Screening Of Single Nucleotide Polymorphisms Research Articles

Rapid detection of type II diabetes mellitus in Saudi patients via simultaneous screening of multiple SNPs

A large-scale DNA-based assay, namely SNaPshot, was developed and validated for the determination of allelic polymorphisms of genes associated with type II diabetes (T2D) mellitus disease in Saudi patients. The new approach is alternative to phenotyping and physiological diagnosis. The assay includes single nucleotide polymorphisms (SNPs) of seven most common genes namely, GNB-3, GCK, SLC30A, KNJ11, TCF7L2, CDKN2A/B and HNF4A. The study proved that the new genotyping assay is rapid, easy to perform and robust for screening of high risk T2D family members. The new assay can be applied in genetic testing as an important tool for identifying significant genetic factors that help in predicting diabetes at early stages of life to avoid subsequent complications. This assay can also be used to evaluate the gene risk variants in populations of other geographic origins.

Assessment of the Effect of HMGCR Variant Alleles on Response to Atorvastatin Treatment in Type 2 Diabetic Egyptian Patients

Use of 3 hydroxyl-3-methylglutaryl-3-coenzyme A reductase (HMGCR) inhibitors, or statins, reduces the progress and the complications of diabetes mellitus by modifying the lipid profile. The aim of this study was to assess the association between variation in statin response and the single-nucleotide polymorphism (SNP) rs12916 C/T in the gene encoding HMGCR, a rate limiting enzyme in cholesterol synthesis and the target enzymatic reaction of statins. A total of 96 Egyptian patients with type 2 diabetic dyslipidemia were treated with atorvastatin 20 mg/day for 3 months. Total cholesterol, triglyceride, low density lipoprotein–cholesterol (LDL-C), high density lipoprotein–cholesterol (HDL-C) plasma concentrations were measured at baseline and at the end of the treatment period together with genetic screening for SNP rs 12916 C/T. It was found that individuals with the CC genotype showed a mean reduction in LDL-C level of about 31.57 ± 87.52 mg/dl (p=0.24), while the reduction in the CT and TT genotypes was 28.50 ± 72.74 mg/dl (P=0.04) and 26.15 ± 101.45 mg/dl (p=0.06), respectively. It is concluded that there is no significant association between this SNP rs12916 C/T and response to atorvastatin therapy in type 2 dyslipidemic diabetic Egyptian patients.

Association Between Chromosome 4Q25 Polymorphism RS2200733 and the Incidence of Atrial Fibrillation in Bulgarian Patients

Summary Atrial fibrillation (AF) is the commonest type of arrhythmia seen in everyday clinical practice, which leads to a significant increase in both morbidity and mortality. Its incidence increases with age and tends to turn into an epidemic. The cause of AF in 10-20% of cases remains unknown. Several mutations and polymorphism that might be responsible for the development of AF have been found, including single nucleotide polymorphisms (SNPs) - rs2200733 and rs10033464 in the long arm of the fourth chromosome. These polymorphisms are selected o the basis of genome- wide association study in Iceland from 2007, the results from which were later confirmed in 4 other large populations. The rs2200733 is a common noncoding polymorphism, described in National Center for Biotechnology Information (NCBI) database dbSNP like NC_000004.12:g.110789013C>T, with a frequency of the less common allele between 0.1 and 0.24. In order to investigate the association between the rs2200733 polymorphism in chromosome 4q25 and the development of AF, we studied the frequency of this polymorphism in patients with heart diseases from the Pleven region, and thus evaluate the relationship between the individual genotype and the clinical condition of the patients. We carried out a case-control study on 80 patients: 40 with AF and 40 without AF- from the Pleven region. None of these had structural heart disease. The study was conducted between November 2015 and November 2017. With deoxyribonucleic acid (DNA) analysis, we determined rs2200733 polymorphism, using a TaqMan-based polymerase chain reaction (PCR). The Cochran-Armitage trend test, the Chi-Squared Pearson correlation, Fisher test we used confirmed the statistically significant association between the rs2200733 polymorphism in chromosome 4q25 and the development of AF. In the population examined, the genotypic frequencies were as follows: CC - 45 (56.2%), CT - 19 (23.8%), TT - 16 (20%), with value of Chi-Square (χ2) 24.496, df=2, p<0.001. Screening for SNPs could be a useful marker for the detection of patients predisposed to AF.

