Restriction Fragment Length Polymorphism Research Articles

Interactions between DASH-style diet and ADIPOQ and ARL15 genes polymorphisms on blood pressure and central obesity in Iranian adults

This study aimed to investigate the interactions between the Dietary approaches to stop hypertension (DASH) and the rs1501299 and rs6450176 SNPs in terms of cardiometabolic risk factors. This cross-sectional study was conducted on 387 adults (20–70 years old) residing in Yazd, Iran. The participants were selected from participants in the recruitment phase of the Yazd Health Study (YaHS) which is a population-based cohort of 9,962 adults. The genotypes of rs1501299 and rs6450176 SNPs were determined using the restriction fragment length polymorphism (RFLP) method. Unlike for the G allele carriers of rs1501299, high adherence to the DASH dietary pattern was related to the lower diastolic blood pressure (DBP; P-interaction = 0.014) and systolic blood pressure (SBP; P-interaction = 0.051) in the participants with TT genotype. Moreover, in individuals with the AA genotype of rs6450176 SNP, adherence to this diet was associated with lower SBP (P-interaction = 0.012), waist-to-hip ratio (WHR; P-interaction = 0.026) and waist-to-height ratio (WHtR; P-interaction = 0.040), while in G allele carriers, the mean of these items was greater in individuals with high adherence to the DASH diet. The results suggest that following the DASH diet may be related to lower blood pressure and central obesity indices only in individuals with TT genotype of rs1501299 and AA genotype of rs6450176, respectively, whereas it can be associated with higher WHR and WHtR in G allele carriers of rs6450176.

Nationwide carrier screening for congenital adrenal hyperplasia: integrated approach of CYP21A2 pathogenic variant genotyping and comprehensive large gene deletion analysis

BackgroundCongenital Adrenal Hyperplasia (CAH) due to 21-hydroxylase deficiency (21-OHD CAH) is an autosomal recessive disorder resulting from pathogenic variants in the CYP21A2 gene. The disorder exhibits variable clinical severity, with the classical form manifesting as salt-wasting crisis in neonates, while inducing ambiguous genitalia in females and precocious puberty in males through simple virilization. Identifying at-risk couples during the preconception stage holds significance for optimizing reproductive choices. Methods: This study included 204 unrelated preconception individuals undergoing carrier screening. A robust molecular approach was devised for rapid detection of nine prevalent CYP21A2 pathogenic variants, utilizing Amplification-Refractory Mutation System (ARMS) PCR and mass spectrometry (MS) genotyping. Complementary quantitative real-time PCR (qPCR) and PCR-based Restriction Fragment Length Polymorphism (PCR-based RFLP) assays were employed for comprehensive gene deletion analysis. The concordance of pathogenic variant detection between ARMS-PCR and MS, as well as the consistency observed in molecular insights from qPCR and PCR-based RFLP, fortified the accuracy of our methodologies. Results: Our combined method could detect common pathogenic variants and large gene deletions with high concordance between ARMS-PCR, MS genotyping, qPCR, and PCR-based RFLP assays. Remarkably, two carriers exhibited significant large-scale deletions, while another manifested a carrier state due to minor-scale gene conversion. The estimated carrier frequency in our cohort using these methods was approximately 1 in 65 individuals. Conclusions: The methods used for 21-OHD CAH carrier screening offer a reliable, swift, and cost-effective approach for detecting common pathogenic variants and large deletions. Despite some limitations, such as the inability to detect all rare mutations, the techniques provide a practical solution for carrier screening, with an estimated carrier frequency of 1 in 65 in our study population. These findings support the potential adoption of these methods in national carrier screening programs, offering a practical balance between efficiency and affordability.

Neural Network Analysis of Age-Related Changes in the Biosystem of the Intestinal Microbiota of Broiler Chickens

The study aims to evaluate age-related changes and mass accumulation activity in the biosystem of the intestinal microbiota of broiler chickens using fractal frequency-taxonomic profiles and neural network analysis. The molecular genetic terminal restriction fragment length polymorphism method and polymerase chain reaction were used to analyze the frequency-taxonomic profiles of the intestinal microbiota. These profiles were processed using the NONN computational neural network and trained to correctly convert digital profile data into age-related activity index (CSIfract) and mass accumulation activity index (CSImass). The visualization of the age dependence of mass accumulation activity was carried out using a two-dimensional Scale matrix. With the increasing age of broilers, a decrease was observed in the biochemical activity of their intestinal microbiota, associated with the accumulation of functional disorders in the microbial biosystem. Mass accumulative activity reached a maximum at 2-3 weeks of development and then gradually decreased. The quantitative composition of microorganisms changes with age: the number of lactobacilli and bacilli decreased, accompanied by increased clostridium and other microorganisms that increase the risk of diseases in populations. The study showed that age-related changes in broilers' intestinal microbiota significantly affected their biochemical activity and disease risk. To maintain optimal microbial activity, using probiotic supplements in feed is recommended. Keywords: Fractal profile, age-related activity and mass accumulation activity of the biosystem of microorganisms, computational neural network.

