Identification of Ovine KRTAP28-1 and Its Association with Wool Fibre Diameter.

Keratin-associated proteins (KAPs) are a diverse group of proteins and form a matrix that cross-links keratin intermediate filaments in hair and wool fibres. From over 100 KAP genes (KRTAPs) identified in mammalian species, KRTAP25-1 is a high sulphur (HS)-KAP gene, which has recently been described in humans. Here, we report the absence of KRTAP25-1 in sheep, and describe a new HS-KRTAP (named KRTAP28-1) in the chromosome region where KRTAP25-1 was expected to be found. Six variants (A-F) of KRTAP28-1 containing eight single nucleotide polymorphisms (SNPs) and a TG repeat polymorphism were detected. One was positioned 30 bp upstream of the transcription start codon and all the others were non-synonymous SNPs, including a nonsense SNP. The TG repeat polymorphism would lead to a reading frame shift at the carboxyl-terminal end. The effect of KRTAP28-1 on wool traits was investigated with 383 Southdown × Merino-cross lambs from seven sire lines. Of the four genotypes with a frequency of over 5%, lambs of genotypes AB and BD produced wool of a smaller MFD than lambs of genotype BC. This shows that KRTAP28-1 is associated with wool fibre diameter, and that variation in this gene might have potential for use as a gene marker for reducing wool fibre diameter.

Mutations analysis of BRCA1 gene in patients with breast cancer in South Khorasan province, East Iran.

Background: Breast cancer (BC) is well-known as the most common malignancy and the first leading cause of cancer-related death among women worldwide. Evidence suggests that familial history and age are important risk factors for the development of this disease in Iran. Mutations in BRCA1 and BRCA2 genes are the cause of 5 to 10% of hereditary BC. Recent studies demonstrated that mutations in BRCA1 were observed in high-risk women with family histories of BC. However, to date, the mutations have not been elucidated in BC patients from east of Iran. The purpose of this study was to analyze BRCA1 mutations in BC patient from South Khorasan Province. Methods: In the present study, 88 BC patients (11 positive family history) were screened for mutations in BRCA1. The analysis of BRCA1 was carried out by SSCP (single-strand conformation polymorphism) for shorter exons and direct sequencing in the case of longer ones. Results: Twenty-eight of the patients (31.8%) had a synonymous mutation (c.4308T>C) in exon 13. A missense mutation (c. 4837A>G) was presented in exon 16 with a frequency of 56.8 %. In exon 11 three missense mutations were observed, and the frequency rate for c.3113A>G was 32.5%, for c.3119G>A was 5%, and the highest frequency belonged to c.3548A>G with 72.4% in familial BC and 45.4% in the non-familial group. Conclusion: In our study, five mutations were found, but none of the founder mutations were identified in this population. Two missense mutations in exon 16 (56.8%) and in exon 11 (65%) had the highest frequency in South Khorasan Province.

Luteinizing hormone beta gene polymorphism and its effect on semen quality traits and luteinizing hormone concentrations in Murrah buffalo bulls.

OBJECTIVE: Present investigation was aimed to study the Single Nucleotide Variants of the luteinizing hormone beta (LHß) gene and to analyze their association with the semen quality (fresh and post-thawed frozen semen) and luteinizing hormone (LH) concentrations in Murrah buffalo bulls. METHODS: Polymerase chain reaction-single stranded conformational polymorphism (PCR-SSCP) and Sanger sequencing method is used to study genetic variability in LHß gene. LH assay was carried out using enzyme-linked immunosorbent assay method. A fixed general linear model was used to analyze association of single nucleotide polymorphism (SNP) of LHß gene with semen quality in 109 and LH concentrations in 80 Murrah bulls. RESULTS: LHß gene was found to be polymorphic. Total six SNPs were identified in LHß gene g C356090A, g C356113T, g A356701G, g G355869A, g G356330C, and g G356606T. Single Stranded Conformational Polymorphism variants of pattern 2 of exon 1+pattern 2 of exon 2+pattern 1 of exon 3 had highly significant (p<0.01) effect on sperm concentration (million/mL), percent mass motility, acrosome integrity and membrane integrity in fresh and frozen semen whereas significant (p<0.05) effect was observed on percent live spermatozoa. SSCP variants of pattern 2 of exon 1+pattern 2 of exon 2+pattern 1 of exon 3 had highly significant (p<0.01) effect on luteinizing hormone concentrations too. CONCLUSION: The observed association between SSCP variants of LHß gene with semen quality parameters and LH concentrations indicated the possibilities of using LHß as a candidate gene for identification of markers for semen quality traits and LH concentrations in Murrah buffaloes.