Interactions Between Regulatory Variants in CYP7A1 (Cholesterol 7α-Hydroxylase) Promoter and Enhancer Regions Regulate CYP7A1 Expression.

CYP7A1 (cholesterol 7α-hydroxylase) catalyzes the rate-limiting step in bile acid biosynthesis from cholesterol-a main pathway for cholesterol removal from the body. CYP7A1 single-nucleotide polymorphisms (SNPs) are associated with total cholesterol and LDL (low-density lipoprotein) levels, risk of cardiovascular diseases, and other phenotypes; however, results are inconsistent, and causative variants remain uncertain, except for a frequent promoter SNP (rs3808607). We used chromatin conformation capture (4C assay), chromatin immunoprecipitation qPCR assay in hepatocytes, and CRISPR (clustered regularly interspaced short palindromic repeats)-mediated genome editing in hepatocellular carcinoma cell line cells to identify regulatory regions for CYP7A1. We then screened for SNPs located in regulatory regions, testing effects on reporter gene assays and on hepatic CYP7A1 expression by measuring allelic mRNA expression imbalance. 4C assays showed several regions interacting with CYP7A1 promoter. CRISPR-mediated genome editing in hepatocellular carcinoma cell line cells revealed a novel CYP7A1 enhancer and a repressor region, located >10 kb downstream of the CYP7A1 promoter. SNP screening with an allelic mRNA expression imbalance in human livers and reporter gene assays identified a frequent functional SNP (rs9297994) located in the downstream CYP7A1 enhancer region. SNP rs9297994 is in high linkage disequilibrium with promoter SNP rs3808607 but has opposite effects on CYP7A1 mRNA expression. Their combined effects using a 2-SNP model robustly associate with hepatic CYP7A1 mRNA expression, ranging >2 orders of magnitude. Moreover, only the 2-SNP model, but not each SNP alone, is significantly associated with LDL levels, risk of coronary artery disease, statin response, and diabetes mellitus in several clinical cohorts, including CATHGEN (Catheterization Genetics) and Framingham. Two interacting regulatory SNPs modulate CYP7A1 expression and are associated with risk of coronary artery disease and diabetes mellitus.

Subclinical thyroid disease and single nucleotide polymorphisms in reproductive-age women in areas of Shanxi Province, China, where iodine exposure is excessive.

When iodine intake is in excess, a susceptible population that has a genetic predisposition will have an increased risk of hypothyroidism or autoimmune thyroiditis. This study evaluated the vulnerability to iodine excess and subclinical thyroid disease through screening of single nucleotide polymorphisms (SNPs) in reproductive-age women to provide evidence to be used for the prevention of subclinical thyroid disease. In Shanxi province, four areas where a range of iodine exposures from low to high were chosen in each region, 60 women were anticipated to enrol, including 20 pregnant women, 20 lactating women, and 20 non-pregnant, non-lactating women. Genotyping was performed using whole-blood samples, and the genotypes of 21 SNPs were determined and compared among areas with different water iodine and between controls and patients with subclinical thyroid disease. In total, 241 participants were enrolled. Among the 21 candidate SNPs, no difference was found among areas with various water iodine, whereas, TG (rs2252696), TSHR (rs4903957), CTLA-4 (rs231775), CAPZB (rs1472565), PDE4D (rs27178), and HLA (rs2517532) were significantly associated with various subclinical thyroid diseases; in particular, the PDE4D (rs27178), ad hoc TT allele, was associated with all examined subclinical thyroid diseases. Vulnerability to subclinical thyroid diseases is influenced by the presence of gene polymorphisms. There is a need for screening of suspected genes to effectively prevent and reduce the occurrence of thyroid diseases. People with the TT allele in PDE4D (rs27178) should be made aware of an increased risk of subclinical thyroid disease.

Genomic Insights Into the Multiple Factors Controlling Abdominal Fat Deposition in a Chicken Model.

