanZu Chinese and 56 unrelated African Blacks, then cloned polymerase chain reaction (PCR) products into T-vector and sequenced it by M13 currency primer, correspondingly. RESULTS: (1) There was no polymorphism at position -418A/A and -384C/C in control group. Only one CHD patient's genotype determined was -418G/G, other were -418A/A and -384C/C in CHD patients. (2) Only two African Blacks' genotype determined was -418G/G, other were -418A/A and -384C/C in African Blacks. (3) However, the Apo(a) promoter sequence was in coincident with the sequence publicized in GenBank and the base at positions -418 was adenine (A) and -384 was cytosine (C). CONCLUSION: The mutation frequencies at position -418 and -384 were low in the Chinese Han Population of Hubei and perhaps no single nucleotide polymorphisms was at two positions. No association with serum lipid levels and CHD was observed. There were great variabilities to the SNPs in the Apo(a) promoter among different races.     

Association of OPG gene single nucleotide polymorphisms with susceptibility to rheumatoid arthritis in Chinese Han population

AIM：To investigate the association of osteoprotegerin (OPG) gene single nucleotide polymorphisms (SNPs), 163A/G (rs3102735) and 245T/G (rs3134069), with susceptibility to rheumatoid arthritis (RA) in Chinese Han population. METHODS：A total of 205 patients with RA and 171 healthy control subjects were enrolled into this study. Genotyping was performed by polymerase chain reaction-based restriction fragment length polymorphism and subsequently confirmed by DNA sequencing. Odds ratio (OR) and 95% confidence intervals (CI) were calculated for the risk genotypes and alleles. RESULTS：OPG gene polymorphisms 163A/G and 245T/G were conformed to the Hardy-Weinberg equilibrium. The statistical differences in the genotypes of AA, AG and GG at 163A/G locus were found in RA and controls. The G allele was associated with an increased risk of RA, with OR of 1.219 (95％ CI: 1.066~2.339). No significant difference was observed between RA group and control group with respect to genotypic and allelic frequencies of OPG gene 245T/G (P>0.05）. CONCLUSION：The OPG gene 163A/G SNP may be associated with RA susceptibility, and G allele may be the risk factor for developing RA.     

Association of endothelial lipase gene Thr111Ile and Gly26Ser polymorphism with lipoprotein in patients with coronary heart disease

AIM: To investigate the association of endothelial lipase gene (LIPG) Thr111Ile and Gly26Ser polymorphism with lipoprotein in patients with coronary heart disease (CHD) in Chinese.METHODS: 438 patients were classified as 242 CHD group and 196 controls group by selective coronary angiography.Plasma level of lipoprotein was determined and the Thr111Ile and Gly26Ser polymorphism was screened by PCR-RELP.RESULTS: The frequencies of Thr111Ile genotype in Chinese were CC 76.7%,CT 23.3％,TT 0.0％.The frequencies of allele were C 88.3％,T 11.7%.The plasma level of HDL-c in CT group was significantly higher than that in CC group (P<0.05) on logistic regression analysis.However,logistic regression analysis revealed that there was no significant difference between CHD group and control group for Thr111Ile polymorphism (P>0.05).No Gly26Ser mutation was observed in this study.CONCLUSION: The polymorphism of Thr111Ile is present in patients with CHD in Chinese,and T allele is related to high HDL-c level.There is no significant association between the polymorphism of Thr111Ile and CHD.The Gly26Ser mutation has not found in this study.     