ASSOCIATION BETWEEN MURINE DOUBLE MINUTE 2 - T309G polymorphism and recurrence of hepatocellular carcinoma after surgical treatment

Background - Discovery and incorporation of biomarker panels to cancer studies enabled the understanding of genetic variation and its interference in carcinogenesis at molecular level. The potential association between single nucleotide polymorphism (SNP) 309 and increased development of tumors, such as hepatocellular carcinoma, has been subject to several studies. This is the first study on this association conducted in Brazil. Methods - 62 cases of cirrhotic patients with hepatocellular carcinoma surgically treated by partial hepatectomy (HPT) or by liver transplantation (LTX) from 2000 to 2009 at Santa Casa Hospital Complex, in the city of Porto Alegre, were retrospectively analyzed. Tumor samples from surgical specimen were collected and prepared for study in paraffin blocks. Results - Overall survival was 26.7 months in the HPT group and 62.4 months in the LTX group (P <0.01). Overall tumor recurrence was 66.7% in the HPT group (10/15) and 17% in the LTX group (8/47) (X²=13.602, P <0.01). Alpha-fetoprotein levels >200ng/mL, microvascular invasion and histological grade were associated with tumor recurrence (P <0.01). Recurrence rates in each surgical group and analysis of factors associated with tumor recurrence, when stratified for each genotypic pattern, were both not statistically significant. Conclusion - G/G genotype was not associated with tumor recurrence after surgical treatment and it did not show any correlation with other prognostic factors.

Contexto - A descoberta e incorporação de painéis de biomarcadores aos estudos do câncer permitiram o conhecimento de variações genéticas e sua interferência no processo de carcinogênese. A possibilidade de associação do polimorfismo de nucleotídeo simples T309G do gene MDM2 com o aumento da formação de tumores, dentre eles o hepatocarcinoma, tem sido alvo de diversos estudos. Objetivo - Analisar a influência do polimorfismo T309G do gene MDM2 na recidiva tumoral de pacientes cirróticos com hepatocarcinoma submetidos a tratamento cirúrgico. Métodos - Foram analisados retrospectivamente pacientes cirróticos com carcinoma hepatocelular submetidos a tratamento cirúrgico (hepatectomia parcial ou transplante hepático) no período de 2000 a 2009, na Santa Casa Hospital Complex in Porto Alegre, South Brazil. Foram coletadas amostras de fragmentos tumorais da peça operatória (fígado explantado ou segmento hepático), as quais foram preparadas para estudo em bloco parafinado. Resultados - A sobrevida global foi de 26,7 meses para o grupo hepatectomias e 62,4 meses para o grupo transplante hepático (P <0,01), havendo 66,7% de recidiva global no grupo hepatectomias (10/15), e 17% no grupo transplante hepático (8/47) (X²=13,602, P <0.01). Níveis de AFP>200ng/mL correlacionaram-se com a recidiva tumoral em ambos os subgrupos cirúrgicos. Observou-se que 83,3% dos pacientes com recidiva também apresentaram invasão microvascular ao exame anátomo-patológico (P <0,01). Não houve significância estatística quando a recidiva neoplásica foi avaliada para os diferentes genótipos e analisada para cada subgrupo cirúrgico. A análise dos fatores prognósticos relacionados à recidiva do hepatocarcinoma, quando estratificada para cada padrão genotípico, também não se mostrou significante. Conclusão - O nosso estudo revelou que o genótipo G/G não esteve associado à recidiva tumoral após o tratamento cirúrgico, seja nas hepatectomias parciais ou transplante hepático. Além disso, a presença desse genótipo não mostrou correlação com os fatores prognósticos estudados.

