Amplification Refractory Mutation System Research Articles

Association of histidine-rich glycoprotein C633T single nucleotide polymorphism and recurrent miscarriage in Iranian women.

Recurrent miscarriage (RM) is defined as the occurrence of at least two or three subsequent miscarriages within the 20th -24th weeks of pregnancy. The primary objective of this study was to investigate whether histidine-rich glycoprotein C633T single nucleotide polymorphism (HRG C633T SNP) statistically correlates with the occurrence of RM among Iranian women. Blood samples from 200 women were taken at the outset of the study. Then, the blood samples of 100 women who had a record of RM (case group) were compared with the other 100 women's blood samples who had no record of RM (control group). Following DNA extraction, the polymorphism of histidine-rich glycoprotein C633T (HRG C633T) for every case was specified and all women were genotyped by the amplification-refractory mutation system (ARMS) method. The results of the study revealed that there was a statistically significant difference between T/T genotype (OR = 3.5, CI (1.39-8.77), p = 0.007), and C/T genotype (OR = 1.83, CI (0.99-3.37), p = 0.05) in the case and control groups. Also, a statistically significant association was observed in T allelic frequency in the RM participants compared to the control group (OR = 2.01, CI (1.31-3.09), p = 0.01). The present study determined that there was a statistically significant relationship between HRG C633T SNP and increased RM regarding allelic and genotypical aspects. Moreover, it became apparent that women with homozygous T/T genotype were more susceptible to the risk of RM.

Potential Association of The Pathogenic Kruppel-like Factor 14 (KLF14) and Adiponectin (ADIPOQ) SNVs with Susceptibility to T2DM.

To evaluate the associations of the pathogenic variants in Kruppel-like Factor 14 (KLF 14) and Adiponectin (ADIPOQ) with susceptibility to type 2 diabetes mellitus (T2DM). Type 2 diabetes mellitus (T2DM) is a pandemic metabolic disease characterized by increased blood sugar and caused by resistance to insulin in peripheral tissues and damage to pancreatic beta cells. Kruppel-like Factor 14 (KLF-14) is proposed to be a regulator of metabolic diseases, such as diabetes mellitus (DM) and obesity. Adiponectin (ADIPOQ) is an adipocytokine produced by the adipocytes and other tissues and was reported to be involved in T2DM. To study the possible association of the KLF-14 rs972283 and ADIPOQ-rs266729 with the risk of T2DM in the Saudi population. We have evaluated the association of KLF-14 rs972283 C>T and ADIPOQ-rs266729 C>G SNV with the risk to T2D in the Saudi population using the Amplification Refractory Mutation System PCR (ARMS-PCR), and blood biochemistry analysis. For the KLF-14 rs972283 C>T SNV we included 115 cases and 116 healthy controls, and ADIPOQ-rs266729 C>G SNV, 103 cases and 104 healthy controls were included. Results indicated that the KLF-14 rs972283 GA genotype and A allele were associated with T2D risk with OR=2.14, p-value= 0.014 and OR=1.99, p-value=0.0003, respectively. Results also ADIPOQ-rs266729 CG genotype and C allele were associated with an elevated T2D risk with an OR=2.53, p=0.003 and OR=1.66, p-value =0.012, respectively. We conclude that SNVs in KLF-14 and ADIPOQ are potential loci for T2D risk. Future large-scale studies to verify these findings are recommended. These results need further verifications in protein functional and large-scale case control studies before being introduced for genetic testing.

Association of polymorphisms within P2RX4 with type 2 diabetes mellitus: a preliminary case-control study

Objective Type 2 diabetes mellitus (T2DM) is a complex heterogenic metabolic with a wide range of etiology. Purinergic receptors have pivotal roles in different processes and are hypothesized to have roles in the pathogenesis of T2DM. Materials and Methods Three hundred subjects affected by T2DM and 300 healthy subjects were genotyped by amplification refractory mutation system polymerase chain reaction (ARMS-PCR). SPSS V16.0 was recruited for statistical analysis. Results The findings showed that the G allele of rs25644A > G increases the risk of T2DM in our population statistically (OR = 1.51, 95% CI = 1.14–1.99, p = 0.003). This allele in some genotype models, including the dominant model, caused an increase in the risk of T2DM. The interaction of genotypes between studied variants in the P2XR4 gene increased the risk of T2DM. Haplotype analysis showed that Ars1169727/Grs25644 haplotype caused an increase in the risk of T2DM. Conclusions The findings suggest that rs25644A > G plays a role in our population’s increased risk of T2DM.