Relationship between SNP of TNFR gene and incidence/severity of pneumonia

AIM：To analyze the relationship between the single nucleotide polymorphism (SNP) of tumor necrosis factor receptor (TNFR) gene and the incidence and severity of pneumonia. METHODS：Total 132 Chinese individuals were enrolled in this study. There were 66 patients with pneumonia and 66 healthy subjects. The SNPs of TNFR gene including TNFR1+36A/G, TNFR1-609G/T, TNFR2+676T/G, TNFR2+1663T/G, TNFR2 +1668A/G and TNFR2 +1690C/T were genotyped by polymerase chain reaction-restriction fragment length polymorphism or gene sequencing for all subjects. Polymorphisms affecting pneumonia incidence and severity were calculated by SPSS. RESULTS：The frequencies of TNFR1-609G and T alleles in pneumonia patients were 40.9% and 59.1%, while those in healthy subjects were 53.8% and 46.2%. The frequency of TNFR1-609T in pneumonia patients was higher than that in healthy subjects (P<0.05). Besides, the frequencies of TNFR1-609G and T alleles in severe pneumonia patients were 25.0% and 75.0%, while those were 46.0% and 54.0% in non-severe pneumonia patients. The frequencies of TNFR2 +1690C and T alleles in severe pneumonia patients were 81.1% and 18.9%, while those were 61.0% and 39.0% in non-severe pneumonia patients. The frequencies of TNFR1-609T and TNFR2 +1690C in severity pneumonia subjects were higher than those in mild subjects (P<0.05). CONCLUSION：It appears that TNFR1-609T is associated with high incidence of pneumonia. TNFR1-609T and TNFR2+1690C are the risk factors of severity in pneumonia in Chinese.     

Association of F XⅢVal34Leu with coronary heart disease

AIM: To investigate the distribution of coagulation factor XⅢ (FXⅢ) Val34Leu polymorphism in Chinese and the relationship between the polymorphism and coronary heart disease (CHD) or myocardial infarction. METHODS: A total of 195 patients with angiographically confirmed CHD and 203 controls were genotyped for the Val34Leu polymorphism by polymerase chain reaction-single strand conformational polymorphism (PCR-SSCP) analysis. RESULTS: The FXⅢ Val34Leu was found in 19 out of all 398 samples (4.8%) representing a Leu allele frequency of 2.4%. The distribution of FXⅢ genotype and allele was not significantly different between all patients and controls (P>0.05). The Val/Leu genotype and Leu allele frequencies in subjects without MI were significantly higher than that in subjects with MI (P<0.05). CONCLUSION: There is F XⅢ Val34Leu polymorphism in Han ethnic group.     

Association between HBV infection and HLA-DPB1 gene in population of Guangzhou Chinese

AIM: To investigate the association between HBV infection and HLA-DPB1 gene in population of Guangzhou Chinese. METHODS: 58 unrelated patients (test positive of HbsAg, HBeAg, HbcAb) and 75 unrelated healthy control individuals were typed by sequencing based typing (SBT) method in their HLA-DPB1 gene. RESULTS: The phenotype frequencies of HLA-DPB1 alleles of patients and control have no significant difference. CONCLUSION: These results indicate that there is no association between HLA-DPB1 gene and HBV infection.     

6个甜樱桃品种ACO基因多态性的检测

乙烯在植物果实成熟过程中起着重要的作用,ACC合酶(1-aminocyclop ropane-1-carboxylic acid synthase,ACS)和ACC氧化酶(1-aminocyclop ropane-1-carboxylic acid oxidase,ACO)是植物乙烯生物合成途径的限速酶。通过DNA序列分析,以不同果实成熟期的6个甜樱桃品种(Prunus avium L.)为材料,检测ACO基因的多态性。获得甜樱桃ACO基因约1 kb,与桃(P.persica)ACO基因(GenBank登录号:AF532976)序列的同源性达96%,其预测的氨基酸序列与桃、梅(P.mume)、美洲李(P.armeniaca)和欧洲李(P.domestica)等ACO的氨基酸序列同源性超过95%。该片段包括4个外显子和4个内含子,内含子符合GT-AT规律。用DNAMAN进行多序列比对分别在内含子2和内含子4内发现2个多态性简单重复序列(AT)n。内含子2有3种片段:即(AT)6、(AT)7和(AT)8;内含子4有2种片段,即(AT)5和(AT)6,组合后共得到4种ACO单倍型。研究在甜樱桃ACO基因座上发现2个SSR标记,为进一步研究ACO基因多态性与果实成熟期相关性奠定基础。     