Hydroxymethylbilane synthase gene mutations and polymorphisms in Brazilian families with acute intermittent porphyria.

Acute intermittent porphyria (AIP), an autosomal dominant disorder, is caused by a deficiency of hydroxymethylbilane synthase (HMBS). In the present study, we sought to establish a correlation between HMBS activity with the presence of mutations and polymorphisms. Enzyme activity was measured in red blood cells of four Brazilian unrelated AIP families (n = 124) and in blood donors (n = 80). The HMBS mutations in AIP family members were studied by PCR-SSCP followed by direct sequencing. Six intragenic SNPs (1345 G>A, 1500 T>C, 2377 C>A, 2478 A>G, 3581 A>G, and 7064 C>A) were determined by PCR-RFLP. Abnormal SSCP patterns in exons 7, 9, 12, and 15 were observed. DNA sequencing analysis revealed one nonsense mutation, R149X, two missense mutations, G111R and L338P, and one deletion, CT 730-731. All mutation carriers had lower enzyme activity. All polymorphisms, except 2377 C>A and 7064 C>A, showed no significant differences compared with previous reports. Mutation screening allowed the detection of the missense mutation, L338P, and the 730_731delCT deletion, two as yet unreported mutations in Brazilian AIP patients. Our findings also showed a high frequency of 2478 A>G and 3581 A>G polymorphism combinations suggesting that these polymorphisms contributed to enzymatic activity reduction in our study population.

Mutation analysis of TNP1 gene in infertile men with varicocele.

BACKGROUND: Varicocele is associated with the failure of ipsilateral testicular growth and development, and the symptoms of pain and reduced fertility. The highly condensed structure of the sperm nuclear chromatin is provided by proper expression of Transition Nuclear Protein (TNP) genes, so any dysregulational expression of these genes results in abnormal spermatogenesis and infertility. OBJECTIVE: The aim of present study was to assess the association between TNP1 mutations and varicocele in Iranian infertile men. MATERIALS AND METHODS: Analysis of association between TNP1 gene mutation and varicocele phenotype was performed using PCR and Single-Stranded Conformational Polymorphism technique and DNA sequencing in 82 varicocele infertile men and 80 control subjects. RESULTS: Sequence analysis was identified one variant in this gene that found in 15 infertile men and was absent in control group. This variant was a single nucleotide polymorphism that were identified in the intron region of this gene at position g.IVS1+75T>C. CONCLUSION: The effect of this nucleotide substitution in intronic region of the TNP1 gene and their role on expression remains to be determined.

Identification and genetic effect of haplotype in the bovine BMP7 gene.

Bone morphogenetic proteins (BMPs) are peptide growth factors belonging to the transforming growth factor-beta (TGF-ß) superfamily, and some members of the BMP family support white adipocyte differentiation. In this study, we focused on the BMP7 which singularly promotes the differentiation of brown preadipocytes. Haplotypes involving 5 single nucleotide polymorphism (SNP) sites in the bovine BMP7 gene were identified and their effect on body weight was analyzed. 16 haplotypes and 18 combined haplotypes were revealed and the linkage disequilibrium was assessed in the cattle population with 602 individuals representing three main cattle breeds from China. The results showed that haplotypes 3, 10 and 14 were predominant and accounted for 75.64%, 69.85%, and 83.36% in Nanyang, Qinchuan and Jiaxian cattle breeds, respectively. The statistical analyses indicated that the SNP 1, 4, and 5 are associated with the body weight, body length, and heart girth at 12 and 24 months in Nanyang cattle population (P<0.05), whereas there is no significant association between their 16 haplotypes and 18 combined haplotypes. Our results provide evidence that some SNPs and haplotypes in BMP7 are associated with growth traits, and may be utilized as a genetic marker in marker-assisted selection for beef cattle breeding programs.

Screening of C-kit gene Mutation in Acute Myeloid Leukaemia in Northern India.