POLYMORPHISM OF THE BETA-CASEIN GENE IN HOLSTEIN-ZEBU COWS IN THE NORTH OF MINAS GERAIS, BRAZIL

This study aimed to investigate A1 and A2 polymorphisms of the beta-casein gene in crossbred Holstein-Zebu (HZ) cows from four herds in Northern Minas Gerais, Brazil. Blood samples were collected from 112 randomly selected cows, of wich 66 were at ½ HZ, and 46 were at ¾ HZ. DNA extraction was conducted using the phenol-chloroform method, and polymorphisms were evaluated using the amplification refractory mutation system - polymerase chain reaction technique. The results demonstrated that 72/112 (64.3%) cows were heterozygous (A1A2), 33/112 (29.5%) were homozygous for A2A2, and 7/112 (6.2%) were A1A1 homozygous. The percentage of A2A2 cows ranged from 10.7 to 41.4% among the evaluated herds. The frequencies of the A1 and A2 alleles were 38.4 and 61.6%, respectively. The A2 allele was the most frequent in both breeds. The ½ HZ cows were 1.8 times more likely (Odds Ratio) to belong to the A1A1 genotype. The distribution of genotypes was in Hardy-Weinberg equilibrium in three of the herds; however, the total of samples and the stratified samples according to breeds were not in equilibrium considering α = 0.05.

Effect of strong programmed cell death ligand-1 (PD-L1) expressions on clinical outcomes in advanced non-small cell lung cancer treated with third-generation epidermal growth factor receptor-tyrosine kinase inhibitors.

e20500 Background: Third-generation tyrosine kinase inhibitors (TKIs) are the standard treatment for advanced epidermal growth factor receptor (EGFR) mutation-positive lung adenocarcinoma. In first/second generation EGFR-TKIs, strong programmed death ligand 1 (PD-L1) expression contributes to primary resistance, significantly affecting patient prognosis. Despite this, the relationship between PD-L1 expression levels and third-generation TKIs remains unclear. Methods: This retrospective cohort study reviewed patients with advanced NSCLC who received third-generation EGFR-TKIs as first-line systemic therapy at the Shandong Cancer Hospital between March 2019 and June 2022. The EGFR status of the patients was assessed using amplification refractory mutation system fluorescence quantitative polymerase chain reaction, and the PD-L1 expression level was evaluated using Dako 22 C3 immunohistochemical staining. The Kaplan–Meier method was used for survival analysis. The log-rank test was used to compare differences in survival curves between the two groups. Results: Overall, 150 patients were included in this study. PD-L1 expression was negative (PD-L1 tumor proportion score < 1%) in 89 cases, weak (1–49%) in 42 cases, and strong (≥50%) in 19 cases. The median follow-up period for the entire cohort was 22.12 months (median progression-free survival [mPFS]: 24.33 months); the median overall survival was not reached. mPFS for patients with negative, weak, and strong PD-L1 expressions was 23.60, 26.12, and 16.60 months, respectively. The mPFS for strong PD-L1 expression was significantly shorter than that for with weak PD-L1 expression but was not associated with negativity, particularly in the 19DEL and 21L858R subgroups. PFS was significantly shorter in patients with strong PD-L1 expression in both subgroups (19DEL and 21L858R) than in those with weak PD-L1 expression. Conclusions: Strong PD-L1 expression in tumor cells influenced the clinical outcomes of patients with advanced NSCLC treated with third-generation EGFR-TKIs. Stronger PD-L1 expression in TKI-treated patients with advanced first-line EGFR-mutated NSCLC was associated with worse PFS.