Genetic selection for an increased growth rate in meat-type chickens has been accompanied by excessive fat accumulation particularly in abdominal cavity. These progressed to indirect and often unhealthy effects on meat quality properties and increased feed cost. Advances in genomics technology over recent years have led to the surprising discoveries that the genome is more complex than previously thought. Studies have identified multiple-genetic factors associated with abdominal fat deposition. Meanwhile, the obesity epidemic has focused attention on adipose tissue and the development of adipocytes. The aim of this review is to summarize the current understanding of genetic/epigenetic factors associated with abdominal fat deposition, or as it relates to the proliferation and differentiation of preadipocytes in chicken. The results discussed here have been identified by different genomic approaches, such as QTL-based studies, the candidate gene approach, epistatic interaction, copy number variation, single-nucleotide polymorphism screening, selection signature analysis, genome-wide association studies, RNA sequencing, and bisulfite sequencing. The studies mentioned in this review have described multiple-genetic factors involved in an abdominal fat deposition. Therefore, it is inevitable to further study the multiple-genetic factors in-depth to develop novel molecular markers or potential targets, which will provide promising applications for reducing abdominal fat deposition in meat-type chicken.

Electrochemical screening of single nucleotide polymorphisms with significantly enhanced discrimination factor by an amplified ratiometric sensor

The detection of single nucleotide polymorphisms (SNPs) is of great clinical significance to the diagnosis of various genetic diseases and cancers. In this work, the development of an ultrasensitive ratiometric electrochemical sensor for screening SNP with a significantly enhanced discrimination factor is reported. The ferrocene (Fc) and methylene blue (MB) dual-tagged triple helix complex (THC) probes are self-assembled on the gold electrode to construct the sensing interface. The addition of the mutant p53 gene causes the disassembly of the THC probes with the release of the Fc-tagged sequence and the folding of the MB-labeled sequence into a hairpin structure, causing the change in the current response ratio of MB to Fc for monitoring the mutant p53 gene. Such ratio is dramatically enhanced by the toehold-mediated displacement reaction-assisted target recycling amplification with the presence of an assistance hairpin sequence. With the significant signal amplification and the advantageous specificity of the THC probes, sub-femtomolar detection limit and a highly enhanced SNP discrimination factor for the mutant p53 gene can be obtained. Besides, the proof-of-demonstration application of the sensor for diluted real samples has been verified, offering such sensor new opportunities for monitoring various genetic related diseases.

Spatio-temporal distribution of PfMDR1 polymorphism among uncomplicated Plasmodium falciparum malaria cases along international border of north east India

PfMDR1 single nucleotide polymorphisms (SNP) are good correlate markers for antimalarial drug resistance worldwide. Present study is a comprehensive view of screening of PfMDR1 polymorphism to antimalarials practiced with geography and time. Study sites Mizoram, Tripura, Meghalaya chosen are at multivariate drug pressure due to cross border migration and transmission. Mizoram is gateway to south east Asia through Myanmar whereas Tripura, Meghalaya share porous border with Bangladesh. Baseline finger pricked blood stained filter paper for confirmed uncomplicated Plasmodium falciparum infected patients (year 2015) were obtained from National Institute of Malaria Research, New Delhi, India. PfMDR1 polymorphism for codon N86Y, Y184F, D1246Y was determined by PCR-RFLP, further confirmed by sequencing. There observed marked predominance of Plasmodium isolates with PfMDR1 wild type alleles for all codons under study i.e. 86, 184, 1246. Spatially, Plasmodium isolates from Mizoram were most diverse with co-existence of PfMDR1 genotype with NYD, YYD, NFD haplotypes, followed by Tripura. Isolates from Meghalaya were of all NYD haplotype. Reports, referring to screening of PfMDR1 SNPs to CQ/SP/AS-SP across India, were archived. Temporal study show distinct rise in proportion of PfMDR1 wild type N86 allele since introduction of Artemether-Lumefantrine as first line antimalarial. Hence spatio-temporal screening of Plasmodium population with PfMDR1 single nucleotide polymorphism accounts for its association with antimalarial susceptibility and validate PfMDR1 SNPs as antimalarial drug resistant marker.