BACKGROUND: Acute Myeloid Leukaemia (AML) is a cancer of blood-forming cells in bone marrow. C-kit gene is a Receptor Tyrosine Kinase class III (RTK) that is expressed by early hematopoietic progenitor cells and plays an important role in hematopoietic stem cell proliferation, differentiation and survival. It is known that c-kit is a proto-oncogene and the activating c-kit mutations are likely to contribute in the development of leukaemia in humans. Exon 11 of c-Kit gene is the frequent site for mutations in different kinds of tumours. METHODS: In order to determine the frequency and prevalence of exon 11 mutations in 51 AML cases, we have done polymerase chain reaction-single-strand conformational polymorphism followed by direct DNA sequencing. RESULTS: The c-kit mutations in exon 11 were detected in 15.68% (8/51) in AML cases. We have detected totally ten missense mutations in eight AML cases those include Lys550Asn, Tyr568Ser, Ile571Leu, Tyr578Pro, Trp582Ser and Arg588Met and novel missense mutations at codons Ile563Lys and Val569Leu. Mutations at codons Ile571Leu and Trp582Ser was found in two independent cases. CONCLUSION: The presence of c-kit mutations in our study adds to investigative spectrum of AML cases. Since the c-kit mutations are seen in other malignancies, mutations in exon 11 of the c-kit gene might be involve in pathogenesis and represent useful predictive genetic marker in AML. Further studies in larger group of cases possibly will be required to determine the prognostic implications and to investigate how these mutations are co-related to the progression and pathogenesis of AML.

Detection of miRNA gene sequence variations in multiple myeloma and its significance / 白血病·淋巴瘤

Objective To investigate sequence variations of 12 miRNA genes in multiple myeloma(MM) in order to find whether sequence variations in miRNA genes are associated with tumorigenesis and discuss the clinical significance of MM associated with miRNA genes mutations. Methods The miRNA gene mutations in 20 cases of MM, 4 MM-derived cell lines and 20 controls were detected by the methods of polymerase chain reaction single stranded conformation polymorphism (PCR-SSCP) and silver staining technique. Both clinical features and laboratory results were analyzed simultaneously. Results The electrophoretic patterns showed a total of three variations in miR-19a, miR-19b and miRNA-335,which were observed in 3 MM cells (15 %, 3/20). We also found variations of miRNA-335 in MM-derived cell lines KM-3and RPMI8226. However, no sequence alteration in the miRNA genes was observed in our set of controls. One of the three MM patients died, and two of them were detected mutations at the terminal stage of the disease.Conclusion A relative high frequency of miRNA gene mutation was found in MM and MM derived cell lines, which suggests possibility of a main mechanism underlying tumorigenesis. And, detecting miRNA gene mutations in MM might be benefit to evaluate the progression and prognosis of disease.

Gene diagnosis of X-linked spondyloepiphyseal dysplasia tarda / 中华检验医学杂志

ObjectiveTo investigate the molecular pathogenesis of a pedigree of X-linked spondyloepiphyseal dysplasia atarda (SEDL) and to establish methods of gene diagnosis. Methods Clinical diagnosis was made based on height measurement, radiological examination and pedigree analysis. Peripheral blood samples of relevant family members were collected. After genomic DNA extraction, single strand conformation polymorphism (SSCP) followed with DNA sequencing was used to detect SEDL gene exons 36. Microsatellite marker DXS16 was selected for linkage analysis. Results The abnormal electrophoretic bands were detected in exon 4 of probands by PCR-SSCP. A c. 218C ＞ T mutation in exon 4 of SEDL gene was found in three probands, which resulted in a change in amino acid sequence S37L. The heterozygous exon 4 mutation was identified in three carriers, but not in healthy individuals, and no mutations were detect in exon 3, 5 and 6 of probands. Three unmarried young females (Ⅲ10, Ⅳ6 and Ⅳ7) were found to harbor the mutation by DNA sequencing analysis. ConclusionsA c. 218C ＞ T missense mutation in exon 4 of SEDL gene is the cause of molecular pathogenesis of the pedigree. SSCP and DNA sequencing can be used for prenatal gene diagnosis.