Application of the TaqMan ARMS-PCR Approach for Genotyping Drug-Induced Hearing Loss Using Dried Blood Samples.

A single nucleotide variant in mitochondrial DNA (mtDNA) 1555A>G is associated with drug-induced hearing loss. For the 1555A>G mutation site, 1555A wild-type and 1555G mutant-type plasmids were constructed, respectively. In this study, a PCR method based on the TaqMan amplification refractory mutation system was proposed to detect mtDNA 1555A>G. A common upstream primer, a common TaqMan probe, and two downstream allele-specific primers with mismatched bases were designed. One-step amplification and detection of the wild-type and mutant type at the 1555 site were realized for the deafness-related gene through two reactions. Based on this detection method, the minimum detection limit of the wild-type and mutant type detection systems for plasmids was 50 copies/μL. The minimum sensitivity for the detection of nucleic acids in real dried blood spot (DBS) samples was 0.1 ng/μL. In the normal DBS DNA sample, the detection limit of the mutation abundance reached 0.78%. The specificity of the detection method was 100%, and the coefficient of variation was less than 3.36%. This approach was validated using clinical DNA extracted from 113 DBS samples of newborns. Additionally, it showed 100% agreement with bi-directional Sanger sequencing. It can be used as an optional method for the clinical detection of deafness-related genes.

Effect of DNMT3A R882H Hot Spot Mutations on DDX43 Promoter Methylation in Acute Myeloid Leukemia.

Epigenetic alterations have been observed in many hematological malignancies, including acute myeloid leukemia (AML). Many of these alterations result from mutations in DNA methyl transferase (DNMT) enzymes, disabling them to methylate target genes in a proper way. In this case-control study, we investigated the association between R882H mutation in DNMT3A gene and DDX43 gene methylation in patients with AML. 47 AML patients and 6 controls were included in this study. After DNA extraction, amplification refractory mutation system (ARMS)-PCR was used to evaluate R882H mutations in DNMT3A gene. The high-resolution melting (HRM) method was used to determine the methylation changes of the DDX43 gene promoter. R882H mutation was only found in 10.6% (5 out of 47) of AML patients. The frequency of DDX43 gene methylation was significantly higher in patients without R882H mutations compared to patients with R882H mutations (P < 0.05). The DNMT3A R882H mutation is typically present in a minority of AML patients. Nevertheless, this mutation is associated with a reduced frequency of methylation in the DDX43 promoter region.

Multiplex Asymmetric PCR by Combining the Amplification Refractory Mutation System with the Homo-Tag-Assisted Nondimer System.

Asymmetric PCR is widely used to produce single-stranded amplicons (ss-amplicons) for various downstream applications. However, conventional asymmetric PCR schemes are susceptible to events that affect primer availability, which can be exacerbated by multiplex amplification. In this study, a new multiplex asymmetric PCR approach that combines the amplification refractory mutation system (ARMS) with the homo-Tag-assisted nondimer system (HANDS) is described. ARMS-HANDS (A-H) PCR utilizes equimolar-tailed forward and reverse primers and an excess Tag primer. The tailed primer pairs initiate exponential symmetric amplification, whereas the Tag primer drives linear asymmetric amplification along fully matched strands but not one-nucleotide mismatched strands, thereby generating excess ss-amplicons. The production of ss-amplicons is validated using agarose gel electrophoresis, sequencing, and melting curve analysis. Primer dimer alleviation is confirmed by both the reduced Loss function value and a 20-fold higher sensitivity in an 11-plex A-H PCR assay than in an 11-plex conventional asymmetric PCR assay. Moreover, A-H PCR demonstrates unbiased amplification by its allele quantitative ability in correct identification of all 31 trisomy 21 samples among 342 clinical samples. A-H PCR is a new generation of multiplex asymmetric amplification approach with various applications, especially when sensitive and quantitative detection is required.

