Single-strand Conformation Polymorphism Research Articles

Single Nucleotide Polymorphism-based Identification of Bacterial Artificial Chromosome-mediated Homologous Recombination.

Bacterial Artificial chromosome (BAC) recombineering is a powerful genetic manipulation tool for the efficient development of recombinant genetic resources. Long homology arms of more than 150 kb composed of BAC constructs not only substantially enhance genetic recombination events, but also provide a variety of single nucleotide polymorphisms (SNPs) that are useful markers for accurately docking BAC constructs at target sites. Even if the BAC construct is homologous to the sequences of the target region, different variations may be distributed between various SNPs within the region and those within the BAC construct. Once the BAC construct carrying these variations was precisely replaced in the target region, the SNP profiles within the target genomic locus were directly replaced with those in the BAC. This alteration in SNP profiles ensured that the BAC construct accurately targeted the designated site. In this study, we introduced restriction fragment length polymorphism or single-strand conformation polymorphism analyses to validate and evaluate BAC recombination based on changes in SNP patterns. These methods provide a simple and economical solution to validation steps that can be cumbersome with large homologous sequences, facilitating access to the production of therapeutic resources or disease models based on BAC-mediated homologous recombination.

Polymorphisms in the Growth Hormone Gene and Their Association with Pre-pullet Body Weight in Kuttanad Ducks of Kerala, India

Background: Kuttanad ducks are dual purpose native ducks found in Kerala state of India. Identification of single nucleotide variations in candidate genes like growth hormone gene and exploring their association with body weight at grower and pre-pullet stages is essential for the marker assisted selection of breeder ducks for genetic improvement. Methods: Genomic DNA was extracted from blood samples collected from 101 Kuttanad ducks. PCR-amplification of two duck growth hormone gene (dGH) loci viz, exon 3 and partial intron 3 (227 bp), exon 5 and partial intron 4 (366 bp) were undertaken using custom-synthesised primers. The 366 bp region was genotyped using Single Strand Conformation Polymorphism. The 227 bp region was genotyped using DNA pooling assay to yield a 139 bp product for subsequent High-Resolution Melting analysis. The association of dGH genotypic variants for these loci with fortnightly body weight at pre-pullet stages up to 16 weeks of age was analysed. Result: In the 366 bp fragment, GG and GA genotypes were revealed with one novel SNP (g 74G greater than A). The GG homozygotes had significantly higher body weight at all stages from two to 14 weeks of age (p≤0.01). A novel SNP (g 71A greater than G) was revealed at the 139 bp locus within the 227 bp region with three genotypes viz., AA, AG and GG. The GG homozygotes had significantly higher body weight at six, 14 and 16 weeks of age (p≤0.05). The study revealed that dGH polymorphic loci have jointly contributed to the variation in growth and hence can be used as markers for the primary selection of growers at sixth week of age followed by their secondary selection at 14-16 weeks of age during the multi-stage selection programmes for breeder ducks.

A Pilot Study Based on the Correlation Between Whole Exome and Transcriptome Reveals Potent Variants in the Indian Population of Cervical Cancer.

Cervical malignancy (CC) is the 2nd most prevalent malignancy among females, leading to cancer mortality. Primary detection of CC tumors results in an improved prognosis. CC is a malignant gynecological tumor, with few treatment options. New diagnostic and therapeutic agents are required to expand patient survival and quality of life. If CC tumors can be found at an early stage, the prognosis is much brighter. New diagnostic and therapeutic agents are needed to increase patient survival and quality of life. In this work, we performed whole-exome sequencing utilizing V5 (Illumina platform) 10 samples, 5 control and 5 CC tumour tissue, and we compared the results with transcriptome studies. KMT2C variations were shown to be among the most vicious in this analysis. From an Indian viewpoint, we found a plethora of SNVs and mutations, including those with known, unknown, and possible effects on health. Based on our findings, we know that the KMT2C gene is on chr. Seven and in exon 8, all three recognized variants are missense, synonymous, coding synonymous, non-coding variants, and GnomAD MAF (- 0.05). The variation at position (7:152265091, T > A, SNV 62478356) in KMT2C is unique, potent, and pathogenic. The missense coding transcript CIQTNF maps to chromosome 7 and displays T > C SNV. In addition, we performed single strand conformational polymorphism analysis on 64 samples and further confirmed them using Sanger sequencing to understand and verify the mutations. KMT2C shows a log FC value of - 1.16. Understanding emerging harmful mutations from an Indian viewpoint is facilitated by our bioinformatics-based, extensive correlation studies of WES analysis. Potentially harmful and new mutations were found in our preliminary analysis; among these ten top mutated genes, KMT2C and CIQTNF were altered in ten cases of CC with an Indian phenotype.