PRRSV-2 variant classification: a dynamic nomenclature for enhanced monitoring and surveillance.

Existing genetic classification systems for porcine reproductive and respiratory syndrome virus type 2 (PRRSV-2), such as restriction fragment length polymorphisms and sub-lineages, are unreliable indicators of close genetic relatedness or lack sufficient resolution for epidemiological monitoring routinely conducted by veterinarians. Here, we outline a fine-scale classification system for PRRSV-2 genetic variants in the United States. Based on >25,000 U.S. open reading frame 5 (ORF5) sequences, sub-lineages were divided into genetic variants using a clustering algorithm. Through classifying new sequences every 3 months and systematically identifying new variants across 8 years, we demonstrated that prospective implementation of the variant classification system produced robust, reproducible results across time and can dynamically accommodate new genetic diversity arising from virus evolution. From 2015 to 2023, 118 variants were identified, with ~48 active variants per year, of which 26 were common (detected >50 times). Mean within-variant genetic distance was 2.4% (max: 4.8%). The mean distance to the closest related variant was 4.9%. A routinely updated webtool (https://stemma.shinyapps.io/PRRSLoom-variants/) was developed and is publicly available for end users to assign newly generated sequences to a variant ID. This classification system relies on U.S. sequences from 2015 onward; further efforts are required to extend this system to older or international sequences. Finally, we demonstrate how variant classification can better discriminate between previous and new strains on a farm, determine possible sources of new introductions into a farm/system, and track emerging variants regionally. Adoption of this classification system will enhance PRRSV-2 epidemiological monitoring, research, and communication, and improve industry responses to emerging genetic variants.IMPORTANCEThe development and implementation of a fine-scale classification system for PRRSV-2 genetic variants represent a significant advancement for monitoring PRRSV-2 occurrence in the swine industry. Based on systematically applied criteria for variant identification using national-scale sequence data, this system addresses the shortcomings of existing classification methods by offering higher resolution and adaptability to capture emerging variants. This system provides a stable and reproducible method for classifying PRRSV-2 variants, facilitated by a freely available and regularly updated webtool for use by veterinarians and diagnostic labs. Although currently based on U.S. PRRSV-2 ORF5 sequences, this system can be expanded to include sequences from other countries, paving the way for a standardized global classification system. By enabling accurate and improved discrimination of PRRSV-2 genetic variants, this classification system significantly enhances the ability to monitor, research, and respond to PRRSV-2 outbreaks, ultimately supporting better management and control strategies in the swine industry.

5,10-methylenetetrahydrofolate reductase C677T gene polymorphism as a risk factor for premature coronary artery disease in patients with type 2 diabetes mellitus

BackgroundAfrica, like the rest of the world, is experiencing an increasing prevalence of diabetes mellitus. Diabetes increases the risk for coronary artery disease (CAD) by fourfold compared to people without diabetes. C677T polymorphism in methylenetetrahydrofolate reductase (MTHFR) and hyperhomocysteinemia were reported by many studies as risk factors for CAD among patients with type 2 diabetes mellitus (T2DM). Early detection of modifiable risk factors for CAD is an important aspect of management of diabetes. This is the only study in Sudan which investigates the association between MTHFR genotypes and plasma homocysteine levels, and their role in premature CAD (PCAD) among patients with T2DM.MethodsThis study is a comparative study. We enrolled 226 Sudanese patients with T2DM, age range 25-60 years, recruited from Alshaab and Omdurman teaching hospitals in Khartoum State. 113 patients had CAD confirmed by angiography and electrocardiography (ECG) and 113 had no evidence of CAD. Polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP), using Hinf1 restriction enzyme, were used to determine MTHFR genotypes. Plasma homocysteine levels were determined by enzymatic assay on the Hitachi Cobas Integra® 400 plus. Data was analyzed using statistical package for Social Sciences (SPSS) 23, using Mann-Whtney U test, general linear model, Chi-square test and logistic regression analysis.ResultsThe frequencies of TT, CT, and CC genotypes were 16,40 and 44% among T2DM patients with PCAD. In T2DM patients without PCAD, the frequencies of TT, CT, and CC genotypes were 00,19 and 83%. The T allele showed strong association with PCAD among T2DM patients, p <0.001, odds ratio (OR) 6.2, 95% CI (3.4-11.6). Patients with PCAD showed higher plasma homocysteine levels than patients without PCAD (13.5 µmol/L versus 10 µmol/L, p < 0.001). The T allele had significant effect on homocysteine level, (p <0.001). Plasma homocysteine levels were higher in individuals with TT genotype than those with CT or CC genotypes in patients with PCAD (16.2 + 5.3, 14.3 + 5.7 and 12.9 + 5.02 µmol/L, p=0.017). Homocysteine levels showed a significant association with CAD, p<0.001, OR 3.2, 95% CI (1.9—5.5).ConclusionsOur study suggests that C677T polymorphism of MTHFR gene and hyperhomocysteinemia are risk factors for PCAD in Sudanese population with T2DM.