The Mutation Rate and the Age of the Sex Chromosomes in Silene latifolia

Many aspects of sex chromosome evolution are common to both plants and animals [1], but the process of Y chromosome degeneration, where genes on the Y become non-functional over time, may be much slower in plants due to purifying selection against deleterious mutations in the haploid gametophyte [2, 3]. Testing for differences in Y degeneration between the kingdoms has been hindered by the absence of accurate age estimates for plant sex chromosomes. Here, we used genome resequencing to estimate the spontaneous mutation rate and theage of the sex chromosomes in white campion (Silene latifolia). Screening of single nucleotide polymorphisms (SNPs) in parents and 10 F1 progeny identified 39 de novo mutations and yielded a rate of 7.31× 10-9 (95% confidence interval: 5.20× 10-9 - 8.00× 10-9) mutations per site per haploid genome per generation. Applying this mutation rate to the synonymous divergence between homologous X- and Y-linked genes (gametologs) gave age estimates of 11.00 and 6.32 million years for the old and young strata, respectively. Based on SNP segregation patterns, we inferred which genes were Y-linked and found that at least 47% are already dysfunctional. Applying our new estimates for the age of the sex chromosomes indicates that the rate of Y degeneration in S.latifolia is nearly 2-fold slower when compared to animal sex chromosomes of a similar age. Our revised estimates support Y degeneration taking place more slowly in plants, a discrepancy that may be explained by differences in the life cycles of animals and plants.

Screening of Single Nucleotide Polymorphism in CD95 (APO-1/FAS) Promoter Region (G-1377A) in Children with Acute Leukemia

CD95 is a cell surface receptor involved in apoptotic signal transmission. Deregulation of this pathway results in down regulation of apoptosis and subsequent persistence of a malignant clone. A single nucleotide polymorphism (SNP) resulting in guanine-to-adenine (G > A) transition in the CD95 promoter region (position ﹣1377) is thought to reduce stimulatory protein 1 (SP1) transcription factor binding and decrease CD95 expression. The purpose of this study was to examine a genetic polymorphism in the core promoter of CD95 is associated with the frequency. G-1377A in the CD95 promoter region was genotyped by polymerase chain reaction and restriction endonuclease analysis and sequenced of the CD95 gene (Beckman Coulter, USA). Among 146 patients, CD95 G-1377A (rs2234767) SNP carriers frequencies has been identified as 25% GA (n:37) and AA 4% (n:6), respectively. This polymorphism of the distribution of the CD95 gene in acute leukemic children will be a guide for future studies.

Screening of single nucleotide polymorphism in CD95 (APO-1/FAS) promoter region (G-1377A) in children with acute leukemia

Screening of single nucleotide polymorphism in <i>CD95</i> (APO-1/FAS) promoter region (G-1377A) in children with acute leukemia

Comparative screening of single nucleotide polymorphisms in beta-casein and and #954;-casein gene in different Yak populations of India

Comparative screening of single nucleotide polymorphisms in beta-casein and and #954;-casein gene in different Yak populations of India

Association of WWOX rs9926344 polymorphism with poor prognosis of hepatocellular carcinoma.

Introduction: The WW domain-containing oxidoreductase (WWOX), widely expressed in human tissues, is considered as a tumor suppressor gene and plays an important role in the incidence and progression of human cancer, HCC included. This study was to investigate the correlation between single nucleotide polymorphisms (SNPs) of the WWOX gene and the prognosis of hepatocellular carcinoma (HCC) patients.Materials and Methods: After a total of 152 HCC patients were recruited, 8 cases with tumor recurrence within 2-years after operation and 8 cases without recurrence were selected randomly for SNP genotyping and screening using Affymetrix Array 6.0. And then we confirmed candidate SNPs in the remaining 136 patients by time-of-flight mass spectrometry (TOF-MS).Results: In total, 32 SNPs were screened and identified as candidate SNPs with one SNP in particular, (rs9926344), being further verified to be valuable. We found that AA+AG genotype and A allele of WWOX rs9926344 were significantly associated with recurrent risk of HCC (p=0.002 and p=0.001, respectively). The Kaplan-Meier curve showed that patients carrying rs9926344 AA +AG genotype had poor RFS (P=0.004) and OS (P=0.005) compared to those carrying GG genotypes. The multivariate COX regression analysis showed that the AA+AG genotype were an independent prognostic factor for tumor recurrence (HR 1.787, 95% CI 1.042-3.064, P=0.035). Furthermore, IHC analysis showed that the WWOX protein down-regulation is more frequent in patients with AG genotype compared to those with GG genotype (P=0.023).Conclusion: Our findings indicate that WWOX rs9926344 polymorphism is positively correlated with tumor recurrence and can be used as an independent prognostic marker for HCC patients after operation.