Primary mucinous tumors of the renal pelvis: Clinical, histopathological, and molecular analysis of three cases

Primary mucinous tumors of the renal pelvis are extremely rare and pose challenges in terms of diagnosis and treatment. This study reviewed the clinical and pathological characteristics of mucinous tumors of the renal pelvis, including mucinous cystadenocarcinomas and mucinous cystadenomas. Immunohistochemical analysis was conducted in three cases, along with KRAS gene detection using the Amplification Refractory Mutation System (ARMS) method. The results revealed mucinous epithelium with acellular mucinous pools in all cases, and acellular mucinous pools were observed in the renal parenchyma and perirenal fat capsules. All tumors expressed CK20 and CDX2, and one case showed KRAS gene mutation. The study suggests that mucinous cystadenomas of the renal pelvis may exhibit borderline biological behaviors. This study is the first to report a KRAS gene mutation in a mucinous cystadenoma of the renal pelvis, offering valuable insights into the diagnosis and treatment of this rare condition.

Detection of Asymptomatic Sickle Cell Hemoglobin Carriers and Fetal Hemoglobin Regulating Genetic Variants in African Descendants from Oaxaca, Mexico.

Sickle cell anemia has been classified as a noninfectious neglected tropical disease and, although not exclusively, affects African descendants more frequently. This study aimed to detect asymptomatic sickle cell hemoglobin carriers (HbAS) in marginalized and vulnerable populations during a public health screening in African descendants from Oaxaca, Mexico, and to validate an amplification refractory mutation system (ARMS)-PCR methodology to detect fetal-hemoglobin (HbF)-regulating genetic variants in BCL11A toward affordable routine association of single nucleotide variants (SNVs) with HbF concentrations. To this aim, hemoglobin variants were detected by acidic citrate agar and alkaline cellulose acetate electrophoreses. SNVs in the hemoglobin subunit beta gene (HBB) were identified by the β-globin mutation detection assay (β-GMDA) and ARMS-PCR, respectively, and validated by Sanger sequencing. The association between genotypes and HbF concentrations was evaluated using Spearman's correlation coefficient. The results obtained during a directed screening in 140 self-identified African descendants revealed 42 HbS-carriers (30%), of which 39 showed normal total hemoglobin concentrations (92.8%), only 3 presented anemia (7.2%), and 9 showed quantifiable HbF concentration (21.4%). As validated by Sanger sequencing, the designed ARMS-PCR efficiently detected homozygous and heterozygous variants in BCL11A. In a cohort of 42 heterozygous (HbAS) and 27 healthy (HbAA) individuals from the same population, only one SNV (rs766432) showed statistically significant association with increasing HbF concentration, and two new unrelated homozygous silent variants were identified. This study reveals the need to raise coverage of HbS screening in vulnerable populations and shows a feasible low-cost ARMS-PCR methodology to determine the presence of SNVs in quantitative trait loci affecting HbF.

Multiplex fluorescent amplification-refractory mutation system PCR method for the detection of 10 genetic defects in Holstein cattle and its comparison with the KASP genotyping assay.

The common deleterious genetic defects in Holstein cattle include haplotypes 1-6 (HH1-HH6), haplotypes for cholesterol deficiency (HCD), bovine leukocyte adhesion deficiency (BLAD), complex vertebral malformation (CVM) and brachyspina syndrome (BS). Recessive inheritance patterns of these genetic defects permit the carriers to function normally, but homozygous recessive genotypes cause embryo loss or neonatal death. Therefore, rapid detection of the carriers is essential to manage these genetic defects. This study was conducted to develop a single-tube multiplex fluorescent amplification-refractory mutation system (mf-ARMS) PCR method for efficient genotyping of these 10 genetic defects and to compare its efficiency with the kompetitive allele specific PCR (KASP) genotyping assay. The mf-ARMS PCR method introduced 10 sets of tri-primers optimized with additional mismatches in the 3' end of wild and mutant-specific primers, size differentiation between wild and mutant-specific primers, fluorescent labeling of universal primers, adjustment of annealing temperatures and optimization of primer concentrations. The genotyping of 484 Holstein cows resulted in 16.12% carriers with at least one genetic defect, while no homozygous recessive genotype was detected. This study found carrier frequencies ranging from 0.0% (HH6) to 3.72% (HH3) for individual defects. The mf-ARMS PCR method demonstrated improved detection, time and cost efficiency compared with the KASP method for these defects. Therefore, the application of mf-ARMS PCR for genotyping Holstein cattle is anticipated to decrease the frequency of lethal alleles and limit the transmission of these genetic defects.