Changes of OX40 ligand expression in patients with acute coronary syndromes

AIM: To investigate the changes of OX40 ligand expression on monocytes and serum soluble OX40 ligand in patients with acute coronary syndromes (ACS).METHODS: The present study included healthy controls (n=30), patients with stable angina (SA, n=40) and patients with ACS, including unstable angina (UA, n=50) and acute myocardial infarction (AMI, n=30). The expression of OX40L on platelets was analyzed with flow cytometry and serum level of soluble OX40L (sOX40L) was determined with ELISA.RESULTS: The expression of OX40L on monocytes and serum level of sOX40L were significantly higher in patients with UA and AMI compared to healthy controls and patients with SA. In patients with AMI, sOX40L levels showed no significant increase when compared to patients with UA, while AMI patients had a peak level of sOX40L at 24 h after AMI. Percutaneous transluminal coronary angioplasty (PTCA) induced a marked rise in sOX40L levels in all patients. However, OX40L expression on monocytes showed no difference between patients with PTCA, before and after operation.CONCLUSION: The expression of OX40L may participate the mechanism of ACS and represent a marker for the activity of coronary disease.     

Serum contents of E-selectin,sICAM-1, sVCAM-1 in patients with acute coronary syndrome

AIM:To explore the serum levels of certain adhesion molecules and its significance in acute coronary syndrome(ACS). METHODS:The subjects included 40 patients with acute myocardial infarction(AMI) and 40 patients with unstable angina pectoris (UAP). Among the 80 patients, 60 patients accepted a follow-up 4 months. At the same time we selected 40 controls from people who attended a routine health check in the university. Serum levels of E-selectin, sICAM-1, sVCAM-1 were measured by ELISA.RESULTS:Serum levels of E-selectin, sICAM-1, sVCAM-1 were significantly higher in the ACS group(AMI or UAP) than in the control group. Four months later, the levels of E-selectin, sICAM-1 became significantly lower in the follow-up group than in the ACS group, while sVCAM-1 showed no significant difference. CONCLUSION:Serum levels of E-selectin, sICAM-1 may have certain diagnostic value for ACS, and can be a useful marker reflecting the stability of the disease.     

Association of S100B polymorphisms with systemic lupus erythematosus in Guangxi population

AIM: To explore the association of rs9984765, rs2839356 and rs2186358 polymorphisms in S100B gene with the susceptibility to systemic lupus erythematosus (SLE). METHODS: SLE patients (n=313) and age-and sex-matched healthy controls (n=396) were recruited in this study. The genotypes of the 3 sites were determined by single-base extension PCR (SBE-PCR) and DNA sequencing. RESULTS: No difference between the SLE patients and controls in the genotype and allele frequencies of rs9984765 and rs2186358 was observed. However, the frequency distribution of rs2839356 C allele was significantly different in the 2 groups (P=0.040). Stratification analysis showed that the frequency of rs2839356 C allele was higher in the patients with neurologic disorder than the patients without neurologic disorder (P=0.023).CONCLUSION: S100B gene rs9984765 and rs2186358 polymorphisms may not contribute to the susceptibility of SLE in Guangxi population. The rs2839356 C allele might be correlated with the SLE patients with neurologic disorder.     

Genetic polymorphisms of TLR2 locus in Chinese Cantonese population

AIM: Toll-like receptor 2 ( TLR2 ) was a significant pathogen recognition receptor in innate immune system. The aim of this study was to investigate the distribution of TLR2 polymorphisms in the general population of Chinese Cantonese. METHODS: Peripheral blood samples were collected from 200 unrelated healthy Chinese Cantonese individuals. The functional regions of TLR2 locus, including promoter region and all three exons with their surrounding intronic regions were amplified using PCR and sequenced in a random sample of 24 subjects. TLR2 genotyping in other 176 subjects was performed using PCR-sequence specific primer and PCR. RESULTS: A total of 5 single nucleotides polymorphisms (SNPs) were detected, the two of which were novel. SNPs located in the coding region were all synonymous substitutions. The most common SNP was rs3804099 with the minor allele frequency of 26.3%. One 22 bp insertion/deletion (INDEL) polymorphism was found in exon 1 with the deletion allele frequency of 31.8%. All polymorphic sites were consistent with Hardy-Weinberg equilibrium. Neutrality test suggested that TLR2 in Chinese Cantonese did not significantly deviate from the neutral model. Linkage disequilibrium (LD) analysis showed complete LD between SNP-18945 C/T and SNP-18 883 C/G, and strong LD between SNP rs3804099 and SNP rs3804100. CONCLUSION: This is the first report on the distribution of TLR2 polymorphisms in the general population of China. It provided some ethnic specific polymorphisms, which might help in the further studies of disease association in Chinese.     