Implications of cytokine genes polymorphisms in Jordanian patients with obsessive compulsive disorder

BackgroundObsessive–compulsive disorder (OCD) is a common and often highly debilitating chronic neuropsychiatric condition. There is substantial evidence that immune system and genetic changes are involved in OCD pathogenesis. Only a few studies have been encountered in the literature in this field. We aimed at providing experimental evidence for single nucleotide polymorphisms (SNPs) in candidate cytokine genes for the etiology of OCD.MethodsA total of 52 OCD patients and 54 healthy controls were randomly recruited from the Jordanian population. Age ranged between 16 and 55 years (35.5 ± 13.72 and 33.5 ± 10.48 years) for patients and controls, respectively. Five polymorphic positions in four interleukin genes (IL-1β; rs16944 and rs1143634, IL-6; rs1800795, IL-10; rs1800896 and rs1800795) were genotyped using polymerase chain reaction–restriction fragment length polymorphism (PCR–RFLP) method. The tumor necrosis factor-α (TNF-α) SNP (rs1800629) DNA was sequenced by the Sanger method. The obtained data were analyzed using the GraphPad Prism method.ResultsThere were no statistically significant differences between the alleles and genotypes in opposite groups. However, there was a positive association between the incidence of the studied SNP cytokine genes in OCD patients and non-OCD individuals. The alleles of two SNPs (IL-1β + 3954 C > T and TNF-α-308 G > A) were more predominant and more positively correlated with a higher risk in OCD women. In contrast, the link between the gender and the occurrence of the mutant alleles of the other four SNPs IL-1β (− 511 C > T); IL-6 (− 174 G > C); IL-10 (− 1082 A > G); and IL-10 (− 819 C > T was more robust in the males OCD patients than the corresponding females.ConclusionThe observed differences between the alleles in different groups may be due to an association in the tested samples rather than a true association. But, the possibility of a critical effect still exists. One might want to explore this further by repeating the study with a larger sample size.

Development of Breed Traceability for Autochthonous Attappady Black Goats of South India

Attappady Black goats, a meat-type breed raised by the tribal communities of the Attappady hill ranges in Kerala, India, play a pivotal role in the region’s economy, heavily contributing to the livelihoods of its socially disadvantaged population. This study aimed to develop and validate a set of breed-specific genetic markers to enable the precise differentiation of Attappady Black goats from Malabari goats. Breed genetic traceability for Attappady Black goats was developed by adopting deterministic approach using breed specific markers. High throughput genome-wide SNP genotypic data of reference goat population (24 each from Attappady Black and Malabari) obtained by Illumina Goat SNP50 BeadChip genotyping was used to identify candidate breed specific markers. Among the 605 numbers of putative Attappady breed specific markers identified in the reference population, nine putative Attappady breed specific markers were selected and were validated for breed specificity by converting them into Polymerase Chain Reaction -restriction fragment length polymorphism (PCR-RFLP) markers and genotyped in large sample goat population of 447 goats (246 Attappady Black and 201 Malabari). On validation, four markers retained their breed specificity with probability of identification (Pi) ranging from 0.24 to 0.66. Panel of four Attappady Black breed specific markers yielded Pi of 0.92 with a probability of misjudgement (Pm) of 0.04. These breed-specific markers hold significant potential for facilitating breed traceability thus ensuring their conservation and enhancing the socioeconomic status of the tribal communities that rely on them. Future research should focus on the broader application of these markers in routine breed authentication and in marketing niche livestock products.

Investigation of Genetic Polymorphisms Related GSTM1, GSTT1, GSTP1 Genes and their Association with Radiotherapy Toxicity among Head and Neck Cancer Patients.