Determination of IL-23 receptor expression and gene polymorphism (rs1884444) in Iranian patients with ankylosing spondylitis

BackgroundThrough investigating genetic variations, it has been demonstrated that single nucleotide polymorphisms (SNPs) in the IL-23 receptor (IL23R) gene have a critical role in the pathophysiology of ankylosing spondylitis (AS). Here, we investigated whether the IL23R variant (rs1884444) is associated with AS in the Iranian population.Methods and materialIn this research, we analyzed rs1884444 in a group of 425 patients with AS and 400 matched controls. For DNA extraction, the phenol/chloroform technique was utilized. Peripheral blood mononuclear cells (PBMCs) were obtained from the whole blood of 39 patients and 43 healthy controls and total RNA was extracted. Genotyping was performed by amplification-refractory mutation system (ARMS)–PCR method. Afterward, the expression level of IL23R was analyzed by the real-time quantitative (Q)-PCR method.ResultsWe observed no significant association between the distribution of alleles and genotypes of rs1884444 and susceptibility to AS. In addition, the expression level of IL23R did not differ between PBMCs from AS patients compared to the control group (P = 0.167). Furthermore, the relative expression level of IL23R was positively correlated with the BASDAI (P < 0.01) and BASFI (P < 0.05) scores of the patients.ConclusionIt appears that IL23R polymorphism (rs1884444) and the level of gene expression might not contribute to the susceptibility to AS in the Iranian population. The correlation of IL23R expression with the level of BASDAI and BASFI scores in patients may be due to the role of the IL-23/IL-23R signaling cascade in inflammation and exert a critical role in the development of AS.

A novel method of multiplex SNP genotyping assay through variable fragment length allele-specific polymerase chain reaction: Multiplex VFLASP-ARMS

Variable Fragment Length Allele-Specific Polymerase Chain Reaction (VFLASP) and Amplification Refractory Mutation System (ARMS) are reliable methods for detecting allelic variations resulting from single base changes within the genome. Due to their widespread application, allele variations caused by Single Nucleotide Polymorphisms (SNPs) can be readily detected using allele-specific primers. In the context of the current study, VFLASP was combined with ARMS method as a novel strategy to enhance the efficacy of both techniques. Clinically important base variations within SNP regions used in the study were detected by a fragment analysis method. To validate the accuracy of the developed VFLASP-ARMS method, specifically designed synthetic sequences were tested using a capillary electrophoresis system. Allele-specific primers exhibit differences solely at the 3′ end based on the sequence of the SNP. Additionally, to increase the specificity of the primers, a base was intentionally added for incompatibility. Therefore, allele discrimination on fragment analysis has been made possible through the 3–6 bp differences in the amplicons.With the optimization of the system, designed synthetic sequences provided reliable and reproducible results in wild-type, heterozygous, and homozygous genotypes using the VFLASP-ARMS method. Hence, our results demonstrated that VFLASP-ARMS method, offers a novel design methodology that can be included in the content of SNP genotyping assays.

Association of a genetic variant in angiopoietin-like 3 with serum HDL-C and risk of cardiovascular disease: A study of the MASHAD cohort over 6 years.