SSCP screening of mutation in exon 13 of low density lipoprotein receptor gene in Chinese familial hypercholesterolemia patients / 中华检验医学杂志

Objective To investigate the application of polymerase chain reaction and single strand conformation polymorphism analysis(PCR-SSCP)to the screening of gene mutation of exon 13 of the LDLR gene in familial hypercholesterolemia(FH).Methods Peripheral blood DNA of 16 clinically diagnosed FH patients was extracted and the exon 13 coding region of the LDLR gene was amplified by PCR.PCR products were separated by optimized SSCP electrophoresis and visualized by silver staining.DNA fragments with abnormal mobility were sequenced to determine the nature and position of mutations.Results The SSCP electrophoresis conditions were optimized as 8%polyaerylamide(degree of cross linking 49:1)gel without glycerin at a electrophoresis temperature of 10℃ or 8%polyacrylamide gel with 5%glycerin at room temperature,gel thickness of＜0.4 mm,and a voltage of 5 V/cm.DNA fragments were well resolved with the conditions and sequencing of the abnormal bands resuhed in detections of missense mutations of A606T,D601N,Y601D and G636V together with a synonymous mutation of 1959C→T in 4 patients and a sole synonymous mutation of 1959C→T in other 4 patients.Conclusion PCR-SSCP is an effective method for the screening of exon13 mutations of LDLR gene in FH patients.

Association between SLC25A12 and SCN2A2 gene polymorphisms and autism / 吉林大学学报(医学版)

Objective To investigate the association SLC25A12 and SCN2A2 gene single nucleotide polymorphisms(SNPs) and susceptibility to autism among 105 Japanese family trios consisting of fathers,mothers,and affected offsprings with autism.Methods Genomic DNA was isolated from the whole blood samples.The PCR-single stranded conformational polymorphism(SSCP) technique was used to test genotype of SNPs(rs3770448,rs3769955) at SLC25A12 and SCN2A2 genes.Results The distributions of genotypic and allelic frequencies of rs3770448 and rs3769955 were not deviated from the Hardy-Weinberg equilibrium.The results of transmission disequilibrium test(TDT) indicated that the allelic frequency transmitted from the heterozygote parents didn′t deviate 50%.Conclusion The polymorphism of rs3770448 in the SLC25A12 and rs3769955 in the SCN2A2 locus may not be associated with autism.But the association of the other SNPs at the SLC25A12 and SCN2A2 locus with the illness can not be ruled out.

Gene Mutations in a Chinese Family with Hailey-Hailey Disease / 中华皮肤科杂志

Objective To analyse gene mutation in members of a Chinese family with Hailey-Hailey disease(HHD)and study the relationship between the genotype and clinical features of the disease.Meth-ods Genomic DNA of leucocytes were obtained from members of the Chinese family with HHD including4patients and6normal persons.Ten exons of ATP2C1gene were amplified by polymerase chain reaction(PCR)and the products were analysed by single-strand conformation polymorphism(SSCP)and direct DNA sequencing.Results A novel mutation was identified in this family.The sequence of"TGTAGCCAT"(2068→2076)was substituded by"AGATGGAACA",which caused a frame shift of open reading frame and premature termination codon(PTC)in gene ATP2C1.There was no relationship between the genotypes and the phenotypes.Conclusion Gene mutation of ATP2C1gene at exon21is the cause for HHD in this fami-ly.