Analysis of mutational variations in TP53 tumour suppressor gene among Pakistani head and neck cancer patients.

The aim of this study was to determine the frequency of TP53 mutation among Pakistani head and neck cancer (HNC) patients who visited Nishtar Hospital Multan and Nishtar Institute of Dentistry (NID), Multan, Pakistan. While significant research has been conducted on the role of p53 in HNC throughout the world, this study is the first of its kind in Southern Punjab, Pakistan. A total of 242 samples (121 cases and 121 controls) were collected from Nishtar Hospital Multan and NID, Multan, Pakistan. After histopathological analysis to determine the stage type and grade of malignancy, DNA extraction and sequencing were carried out to assess any mutations in the TP53 region (exons 5-8). Genetic screening was performed by the polymerase chain reaction (PCR)-single strand conformation polymorphism (SSCP) technique and Chromas 2.6.6 was used to visualise the sequencing results. The mean age of cases was 50.73 ±16.41 years and controls were 37.55 ± 15.51 years. The frequency of HNC was higher in male patients (65.28%) than in female patients (34.71%). Overall, this cancer was found to be significantly more prevalent in the age group of >35-55 years (p < 0.001). Smoking (51% versus 14%), naswar usage (15.7% versus 6.6%), poor oral hygiene (52.9% versus 29.8%) and anemic status (57.0% versus 4.1%) were significantly associated with cases (p ≤ 0.05) compared to controls. Only 04 samples exon 5 (1 sample), exon 7 (2 samples) and exon 8 (1 sample) with changed mobility patterns were found on the SSCP gel. All mutations were missense and heterozygous. Out of four mutant samples, three mutations were in the hotspot regions (codon 175, 245 and 248) of p53. Based on this study, there may be a weak association between the TP53 exon 5-8 mutation and HNC patients in Southern Punjab, Pakistan.

Four microRNA gene polymorphisms are associated with Iraqi patients with colorectal cancer

BackgroundColorectal cancer (CRC) is a major cause to global cancer-related mortality. The development of colorectal cancer is linked to hereditary variables that exhibit variability.ObjectivesThe objective of this investigation was to assess the potential correlation between microRNA gene polymorphisms and colorectal cancer (CRC) throughout the Iraqi population.MethodsDNA samples were obtained from a cohort of 100 individuals diagnosed with the (CRC) disease, as well as 100 samples as control group. Four primers were designed to amplify four specific high-frequency variants found within microRNA molecules. These variants include Mir146a G\\C, Mir423 A\\C, Mir196a2, and Mir370. The genotyping of the PCR fragments was performed using the single-strand conformation polymorphism (SSCP) method, followed by direct sequencing of each genotype.ResultsGenotyping experiments confirmed the variability of four targeted variants, namely Mir146a G\\C, Mir 423 A\\C, Mir196a2, and Mir370 tend to exhibit a significant association with (CRC). Individuals with Mir146a: GC and Mir 423 A\\C genotype showed a possible association with the increased risk of (CRC), respectively (P = 0.001; OD 0.50; CI 95% 0.33–0.76; P = 0.002; OD 0.53; CI 95% 0.36–0.80). Individuals with Mir196a2: TT and Mir370 GG genotype exhibited a potential association with (CRC) (P = 0.017; OD 0.44; CI 95% 0.22–0.86; P ≤ 0.001; OD 0.24; CI 95% 0.11–0.50).ConclusionsThe study reveals that single nucleotide polymorphisms (SNPs) in microRNA have a notable and distinct correlation with the heightened susceptibility to colorectal cancer (CRC).

KRAS and BRAF genetic alterations in lung cancer: A case - control study.