In this study we explored the association of polymorphisms of glutathione s transferase gene including GSTM1, GSTT1 and GSTP1 with adverse acute normal tissue reactions resulted from radiotherapy in HNC patients. We assessed the association of GSTM1 and GSTT1 null genotypes and Ile105Val of exon-5 and Ala114Val of exon-6 of GSTP1 gene polymorphisms with the risk of acute skin toxicity reactions after therapeutic radiotherapy in HNC patients. Four hundred HNC patients administered with Intensity modulated radiation therapy were enrolled in this study for the evaluation of radiotherapy associated toxicity reactions. The genotyping of GSTM1 and GSTT1 were performed by polymerase chain reaction (PCR). The GSTP1 Ile/Val of exon-5 and Ala/Val of exon-6 polymorphism was determined by PCR followed by restriction fragment length polymorphism (PCR-RFLP). The univariate logistic regression analysis showed that GSTM1 and GSTT1 null genotypes were not associated with either skin reaction or oral mucositis in response to radiotherapy induced after effects. When we studied, A313G polymorphism at exon 5 and C341T polymorphism at exon 6 of GSTP1 gene, majority of genotypes were wild type A/A genotype for exon 5 showed non-significant association with Skin reactions whereas, C/T genotype of exon-6 showed significant negative association with skin reactions. The findings obtained from this study concluded that the null genotypes of GSTM1 and GSTT1 gene polymorphisms showed no association with radiotherapy induced acute toxicities such as dermatitis and oral mucositis. The results indicated negative association of heterozygous C/T genotype of exon-6 of GSTP1 with acute skin reactions.

Investigation of the rs7041 variable of vitamin D-connector protein gene relation with pancreatic involvement in patients with coronavirus disease 2019.

This study aims to examine whether the presence of mutation exists in the vitamin D-connector protein gene rs7041 variant of the pancreatitis table for patients diagnosed with coronavirus disease 2019. A total of 113 patients with normal pancreatic enzyme levels diagnosed with coronavirus disease 2019 and 120 patients with both coronavirus disease 2019 diagnosis and high pancreatic enzyme levels were included in the study. The rs7041 genotyping of the 11th single nucleotide variation in the vitamin D-connector protein gene was determined by polymerase chain reaction and restriction fragment length polymorphism methods. In the patient group with coronavirus disease 2019 diagnosis and high pancreatic enzyme levels, the vitamin D-connector protein gene rs7041 variance GG genotype ratio was determined to be higher than the normal coronavirus disease 2019 patients. As a result of comparisons between the two groups, the difference between the genotype ratios in the relationship was determined to be statistically significant (p=0.004). Coronavirus disease 2019 patients were determined that the rs7041 halves in the vitamin D-connector protein gene could be prone to pancreatitis formation. Different populations and work with more patient groups are needed to verify the results of the study.

Polymorphism of allelic variants of genes in dairy cow’s breed in the conditions of mountain and plain dagestan

In the Republic of Dagestan, where livestock farming is a key area, dairy cattle breeds such as Red Steppe and Caucasian Brown have not been studied in terms of genetic identification. Variations in the PIT1, PRL and GH genes associated with milk productivity can provide important insights into the adaptation of these breeds to various environmental conditions of Dagestan - from plains to mountains and foothills. This can also reveal the unique allelic characteristics of these genes, which is the central focus of our study. The DNA used for the study was extracted from the blood of Caucasian Brown cows bred in mountain (n=74) and foothill (n=52) vertical zonality, as well as the Red Steppe breed bred in the conditions of lowland Dagestan (n=52). The studied cattle populations were genotyped by the polymerase chain reaction method – restriction fragment length polymorphism (PCR-RFLP) on a programmable fourchannel thermal cycler "Tercik" of the company "DNA-technology" (Russia). The polymerase chain reaction (PCR) was carried out using synthesized specific primers. The study of the Caucasian brown cattle breed showed differences in the genetic profiles of individuals living in different ecosystems. In mountainous areas, the allele of the pituitary transcription factor PIT-1A occurs with a frequency of 0.10, while the allele PIT-1B - with a frequency of 0.90. On the contrary, in lowland areas, the proportion of the allele PIT-1A increases to 0.18, and PIT-1B decreases to 0.82. In the mountain zone, the genotypes PIT-1AA, PIT-1BB and PIT-1AB are 2.0%, 82.0% and 16.0%, respectively. In the plain zone, the genotype PIT-1AA is not observed. Cattle raised in certain natural conditions demonstrate unique allelic combinations that are the result of adaptation to the local environment. The data obtained allow us to take these features into account and use them in further breeding work.