Loss-of-function (LOF) variants of the angiopoietin-like 3 (ANGPTL3) gene are reported to be associated with serum triglyceride (TG) and high-density lipoprotein cholesterol (HDL-C) concentrations and thereby affect the risk of cardiovascular disease (CVD). In the present study, we examined the association of rs10789117 in the ANGPTL 3 gene locus and the risk of CVD in the group of people who were part of the Mashhad-Stroke and Heart-Atherosclerotic-Disorders (MASHAD) cohort. One thousand and two healthy individuals enrolled in this study of whom 849 subjects were healthy and 153 subjects developed CVD outcomes after 6 years of follow-up. After a 12-h overnight fasting, 20 mL of blood samples were collected for the measurement of fasting blood glucose and lipid profile. DNA was extracted, and the Tetra-ARMS PCR (amplification refractory mutation system) was used for genotyping of rs10789117 in the ANGPTL3 gene. The genotype frequencies of the variant of rs10789117 in the ANGPTL3 gene were estimated using χ2 tests. Eventually, the statistical analysis was done by SPSS version 20. Individuals with AC/CC genotypes (rs10789117) were found to have to greater risk of CVD events compared to AA genotype (OR = 1.43, 95%CI = 1.01-2.02, p = 0.041). There was a 1.3-fold increase in cardiovascular events in individuals carrying the C allele of rs10789117 variant compared to non-carriers (OR = 1.32, 95%CI = 1.06-1.72, p value = 0.038). There were significant differences between different genotypes for serum triglyceride levels within the control group, but this difference was not significant in the group with CVD. Moreover, there was a significant association between CC genotype and CVD risk in the individuals with a normal serum HDL-C. We have found that a rs10789117 C>A in ANGPTL3 gene polymorphism was associated with incident CVD events, and this may be of value as a risk stratification biomarker in CVD in the Iranian population.

Cytokine Genetic Polymorphism of Interleukin-4 and Risk of Asthma in Some Iraqi Patients

Abstract Background: Asthma, a chronic inflammatory respiratory disorder, is influenced by genetic and environmental factors. Allergic asthma is becoming more common because of higher levels of air pollution. Interleukin-4 (IL-4) plays an important role in allergic inflammation and causes the expression of vascular cell adhesion molecule-1. Objective: The present study aims to shed light on the association between cytokine genetic polymorphisms and asthma in Iraqi patients, and to determine their impact on the risk of disease, under the scope of the following: (1) IL-4 cytokines, in terms of their serum level. (2) Assessment of the polymorphisms in the promoter regions of the IL-4, genes in asthma disease patients was carried out, and then their impact on the profile of investigated cytokines was evaluated. Such a collective evaluation may aid in a better understanding of etiopathogenesis in the asthma disease. Materials and Methods: A case-control study was conducted that included 100 participants divided into a patient group (n = 50) with bronchial asthma and a healthy group as a control (n = 50) without asthma. Blood specimens were collected from participants at Marjan Hospital, Babylon Governorate. Serum levels of IL-4 were estimated by an enzyme-linked immunosorbent assay (ELISA) kit. IL4-590 gene polymorphism was detected using amplification refractory mutation system polymerase chain reaction (PCR) with specific primer sequences. Results: The mean concentrations of serum IL-4 were significantly higher in patients having asthma compared to the healthy group (149.84 vs. 53.50, P &lt; 0.0001). In terms of allele and genotype frequencies, the TT genotype was shown to be less common in asthma group compared to controls (4% vs. 22%), whereas the CT genotype was more common in asthma patients (34% vs. 16%). The CC genotype was similar between the two groups. The T allele was more frequent in healthy (39%) compared to asthmatic group (12%), whereas the C allele frequency was higher in asthma patients (61%) compared to controls (88%). Conclusion: This study suggests that gene polymorphism of IL4-590 is associated with risk of developing asthma. Asthma patients have elevated levels of IL-4 and a lower frequency of the TT genotype, indicating an increased likelihood of developing asthma when carrying the T allele and TT genotype. Conversely, the C allele may have a protective influence against asthma development.

Hematological Profile of Hb Adana Among High School Students in Northeast Peninsular Malaysia.