No Association of Factor XIII Val34Leu Polymorphism with Primary Intracerebral Hemorrhage and Healthy Controls in Korean Population

The polymorphism in the factor XIII A-subunit gene (FXIII Val34Leu) has been recognized as a risk factor for primary intracerebral hemorrhage (PICH). In addition, FXIII Val34Leu has a significant ethnic heterogeneity. FXIII Val34Leu was detected in 41.7-54.8% of the Westerners, but in 2.5% of the Asians. We aimed to evaluate the prevalence of FXIII Val34Leu in patients with PICH and in healthy controls among Koreans. We recruited 58 in-patients with PICH, defined by brain computed tomography or magnetic resonance imaging, and 48 controls matched for age, sex, and risk factors for cerebrovascular diseases. Genomic DNA was extracted from blood. A 183-bp fragment of exon 2/intron B of the factor XIII Asubunit gene was amplified by polymerase chain reaction (PCR). The factor XIII genotype was determined through a single-stranded conformational polymorphism. Fifty-eight patients and 48 controls showed the same band patterns on SSCP. In addition, we directly sequenced six random-selected DNA segments using DNA auto-sequencer. In conclusion, the results of this study suggest that FXIII Val34Leu be absent or rare both in patients with PICH and in healthy controls among Koreans.

Polymorphism in insulin receptor gene exon 17 in women with polycystic ovary syndrome / 中华妇产科杂志

Objective To investigate insulin receptor (INSR) genotype exon 17 frequencies in women with polycystic ovary syndrome(PCOS) and to elucidate its role in the pathogenesis of PCOS. Methods The study involved 33 women with PCOS and 28 healthy control women who were genotyped for polymorphism of INSR gene exon 17 by single strand conformation polymorphism (SSCP) analysis. Body mass index (BMI), insulin sensitive index (ISI), the expression of INSR beta subunit, and serum concentration of luteinizing hormone(LH), total testosterone between the genotypes were compared. Results (1) The T -to- C mutation was observed in the INSR gene exon 17 (1008 bp). The frequency of the C/C genotype was significantly higher in patients (39%) than in the controls (11%) (P

embB mutations in Mycobacterium tuberculosis ethambutol-resistant isolates / 中华检验医学杂志

Objectives To understand the mutations of embB genes in M. tuberculosis isolates, and to evaluate their clinical value. Method 102 clinical isolates were identified for their mycobacterial species, and then analyzed their embB genes with PCR SSCP, PCR RFLP, and PCR direct sequencing. Results Mycobacterium tuberculosis strain H 37 R v was used as a control. 102 clinical isolates all had the same 16S rDNA SSCP profiles as M. tuberculosis . Forty one drug sensitive isolates had normal embB SSCP and RFLP profiles. Of 61 ethambutol resistant isolates, 23 (37.7%) displayed abnormal embB SSCP profiles. Eight isolates had abnormal RFLP profiles. All embB mutations situated at codon 306, whose EMB MICs were more than 20 ?g/ml. Eight isolates had ATG to ATA or ATT mutations at codon 306. Thirty isolates had ATG to GTG or CTG mutations at codon 306, whose EMB MICs were more than 30 ?g/ml. Conclusions Ethabutol resistances in some M. tuberculosis isolates were due to mutations on embB genes. PCR SSCP and PCR RFLP method might become a simple and rapid diagnostic test for genotypes of M. tuberculosis ethabutol resistance.

Detecting p53 gene mutation of breast cancer and defining differences between silver staining PCR-SSCP and immunohistochemical staining

This study detects and defines the patterns of p53 gene mutations in breast cancers. We analyse p53 gene mutations through comparing the results of single-strand-conformation-polymorphism (SSCP) and immunohistochemistry (IHC), and we try to define the differences between the results of SSCP and IHC. Twenty-seven fresh primary breast cancer tissues and eight normal breast tissues were studied. The IHC was done with the usual streptavidin-biotin peroxidase complement method by using monoclonal antibody DO-7. The results of staining was scored. The SSCP method was done by using Cold SSCP Electrophoresis System. Overexpressions of p53 protein were seven (25.9%) among 27 cancer cases on IHC. Four (57.1%) of seven cases were positive in SSCP. In SSCP, the mutations were detected in 10 (37%) among 27 cancer cases. The mutations were two in exon 5, one in exon 8, and seven cases in exon 7. All of 10 mutations were proved by sequencing analysis. Of them, only four (40%) were positive in IHC. We consider the IHC as a screening method for p53 gene mutations.