Relationship of p21WAF1 gene polymorphisms with protein expression in breast carcinomas

AIM: To investigate the relationship between p21^WAF1gene polymorphisms and protein expression in breast carcinoma. METHODS: Polymerase chain reaction single-strand conformation polymorphisms technique (PCR-SSCP) and immunohistochemical assay of S-P immunostaining technique were used to study polymorphisms of p21^WAF1 and protein expression respectively on the specimen of paraffin-embedded tissues in 100 cases of breast carcinomas and 40 benign breast diseases as control. RESULTS: Two p21^WAF1 gene polymorphisms were found in 18% (18/100) of breast carcinomas and 5% (2/40) of control samples. The difference between the two groups was statistically significant (χ²=3.94, P＜0.05). The positive immunohistochemical reaction of p21^WAF1 protein were found in 50% (50/100) of breast carcinomas and 12.5% (5/40) of control samples. The difference between the two groups was statistically significant (χ²=16.84, P＜0.01). The positive immunohistochemical reaction of p21^WAF1 protein were found in 100% (18/18) of breast carcinomas with p21^WAF1 gene polymorphisms and 39% (32/82) of no p21^WAF1 gene polymorphisms. The difference between two groups was statistically significant (χ²=21.95, P＜0.01). The p21^WAF1 gene polymorphisms were correlated with the protein expression in breast carcinomas (r=0.576, P＜0.01). CONCLUSION: p21^WAF1 gene polymorphisms may create the different copies of mRNA and may make relevant protein molecules.     

Relationship between TNF-α gene polymorphism-308 and type 2 diabetes with periodontitis

AIM: To investigate the gene polymorphism-308 of tumor necrosis factor alpha (TNF-α) in the relation with the susceptibility to periodontitis combined with type 2 diabetes mellitus (DM) and its severity.METHODS: Human DNA samples were obtained from 240 DM patients with periodontitis (periodontitis group, n=120) and without periodontitis (control group, n=120). All patients were genotyped by PCR-RFLP analysis. The frequencies of genotypes and alleles were analyzed. Sulcus bleeding index (SBI) and probing depth (PD) in all patients were measured. The polymorphism-308 of TNF-α gene in the relation with the susceptibility to periodontitis combined with DM and its severity was analyzed.RESULTS: No significant difference in the frequency of genotype and allele was found between DM patients with mild periodontitis and DM patients without periodontitis (P>0.05). However, the frequencies of these genotypes and alleles in DM patients with moderate and severe periodontitis were significantly higher than those in DM patients without periodontitis (P<0.01). The findings showed that the level of TNF-α was associated with SBI and PD (P<0.01).CONCLUSION: TNF-α -308 G/A polymorphism is not associated with DM patients with mild periodontitis, whereas it may have a role in pathogenesis and prognosis of moderate and severe periodontitis combined with DM through TNF-α level.     

Gene cloning,prokaryotic expression,purification and characterization of CAD-caspase activated DNase

AIM：To construct a the plasmid pCool-GST-ICAD/CAD that expresses the CAD protein with biological function in E.coli BL21(DE3)strain.METHODS：CAD was amplified by PCR and inserted into pCool-GST-ICAD to create pCool-GST-ICAD/CAD.The plasmid was checked and transfected into BL21 for expression.The expressed protein were purified by using the GST affinity column,ion exchange and the gel filtration.Protein purity was checked by SDS-PAGE.CAD activity was checked by DNA fragmentation test.RESULTS：Plasmid pCool-GST-ICAD/CAD was constructed as design.The expression of GST-ICAD/CAD protein complex at mg level was observed.After purification,ICAD/CAD protein complex were obtained.Two clear protein bands were shown on SDS-PAGE.The DNA fragmentation test shows that the purified CAD protein indeed had the nonspecific DNase activity.CONCLUSION：Cloning and purification of CAD protein was successful with intact biological activity.