Findings in the Molecular and Proteomic Diagnosis of Nocardia brasiliensis

Nowadays, cases of so-called neglected diseases (related to poverty) are increasing. One of them is mycetoma and its variants: Actinomycetoma (caused by bacteria) and eumycetoma (caused by fungi). In the case of actinomycetoma, this disease is often not reported and, as it does not cause pain, people who suffer from it usually go to the doctor when the lesions cause them discomfort when moving or work or social problems. While the diagnosis is usually “easy” since it is mainly based on cultures and stains, many times this is omitted, giving rise to subclinical diagnoses. In this work we strive to make an accurate diagnosis for a patient who suffered from actinomycetoma caused by Nocardia brasilliensis and who was treated with miconazole (an antifungal) for 10 years. After collecting the sample, the corresponding cultures and stains were performed to identify the causal agent, which is Nocardia brasilliensis. Although when talking about the causative agents of actinomycetoma, this aforementioned bacteria is the most studied and reported, we undertook the task of confirming in a more precise way the agent with which we were dealing, for which a procedure was standardized for the identification of the 16S rRNA gene of this bacteria by means of genomic PCR and the realization of the genetic fingerprint mediated by restriction enzymes of Restriction Fragment Length Polymorphism (polymorphisms of the length of restriction fragments) for the 18 strains cultivated in Saburoud Dextrose agar plates. Despite the fact that this gene is present in a huge quantity of bacteria and also that this genus of bacteria has the capacity to mutate certain genes with different organisms that share its environment, such as Aspergillus sp, Fusarium sp, Actinomadura sp, Madrurela sp. This gene does not have homologous regions between the different genera and species that have it, since it has been used for more than 20 years in the identification of this bacteria by PCR with restriction enzymes. In addition, the mutations that could be generated do not occur in the region of this gene since thousands of years have to pass so that they can be expressed and therefore be noticed. This is how, through the extraction, purification, and quantification of the genetic material for its subsequent sequencing with the help of virtual tools such as the BLAST program (Basic Local Alignment Search Tool) that belongs to NCBI, we were able to demonstrate the presence of this 16S rRNA gene in order to have an exact diagnosis of Nocardia brasiliensis.

Association between FTO gene polymorphism and obesity in down syndrome children

Children with Down syndrome (DS) have a higher incidence of overweight and obesity compared to typically developing peers. The fat mass and obesity-associated gene (FTO) is one of the early identified genes linked to obesity in various populations. To date, the FTO rs17817449 gene polymorphism has not been investigated in overweight/obese-DS (ODS) individuals. The current study aimed to explore the potential association between the FTO rs17817449 gene polymorphism and obesity-related markers, and to evaluate the ability of this polymorphism in the prediction of overweight/obesity in DS children and adolescents. This case-control study included 100 DS children under the age of 18, classified into three groups according to BMI-percentile; 50 non-obese DS (NODS), 24 overweight DS, and 26 ODS. Genotyping of FTO gene rs17817449 polymorphism was performed using the restriction fragment length polymorphism (RFLP-PCR) method. Serum lipid and thyroid profiles were also assessed. The results revealed significant increase in the frequency of the FTO rs17817449 T allele among overweight /ODS children compared to NODS children (p=0.0099). Overweight/ODS children exhibited significantly higher frequencies of the FTO rs17817449 GT and TT genotypes compared to NODS children.Conclusion:There is an association between FTO rs17817449 genetic variant and overweight/obesity among the studied DS groups. The FTO rs17817449 GT and TT genotypes, as well as TGs level, were identified as independent risk factors for predicting overweight and obesity in DS children.What is Known:• Overweight and obese-DS (ODS) children displayed higher BMI and atherogenic lipid profile than non-obese DS children (NODS). FTO gene polymorphism rs17817449 contributes to obesity development in general population, but there is conflicting information about the risk allele.What is New:• FTO rs17817449 TT genotype and T allele were considered as independent risk factors for overweight and obesity development in DS children, so they could be used for obesity prediction in DS children.