Allele specific LAMP- gold nanoparticle for characterization of single nucleotide polymorphisms

Due to their relevance as disease biomarkers and for diagnostics, screening of single nucleotide polymorphism (SNPs) requires simple and straightforward strategies capable to provide results in medium throughput settings. Suitable approaches relying on isothermal amplification techniques have been evolving to substitute the cumbersome and highly specialized PCR amplification detection schemes. Nonetheless, identification of an individual’s genotype still requires sophisticated equipment and laborious methods.Here, we present a low-cost and reliable approach based on the allele specific loop-mediated isothermal amplification (AS-LAMP) coupled to ssDNA functionalized gold nanoparticle (Au-nanoprobe) colorimetric sequence discrimination. The Au-nanoprobe integration allows for the colorimetric detection of AS-LAMP amplification product that can be easily interpreted in less than 15min. We targeted a clinical relevant SNP responsible for lactose intolerance (-13910C/T dbSNP rs#: 4988235) to demonstrate its proof of concept and full potential of this novel approach.

Gene-specific sex-linked genetic markers in date palm (Phoenix dactylifera L.)

During the past decade, there have been numerous attempts to identify sex-linked molecular genetic markers that can be used to discriminate among male and female trees in date palm (Phoenix dactylifera L.). In our approach to address this biological problem, we applied a comparative genomics approach and used a candidate sex-linked Tormozembryo Defective (TOZ19) gene found to be male-specific in aspen. Using BLAST against the date palm genome assembly, we found a putative Transducin Beta-like Protein 3 (TBL3) gene in date palm that was highly homologous to the TOZ19 gene and sequenced it in three male and four female trees from four economically important date palm cultivars from Egypt. Based on the obtained multiple nucleotide sequence alignments, male- and female-specific date palm haplotypes were identified by screening single nucleotide polymorphisms (SNPs). Subsequently, a respective gene fragment in additional five date palm samples comprising three females and two males were cloned and sequenced to independently confirm the previously identified putative sex-linked SNPs. The three putative sex-linked SNPs can be used now to discriminate male and female date palms at their seedling stage. This will further enhance and pave the way for commercial date palm cultivation through seeds. The identified molecular markers are relatively easy, cheap, fast, and reproducible sex identification tools. Female date palms are either homozygous or heterozygous, while male date palms are hemizygous at the putative sex-linked loci.

Association of Vitamin D Receptor Polymorphisms With the Risk of Nonmelanoma Skin Cancer in Adults