Detection of H ras mutation in urine exfoliated cells complements cytology in 48 TCC patients / 中华检验医学杂志

Objective To determine the value of detection of H ras oncogene mutation in urine exfoliated cells as clinical indicator of tumor presence, recurrence and stage.Methods Point mutation at codon 12 of H ras gene was assayed by polymerase chain reaction followed by analysis of single strand conformation polymorphism in urine exfoliated cells from 48 patients with transitional cell carcinoma before operation and 28 patients with non urothelial cancer or normal individuals. The mutation was further confirmed by dideoxy mediated chain termination method of DNA sequencing. Cytology analysis was carried out simultaneously. Bladder tumor specimens were obtained from 48 patients during operation, and histologically elevated for tumor content and grading.Results 48%(23 of 48) of the patients were detected by their aberrant band in SSCP. All aberrant bands displayed a mutant H ras sequence, where 15% (7 of 48) of the patients displayed, apositive cytological analysis. Analysis of abnormalities with tumor stage revealed that the greater detection of high pathological stage (Ⅲ Ⅳ) compared with low stage (Ⅰ Ⅱ) was related to the recurrence of transitional cell carcinoma.Conclusion Our results suggest that the detection of H ras mutations may be of clinical value in the detection of TCC.

The study of RHD gene mutation in weak D / 대한수혈학회지

BACKGROUND: The weak D is characterized serologically by a weak or negative agglutination reaction with polyclonal anti-D in an immediate-spin test and agglutination is enhanced in the indirect antiglobulin test. Weak D has a lower number of D antigen or weaker antigen density than are normal D positive red cells. Here we studied the cause of weak D antigenicity at genetic level and compared to that of normal D RBCs. METHODS: The amplification of RHD gene and RHCcEe gene site was done in normal D(n=20), weak D(n=8), D negative group(n=20) by polymerase chain reaction and by based on D typing in these individuals compared to that of serologic D typing. In addition, to detect RHD gene mutation and nucleotide sequence difference of weak D group compared to normal D RBCs, single stranded conformational polymorphism PCR was simultaneuosly perfomed in two group by RHD amplified product(189 bp). We analysis the correlation RHD genotyping and serological phenotyping, and also analysis the difference of nucleotide sequence between two group in genetic level. RESULTS: The RHD genotyping was completely matched normal D(n=20), D negative group(n=20) but weak D group(n=8) showed same genotype of normal D RBCs. In single stranded conformational polymorphism PCR, weak D phenotypes does not show any abnormalities at the genomic level when compared to the RHD gene in normal D phenotypes. CONCLUSIONS: RHD PCR showed good correlation with conventional serologic test but weak D genotype was same as that of normal D RBCs. The weaker immunogenicity of weak D is not explained by genomic DNA difference itself.

Polymorphism within the interleukin-10 receptor cDNA gene in patients with systemic lupus erythematosus / 中华皮肤科杂志

Objective To assess the association between polymorphism within the interleukin-10 receptor cDNA gene (IL-10R) and Chinese patients with systemic lupus erythematosus (SLE). Methods The IL-10R genotypes of 94 SLE patients and 80 healthy subjects were examined by reverse transcription-polymerase chain reaction-single strand conformation polymorphism method (RT-PCR-SSCP), RT-PCR-restriction fragment length polymorphism (RFLP) and DNA sequencing. Results There were significant differences in the IL-10R2 genotype frequencies of these two groups. The IL-10R2 G520/G520 genotype increased the risk of developing SLE (OR = 0.515, 95% CI 0.414-0.579, P = 0.004) and individuals who had G520/A520 genotype also had a higher susceptibility to SLE (OR = 1.968, 95% CI 0.981-3.949, P = 0.055). There was no significant association between SLE and IL-10R1 genotypes. The risk of developing SLE was detected in the individuals who had the combination of IL-10R1 G241/G241 and IL-10R2 G520/G520 (OR = 0.515, 95% CI 0.444-0.597, P = 0.004). Conclusion The IL-10R2 genotypes of G520/G520 and G520/A520 as well as the combination of genotypes IL-10R1G241/G241 and IL-10R2 G520/G520 may increase the susceptibility to SLE in Chinese people.