Mosquito breeding water parameters are important determinants for Microsporidia MB in the aquatic stages of Anopheles species

BackgroundMicrosporidia MB disrupts Plasmodium development in Anopheles mosquitoes, making it a possible biocontrol tool for malaria. As a tool for vector/disease control, its ecological distribution and the factors that determine their occurrence must be defined. We investigated the frequency of Microsporidia MB in Anopheles mosquitoes across selected sites in northern and southern Ghana, as well as the physicochemical parameters of mosquito breeding water that are associated with the occurrence of the fungus, by fitting regression models.MethodsA non-column extraction method was used to extract DNA from the abdomens of 4255 adult Anopheles mosquitoes that emerged from larvae and pupae collected between August and October of 2021 and 2022. Detection of Microsporidia MB was achieved using quantitative PCR (qPCR), while mosquito species were molecularly identified using short interspersed nuclear elements (SINE), restriction fragment length polymorphism (RFLP) methods, and the ANOSPP algorithm.ResultsOverall Microsporidia MB distribution was 2.2% (92/4255). Male mosquitoes exhibited a higher frequency of infections and had a predicted probability of infection that was 85% higher than that of females. Sites in Ghana's Savannah zone had the highest Microsporidia MB distribution (68.5%). Biochemical oxygen demand in mosquito breeding water was estimated to be positively associated with and significantly predicts Microsporidia MB in mosquitoes with an accuracy of 94%. Increasing ammonium ion concentrations reduced the chances of finding Microsporidia MB-positive mosquitoes. According to our data, all Anopheles mosquitoes, including minor species such as An. squamosus, An. pretoriensis and An. rufipes, had equal probability of Microsporidia MB infection.ConclusionsThese results provide preliminary information on micro-ecological factors that potentially support the sustainability of Microsporidia MB infection in mosquitoes during their aquatic life stages. It will be important, therefore, to explore the impact of strategies for larval source management on these factors to ensure that the symbiont's persistence during the host's aquatic stages may not be adversely affected should it be used as an integrated approach for mosquito/disease control.Graphical abstract

Effects of Hinc II Polymorphism in the LDL Receptor Gene on Serum Lipid Levels of Jordanian Individuals with High Risk for Coronary Heart Disease

Coronary heart disease (CHD) has presented high prevalence in the Jordanian population. Nevertheless, studies of genetic risk factors for CHD in our country are insufficiently carried out. We are intended to investigated the effects of Hinc II (exon 12) polymorphisms at the low-density lipoprotein receptor (LDLR) gene on circulating lipids of 150 individuals with high risk for CHD (HRG) and 150 controls (C). Genomic DNA was extracted from white blood cells and amplified by polymerase chain reaction (PCR) and digested with HincII Restriction enzyme. LDL, TC (total cholesterol), TG (triglycerides), and HDL (high density lipoproteins) levels were measured in all subjects. RFLP (restriction fragment length polymorphism) analysis was conducted to identify genotype of LDLR gene in (HRG) and 150 controls (C). The results showed a significant correlation existed between this RFLP locus and (HRG), X2=10.6, P<0.05; H<sup>+</sup> allele frequency: X2=7.88, P<0.05., H<sup>-</sup> allele frequency: X2=7.88, P<0.05. Genotypes H<sup>+</sup>H<sup>+</sup> and H<sup>+</sup>H<sup>- </sup>in high risk group are significantly associated with high levels of LDL, TC, TG, P<0.05 and with low level of HDL P<0.05., while H<sup>-</sup>H<sup>-</sup> is associated with normal values of serum lipids P<0.05. It is inferred that H<sup>+</sup> allele might be associated with high blood cholesterol level, and the H<sup>-</sup> allele with normal level. This study suggests that the differences in LDLRG genotypes might affect the phenotype of lipid metabolism.

Association of rs3755319 and rs4148325 of the <i>UGT1A1</i> gene, rs2328136 of the <i>NUP153-AS</i> gene, and rs16928809 of the <i>SLC22A18</i> gene with benign unconjugated hyperbilirubinemia