Protective effects of UV-B radiation against nonmelanoma skin cancer (NMSC) are exerted via signaling mechanisms involving the vitamin D receptor (VDR). Recent studies have examined single-nucleotide polymorphisms (SNPs) in the VDR, resulting in contradictory findings as to whether these polymorphisms increase a person's risk for NMSC. To examine whether the polymorphisms in the VDR gene are associated with the development of NMSC and the demographic characteristics of the participants. This case-control study recruited 100 individuals who received a diagnosis of and were being treated for basal cell carcinoma or squamous cell carcinoma (cases) and 100 individuals who were receiving treatment of a condition other than skin cancer (controls) at the dermatology clinics at the Kirklin Clinic of the University of Alabama at Birmingham Hospital between January 1, 2012, and December 31, 2014. All participants completed a questionnaire that solicited information on skin, hair, and eye color; skin cancer family history; and sun exposure history, such as tanning ability and number of severe sunburns experienced throughout life. Blood samples for DNA genotyping were collected from all participants. Polymorphisms in the VDR gene (ApaI, BsmI, and TaqI) were assessed to determine the association of polymorphisms with NMSC development and demographic characteristics. χ2 Analysis was used to determine whether genotype frequencies deviated significantly from Hardy-Weinberg equilibrium. Logistic regression was used to calculate odds ratios (ORs) and associated 95% CIs for the identification of factors associated with NMSC diagnosis. A model was created to predict NMSC diagnoses using known risk factors and, potentially, VDR SNPs. A total of 97 cases and 100 controls were included. Of the 97 cases, 68 (70%) were men and 29 (30%) were women, with a mean (SD) age of 70 (11) years. Of the 100 controls, 46 (46%) were men and 54 (54%) were women, with a mean (SD) age of 63 (9) years. All participants self-identified as non-Hispanic white. A model including age, sex, and skin color was created to most effectively predict the incidence of skin cancer. Risk factors that significantly increased the odds of an NMSC diagnosis were light skin color (OR, 5.79 [95% CI, 2.79-11.99]), greater number of severe sunburns (OR, 2.59 [95% CI, 1.31-5.10]), light eye color (OR, 2.47 [95% CI, 1.30-4.67]), and less of an ability to tan (OR, 2.35 [95% CI, 1.23-4.48]). The risk factors of family history of NMSC (OR, 1.66 [95% CI, 0.90-3.07]) and light hair color (OR, 1.17 [95% CI, 0.51-2.71]) did not reach statistical significance. Participants with the BsmI SNP were twice as likely to develop NMSC than participants with no mutation (OR, 2.04 [95% CI, 1.02-4.08]; P = .045). The results of this study are especially useful in the early treatment and prevention of NMSC with chemopreventive agents (for those with the BsmI SNP). A screening for the BsmI SNP may emphasize the importance of sun protection and facilitate skin cancer prevention and, therefore, decrease the skin cancer burden.

Toward Identification of Black Lemma and Pericarp Gene Blp1 in Barley Combining Bulked Segregant Analysis and Specific-Locus Amplified Fragment Sequencing.

Black barley is caused by phytomelanin synthesized in lemma and/or pericarp and the trait is controlled by one dominant gene Blp1. The gene is mapped on chromosome 1H by molecular markers, but it is yet to be isolated. Specific-locus amplified fragment sequencing (SLAF-seq) is an effective method for large-scale de novo single nucleotide polymorphism (SNP) discovery and genotyping. In the present study, SLAF-seq with bulked segregant analysis (BSA) was employed to obtain sufficient markers to fine mapping Blp1 gene in an F2 population derived from Hatiexi No.1 × Zhe5819. Based on SNP screening criteria, a total of 77,542 polymorphic SNPs met the requirements for association analysis. Combining two association analysis methods, the overlapped region with a size of 32.41 Mb on chromosome 1H was obtained as the candidate region of Blp1 gene. According to SLAF-seq data, markers were developed in the target region and were used for mapping the Blp1 gene. Linkage analysis showed that Blp1 co-segregated with HZSNP34 and HZSNP36, and was delimited by two markers (HZSNP35 and HZSNP39) spanning 8.1 cM in 172 homozygous yellow grain F2 plants of Hatiexi No.1 × Zhe5819. More polymorphic markers were screened in the reduced target region and were used to genotype the population. As a result, Blp1 was delimited within a 1.66 Mb on chromosome 1H by the upstream marker HZSNP63 and the downstream marker HZSNP59. Our results demonstrated the utility of SLAF-seq-BSA approach to identify the candidate region and discover polymorphic markers at the specific targeted genomic region.