Background Hb Adana is a non-deletional alpha (α)-thalassaemia variant resulting from mutations in α1- or α2-globin codon 59 (αCD59), leading to the production of unstable α-globin. Clinical manifestations can vary from silent carrier status to dependence on blood transfusions, hepatosplenomegaly, skeletal deformities, and spinal cord compression. Despite the significance of Hb Adana inheritance, studying this variant poses challenges due to the scarcity of molecular tests and the potential for routine diagnoses to be overlooked. This study aims to investigate the prevalence of Hb Adana among local high school students and assess the hematological parameters and hemoglobin analysis of Hb Adana in Malaysia. Methodology This retrospective study analyzed 13,721 blood samples collected from high school students participating in Malaysia's National Thalassaemia Screening Program at Hospital Raja Perempuan Zainab II (HRPZ II). Deletional α-thalassaemia was detected using multiplex gap-polymerase chain reaction (PCR), while common non-deletional α-thalassaemia was identified using multiplex amplification refractory mutation system (ARMS) PCR. Data were extracted from the HRPZ II database for analysis. Results Among the participants, 2327 individuals were found to have either common deletional (n=1037, 44.6%) or non-deletional (n=1290, 55.4%) α-thalassaemia. Hb Constant Spring was the most prevalent non-deletional α-thalassaemia, accounting for 53.03% of cases. Thirty-one participants (1.33%) exhibited αCD59α/αα, and one (0.04%) had αCD59α/-α3.7. Among the 32 subjects with Hb Adana, 87.5% were Malay, and 12.5% were Orang Asli. Additionally, seven cases of HbE/Hb Adana co-inheritance were identified. Hemoglobin levels in heterozygous Hb Adana individuals ranged from mild anemia to normal, between 95 g/L and 153 g/L. Mean corpuscular volume (MCV) and mean corpuscular hemoglobin (MCH) were approximately 73 fL and 23 pg, respectively. Conclusion This study delineates the distribution ofα-thalassaemia mutation patterns among high school students in Kelantan, Northeast Peninsular Malaysia. Our findings indicate that Hb Adana is rare in our region and co-inheritance with an α-gene deletion results in α+-thalassaemia and with HbE, α0-thalassaemia. All heterozygous Hb Adana individuals exhibited low MCVs and MCHs.

Expression Localization of the KRT32 Gene and Its Association of Genetic Variation with Wool Traits.

Changes in keratin gene expression and spatiotemporal regulation determine the compositional content and cellular localization of wool keratin, thereby affecting wool traits. Therefore, keratin gene family member 32 (KRT32) was selected for a study using RT-qPCR, immunofluorescence, and penta-primer amplification refractory mutation system (PARMS) techniques. The results showed that KRT32 mRNA was highly expressed in the skin and localized to the inner root sheath (IRS), outer root sheath (ORS) and dermal papilla (DP). Sequencing results identified eight SNPs in KRT32, and association analyses revealed that the variations were significantly associated with multiple traits in wool (p < 0.05), including MFD, CF and MFC. The constructed haplotype combination H2H3 has higher CF and smaller MFD than other haplotype combination (p < 0.05). In conclusion, KRT32 can be used as a candidate gene for molecular genetic improvement of wool in Gansu Alpine Fine-wool sheep.

IGF2BP2 and IGFBP3 Genotypes, Haplotypes, and Genetic Models Studies in Polycystic Ovary Syndrome.

Insulin resistance has been correlated with the genetic diversity within the insulin-like binding proteins genes. Moreover, insulin resistance is one of the key characteristics of the widespread reproductive endocrine condition known as polycystic ovarian syndrome (PCOS). Hence, this study is aimed to determine the association between IGFBP3 and IGF2BP2 gene variants and PCOS risk. A total of 300 subjects (150 PCOS cases diagnosed based on Rotterdam ESHRE/ASRM consensus criteria and 150 healthy subjects) were recruited in this case-control cross-sectional study. Tetra-primer amplification refractory mutation system polymerase chain reaction (ARMS-PCR) was used for genotyping rs11705701, whereas genotyping of rs1470579 and rs2854744 was done employing PCR-restriction fragment length polymorphism (PCR-RFLP) technique. The CC and AA+AC genotypes of rs1470579 conferred an increased risk of PCOS in our population. Regarding the rs2854744, an increased risk of PCOS was observed under the codominant homozygous (TT vs. GG) model by 2.54 fold. The C allele of rs1470579 and T allele of rs2854744 enhanced PCOS risk by 1.97 and 1.46 folds, respectively. Haplotype analysis showed that the Ars1470579Ars11705701 haplotype conferred a decreased risk of PCOS (odds ratio = 0.53, 95% confidence interval = 0.34-0.83, p = 0.006). The AC/GG/GT, AA/GA/GT, AC/GA/GG, and AC/GA/GT genotype combinations of rs1470579/rs11705701/rs2854744 were associated with a decreased risk of the disease. IGF2BP2 rs1470579 and IGFBP3 rs2854744 enhanced PCOS susceptibility in a Southeastern Iranian population. Further investigation involving larger cohorts representing diverse ethnic backgrounds is needed to confirm the current findings.