Background: Benign unconjugated hyperbilirubinemia, also known as Gilbert's syndrome, is a common in the population moderate increase in total and unconjugated bilirubin concentrations in individuals without liver disease or hemolysis. Aim: To identify associations of rs3755319, rs4148325 of the UGT1A1 gene, rs2328136 of the NUP153-AS gene, and rs16928809 of the SLC22A18 gene with benign unconjugated hyperbilirubinemia. Methods: This case-control study included a group of individuals with benign unconjugated hyperbilirubinemia (n = 414, mean age 36.7 ± 15.9 years, 49.8% men) and a control group (n = 381, mean age 39.1 ± 15.9 years, 52.5% men). The sample was randomly selected from the participants of the MONICA project, screening of young people aged 25–44 years and a cross-sectional study of schoolchildren in Novosibirsk. DNA was isolated from venous blood by phenol-chloroform extraction or an express assay (PROBA-RAPID-GENETICS, DNA-Technology, Russia). Genotyping of the groups by rs3755319, rs4148325 of the UGT1A1 gene, rs2328136 of the NUP153-AS gene, and rs16928809 of the SLC22A18 gene was performed by polymerase chain reaction followed by restriction fragment length polymorphism analysis. Results: No significant differences were found between the individuals with benign unconjugated hyperbilirubinemia and the control group by the genotypes and alleles of the nucleotide sequence variants of rs16928809 of the SLC22A18 gene (p 0.05). The CC genotype and the C allele of rs3755319 of the UGT1A1 gene were more common in the individuals with benign unconjugated hyperbilirubinemia, than in the control group (CC vs AC + AA: odds ratio (OR) = 21.1, 95% confidence interval (CI) 14.7–30.4, p 0.001; C vs A: OR = 12.4, 95% CI 9.4–16.4, p 0.001). The concentrations of total and unconjugated bilirubin were higher in the carriers of the CC genotype of rs3755319, compared to the carriers of the other two genotypes (p 0.05). rs4148325 of the UGT1A1 gene was in the linkage disequilibrium with the rs3064744 UGT1A1 variant. The GG genotype and the G allele rs2328136 of the NUP153-AS gene were more common in the individuals with benign unconjugated hyperbilirubinemia than in the control group (GG vs AG + AA: OR = 1.361, 95% CI 1.002–1.848, p = 0.048; G vs A: OR = 1.33, 95% CI 1.02–1.73, p = 0.034). Conclusion: The CC genotype and the C allele of rs3755319 of the UGT1A1 gene, the GG genotype and the G allele of rs2328136 of the NUP153-AS gene are the genotypes and alleles of risk for benign unconjugated hyperbilirubinemia. The rs16928809 of the SLC22A18 gene is not associated with benign unconjugated hyperbilirubinemia.

Incidence of PAX6 single nucleotide polymorphisms in congenital iridofundal coloboma and other congenital ocular abnormalities: a tertiary care hospital experience from central India

Purpose of this study is to identify selected single nucleotide polymorphisms (SNP) of the PAX6 gene and to assess their correlation with congenital iridofundal colobomas and other congenital ocular anomalies.Material and methods. It was a case-control study done on 45 patients aged from 75 days to 58 years (mean age 29.36 ± 14.4 years) with irido-fundal coloboma and 45 healthy controls aged 35.23 ± 13.92 years. Ocular examination was done by using slit-lamp microscopy inspection, fundoscopy and intraocular pressure measurement. Genotyping was done by using the polymerase chain reaction — restriction fragment length polymorphism (PCR-RFLP) method.Results. Two irido — fundal coloboma patients showed CT (+/-) heterozygous genotype of rs667773 SNP and the rest were wild-type CC (-/-) homozygous genotype. All controls showed CC-/- wild-type homozygous genotype. PAX6 SNP rs3026354 showed CC (-/-) wild-type homozygous genotype condition in all patients. Neither CG (+/-) heterozygous nor homozygous GG (+/+) genotype was reported in patients and controls. SNP rs662702, genotype pattern was CC-/- wild type homozygous in all patients and controls. CC genotype frequency was 95.56 and CT genotype was 4.4% while C allele frequency was 97.78 and T allele frequency was 2.22 % in rs667773 C>T SNP. rs3026354C>G SNP had 100 % CC genotype and C allele frequency in both case and control populations. SNP rs 662702C>T showed 100 % CC genotype and C allele frequency in the case and control respectively.Conclusion. The elevated frequency of the CC genotype with C allele was more common in irido fundal patients. Two heterozygous CT genotype of rs667773C>T SNP were reported in two irido-fundal coloboma patients.

Rare STAT3 haplotypes cause a predisposition to developing congenital anomalies of the kidney and urinary tract disorder.

Congenital anomalies of the kidney and urinary tract (CAKUT) are characterized by renal developmental disorders in the embryonic period. STAT3 is a member of the STAT protein family. The members of this protein family play roles in various cellular mechanisms, such as the early stages of embryonic development, kidney development, and renal diseases. This study aims to determine the frequency of STAT3 rs1053004, rs744166, rs3816769, and rs4796793 polymorphisms in individuals with CAKUT. Two of four polymorphisms, rs744166 (c.-1-13666T>C, NM_001369512.1) and rs4796793 ( c.-1915C>G, NM_001369512.1), were analyzed by a polymerase chain reaction (PCR) and the restriction fragment length polymorphism method. Two other polymorphisms, rs1053004 (c.*1671C>T, NM_001369512.1) and rs3816769 (c.273+314A>G, NM_001369512.1), were analyzed using real-time PCR-melting curve analysis. Our research indicates that individuals with the TT allele for rs1053004 single nucleotide polymorphism have a 1.23 times greater disease risk than those with the CC allele. Those with the CC allele for rs3816769 have a 1.41 times greater risk of disease than those with the TT allele. These findings suggest a potential genetic predisposition to CAKUT. Furthermore, the research identified significant connections between rare haplotypes and CAKUT (p = 0.041). The CCTC haplotype for rs744166, rs4796793, rs1053004, and rs3816769 polymorphism was exclusively present in the CAKUT group, while the CGTT haplotype for the same polymorphisms was only detected in the control group. The presence of rare haplotypes for the rs1053004, rs3816769, rs4796793, and rs744166 polymorphisms may significantly affect the onset or prevention of CAKUT. These findings could potentially have important clinical implications, providing a deeper understanding of the genetic basis of CAKUT and potentially influencing future diagnostic and treatment strategies.