Screening for single nucleotide polymorphisms in highly degraded DNA by using the amplified fragment length polymorphism technique

Short tandem repeat (STR) analysis is generally used for human identification of forensic samples; however, standard STR analysis sometimes fails to generate full profiles since DNA is frequently degraded by various environmental factors. Recently, single nucleotide polymorphism (SNP) analysis has attracted attention for human identification since the shorter amplicons are better suited for degraded samples. Though various SNP loci are used for analysis of degraded samples, it is unclear which ones are more appropriate. To identify SNPs that were resistant to degradation, we artificially degraded DNA obtained from the buccal swabs of six volunteers and the K562 cell line by heat treatment. Subsequently, the amplified fragment length polymorphism (AFLP) technique was used for SNP screening. We focused on the AFLP bands detected in both the heat-treated and untreated samples, and DNA extracted from these bands was directly sequenced. DNA degradation increased as the duration of heat treatment increased, and no STR profiles could be generated after 6h of heat treatment. When the AFLP band patterns were compared between 6h heat-treated and untreated samples, eight common bands were detected. The sequences of the DNA fragments of these common bands had higher adenine-thymine (A-T) content and included 17 SNPs. The SNPs detected in the heat-treated and untreated samples were considered to be resistant to degradation. Although there was a little information available in databases regarding the nine SNPs identified in this study, this study shows that some of these SNPs might be useful for human identification of extremely degraded DNA.

Characterization of NPHS2 gene polymorphisms associated to steroid resistance nephrotic syndrome in Indian children

Nephrotic syndrome (NS) is the common glomerular disease in children. These children are treated with steroids, depending upon their behavior. They are either steroid sensitive (SSNS) or steroid resistant (SRNS). NPHS2 gene mutants are linked to the risk of autosomal recessive SRNS and in some cases to SSNS. The present study has been performed to screen single nucleotide polymorphisms (SNPs) of the NPHS2 gene in a group of 90 Indian children suffering with NS (30 SSNS, 30 SRNS and 30 Controls) by PCR method followed by direct exon sequencing. Effect of SNPs on fold expression changes at transcript level of podocin was checked using quantitative real time PCR (qRT-PCR). SNPs identified through sequencing helps to carry out in-silico analysis. Overall 17 SNPs were identified in NPHS2 gene where 6 were found novel. Three missense SNPs p.R299Q, p.P20L and p.G35D were also identified in this population where SNP, p.G35D was found novel. In addition to sequencing analysis, results of in silico analysis shows that a mutant with these three missense SNPs has least ligand binding efficiency compared to native model. Moreover the significant observation of this study included two intronic SNPs c.451+23C>T and c.451+58A>T present in SRNS group of patients. These SNPs has shown high level of clinical significance within genomic and allelic frequency along with haplotypes and linkage disequilibrium count. The qRT-PCR analysis shows, down expression of podocin protein at transcript level in SRNS patients compared to SSNS patients. All these results support the fact that SNPs present in this population could affect the protein structural stability. Thus it is concluded that the polymorphisms predicted in this study might be disease causing in the NPHS2 gene and may have influence on the therapeutic response of NS patients.

Nonsynonymous changes of equine lentivirus receptor-1 (ELR1) gene in amino acids involved in the interaction with equine infectious anemia virus (EIAV)

Equine lentivirus receptor-1 (ELR1) has been characterized as the specific functional receptor that mediates equine infectious anemia virus (EIAV) entrance to horse macrophages. This receptor is tumor necrosis factor receptor superfamily member 14 (TNFRSF14). The aim of this study was to investigate the occurrence of allelic variants in the coding sequence of equine TNFRSF14 gene by screening for single-nucleotide polymorphisms (SNPs) in different equine populations. Forty seven horse samples were randomly selected from a reservoir of EIAV-seropositive and seronegative samples collected from different outbreaks and regions of Argentina. DNA samples were scanned via PCR and direct sequencing of exon 3 and exon 5 of TNFRSF14 gene. A total of 21 SNPs were identified, of which 11 were located in coding sequences. Within exon 5, four SNPs caused nonsynonymous substitutions, while two other SNPs caused synonymous substitutions in crucial residues (Ser112 and Thr114) implicated in the interaction with EIAV. Despite some of exon 5 variants occurred exclusively in EIAV-positive or EIAV-negative horses, critical residues for the function of the mature protein were conserved, accounting for selective pressures in favor of preserving the specific function of TNFRSF members and the host immune response. To our knowledge, this is the first report of the existence of allelic variations involving some crucial amino acid residues in horse ELR1. Further, it could be an initial step to test the possible functional relevance and relationship of these variants with EIAV infection and disease progression as well as to develop preventive strategies.