Study the Association of Fat Mass and Obesity-associated (FTO) Gene Polymorphisms (SNP rs9939609) with Biochemical Markers in Obese Iraqi Patients

This study aimed to evaluate the association of single nucleotide polymorphisms (SNP rs-9939609) of the fat mass and obesity-associated (FTO) gene with body mass index (BMI) and some biochemical markers in obese patients. The fat and mass obesity (FTO) gene variant specific single nucleotide polymorphism (SNP rs-9939609) with the T to A missense mutation may have a powerful association with obesity. This case-control study included 106 obese patients (57 males and 49 females) and 80 healthy control (40 males and 40 females). DNA was extracted from whole blood, and then the tetra-primer amplification refractory mutation system - polymerase chain reactions (ARMS.PCR) technique was used to limit the single nucleotide gene polymorphisms (rs9939609) of the FTO gene. Lipid profile, glycated hemoglobin (HbA1C), fasting blood sugar, insulin and FTO concentrations were measured by standard methods. In this study, the A allele in the rs9939609 was at a higher frequency of 35 (33.1%) in the obese patients and at a significant p-value = 0.039 compared with control 15 (18.7%). It significantly increased in the additive model and allele frequency (p= 0.003, 0.002 respectively). The rs9939608 SNP showed a significant association with increased BMI, insulin and homeostatic model assessment-insulin resistance (HOMO-IR) with p-values of 0.001, 0.001 and 0.028 respectively. After making adjustments for age and sex, lower levels of high-density lipoprotein (HDL) were observed in the AA and TA genotypes compared to the TT genotype (p = 0.004). While, no significant differences were recorded between the rs9939608 SNP and HbA1C, total cholesterol (TC), triglyceride (TG), very low-density lipoprotein (VLDL), and low-density lipoprotein (LDL) in obese patients

Analysis of Clinical and Genetic Factors of Obesity and Psoriasis Concomitance-The Influence of Body Mass Composition, Prevalence of Mood Disorders, Environmental Factors and FTO Gene Polymorphisms (rs9939609, rs1558902).

The study enrolled 30 overweight or obese adult patients with chronic plaque psoriasis and 30 overweight or obese volunteers (northern Poland region, Caucasian population). Mood disorders, body mass composition by using bioelectrical impedance analysis (BIA) and FTO gene polymorphisms (rs9939609, rs1558902) by tetra-primer amplification refractory mutation system PCR (T-ARMS-PCR) were assessed. Results revealed significantly higher visceral adipose tissue levels in psoriatic patients (5.23 ± 2.29 [L] vs. 3.41 ± 1.86 [L]), p = 0.001), especially among men, along with elevated rates of moderate and severe depression (26.67% vs. 6.67% and 13.33% vs. 3.33%, p = 0.048 respectively). Additionally, FTO gene polymorphisms correlated with waist-hip ratio differences in both groups. The study highlights the importance of evaluating body composition beyond body mass index, recognizing its influence on psoriasis and associated conditions like depression. The FTO gene may serve as a potential genetic link between psoriasis and obesity, warranting further research for validation. Adiposity emerges as a key and modifiable risk factor, underscoring the clinical implications of body composition complexities in psoriasis management.