Marker Assisted Selection: A Novel Approach for Crop Improvement

The process of marker-assisted selection, or marker-aided selection (MAS), selects a trait of interest indirectly by considering a marker linked to the trait (e.g., quality, productivity, disease resistance, and biotic stress tolerance) rather than the trait itself. This integration of marker data with traditional selection approaches has become a widely studied and recommended method for advancing breeding programs. This technique has been extensively researched and recommended for animal and plant breeding. Here, we combine marker data with conventional selection to choose the best candidate for a future breeding program. Nowadays, the majority of MAS investigations employ DNA-based markers such as microsatellites, single nucleotide polymorphisms (SNPs), random amplification of polymorphic DNA (RAPD), amplified fragment length polymorphism (AFLP), and restriction fragment length polymorphism (RFLP). In this case, we discussed several kinds, techniques, and other features of marker Assisted Selection.

MTHFR Gene Polymorphisms and DNA Methylation in Idiopathic Spontaneous Preterm Birth

Background and Objectives: Preterm birth (PTB) is a complex condition with various contributing factors, including genetic and epigenetic influences such as DNA methylation. Methylenetetrahydrofolate reductase (MTHFR) plays a critical role in DNA methylation and the remethylation of homocysteine. This study aimed to investigate the association between maternal MTHFR C677T and A1298C polymorphisms, LINE-1 DNA methylation levels, and the risk of idiopathic spontaneous preterm birth (SPTB) in Caucasian women from Croatia and Slovenia. Materials and Methods: A total of 50 women with SPTB (<34 weeks of gestation) and 50 control women were included in the study. MTHFR polymorphisms were analyzed using polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP), and LINE-1 DNA methylation levels were quantified using the MethyLight method. Results: The study found no significant differences in MTHFR C677T and A1298C polymorphisms’ genotype or allele frequencies between women with SPTB and controls. Additionally, no statistical significance of LINE-1 DNA methylation was found between the genotypes of the MTHFR polymorphisms analyzed. Conclusions: The study suggests no conclusive association between MTHFR C677T and A1298C polymorphisms, LINE-1 DNA methylation, and SPTB in Croatian and Slovenian women. Considering prior evidence connecting MTHFR polymorphisms, hyperhomocysteinemia, and PTB, the lack of homocysteine measurements and unassessed impact of folate or vitamin B supplementation limit the conclusions.

Multiethnic prevalence of the APOL1 G1 and G2 variants among the israeli dialysis population

Abstract Background and hypothesis The two apolipoprotein L1 (APOL1) variants, G1 and G2, are common in populations of sub-Saharan African ancestry. Individuals with two of these alleles (G1 or G2) have an increased risk for a spectrum of non-diabetic chronic kidney diseases. However, these variants are typically not observed outside of populations that self-identify as current continental Africans or having clear recent African ancestry such as, most notably, African Americans, and other large population groups in the Americas and several European countries. We hypothesized that the diverse ethnic groups within the Israeli population may exhibit varying levels of recent African ancestry. Therefore, it is plausible that APOL1 risk alleles might be present even in individuals who do not self-identify as having sub-Saharan African descent. Methods We non-selectively screened persons with kidney failure across Israel for APOL1 risk variants using restriction fragment length polymorphism. Results We recruited 1 744 individuals from 38 dialysis units in Israel. We identified eight patients of Moroccan Jewish, Bedouin, or Muslim Arab ancestry, who carry at least one G1 or G2 allele. None of the eight patients carried the protective APOL1 p.N264K variant. Furthermore, despite all Bedouin individuals being G2 heterozygous, the G2 minor allele frequency was significantly enriched in kidney failure cases compared to ethnically matched controls (P value = 0.006). Conclusions These findings show that APOL1 G1 and G2 allelic variants are present in populations previously not appreciated to possess recent sub-Saharan ancestry and suggest that a single G2 risk variant may confer increased risk for chronic kidney disease in certain population contexts.