Lung cancer (LC) is one of the most critical neoplastic abnormalities, having globally a high mortality rate. Knowledge about its genetic mutations and their association with clinically pathological features of LC is very important. Here, we describe the epidemiological molecular study of genetic mutations in KRAS and BRAF genes and their relationship with the demographic and clinical characteristics of Pakistani patients with lung adenocarcinoma. To analyze the mutations of KRAS and BRAF in LC patients among Pakistani population. The study has been carried out at universities and health institutes of Islamabad, Pakistan. Deoxyribonucleic acid (DNA) was extracted from the patient samples by using the standard protocol and amplified by using the specific primers. Later on, the Polymerase Chain Reaction (PCR) products were examined with the help of single stranded conformational polymorphism (SSCP). Relationship between KRAS, BRAF mutations, and LC risk was accessed by conditional logistic regression using SPSS version 24.0. Results were illustrated by odds ratio (OR), 95% confidence interval (CI), and P value. LC is more common in male population and smoking is one of the leading risk factors for (p < 0.0001) LC. KRAS and BRAF mutations were found to be contributing factors toward LC development and showed statistically significant results along with conformation through computational analysis. It can be concluded that smoking is lethal and cancer causing. The concomitant mutations found in KRAS and BRAF were infrequent, and they probably have a very unusual effect on the clinical management of Pakistani patients with lung adenocarcinoma.

FBXW7 polymorphism asserts susceptibility to colorectal cancer

FBXW7, belonging to the F-Box protein family, is considered a candidate cancer susceptibility gene. Our findings indicate that single nucleotide polymorphisms (SNPs) in the FBXW7 gene are linked to cancer risk, strengthening FBXW7′s role in the pathogenesis of colorectal cancer. Our case-control study comprised of 450 patients diagnosed with colorectal cancer (CRC) and an equal number of 450 healthy subjects. FBXW7 SNPs rs2255137C>T and rs6842544C>T were genotyped using PCR-Restriction Fragment Length Polymorphism (PCR-RFLP) and Single-Stranded Conformation Polymorphism (SSCP) techniques and further cross-checked by direct sequencing. Linkage disequilibrium and haplotype analyses of these SNPs were also assessed. The in-silico approach was used to reveal the functional analysis between the nonsynonymous variation (rs6842544) and CRC followed by its validation at the protein level by western blotting and reverse transcription-PCR. A significant association of colorectal cancer was detected with rs6842544 SNP. However, there was no association between FBXW7 rs2255137 polymorphism and CRC. The homozygous individuals carrying the C variant in FBXW7 rs6842544 showed a slightly higher risk for colorectal cancer (OR = 1.590, 95%CI = 0.39 ∼ 2.89, p = 0.011). The haplotype CC identified in this study seemed to be associated with good prognosis (OR = 1.22, 95% CI = 1.00 ∼ 1.47, p = 0.0013) whereas the TT haplotype was found to reduce the CRC risk (OR = 0.642, 95%CI = 0.48 ∼ 0.84, p = 0.039). In-silico prediction proposed that the variant R133G is responsible for the lower expression of FBXW7. Additionally, the expression profiling of FBXW7 nonsynonymous SNP was significantly lower in primary CRC tissues than in the paired non-cancerous tissues at protein and mRNA levels. The study indicates that the FBXW7 rs6842544 is associated with the risk of development of CRC and could serve as a molecular biological marker to screen high-risk groups for CRC.

Detection of new single nucleotide polymorphisms in plasma cells carcinoma (myeloma) by polymerase chain reaction–single strand conformation polymorphism and sequencing technical methods

Abstract Background: MicroRNA (miRNA) is a short, single-stranded, non-coding sequence of RNA that does not translate into proteins. Because of their ability to control gene expression, recent studies have shown that these short, non-coding sequences play an important role in cancer. So that, detection the polymorphisms or variants in miRNA genes may help to identify their correlation to cancer susceptibility. Objectives: Study the polymorphisms in miRNA genes or variants miRNA genes and their correlation with myeloma disease. Materials and Methods: Twenty samples were collected from people who visited Marjan Hospital in Province of Babylon, Iraq, from February to July 2022. In addition, 20 samples from healthy people were collected as control. Genotyping of miRNA was accomplished using the polymorphism techniques of polymerase chain reaction and single strand conformation polymorphism (SSCP). DNA sequencing was also applied to categorize all the ranges of SSCP models detected using only gel imaging. Results: The genotypes obtained in this experiment verified that several single nucleotide polymorphisms were obtained between 5-band and 6-band miRNA National center for biotechnology information Primer3 plus reference. Conclusion: The results explain that the miRNA gene polymorphisms may have correlation with the susceptibility to myeloma disease.

Genetic variants in the TAC1 transcriptional regulatory region affect on trainability and excitability levels in Belgian Malinois dogs.

Trainability in dogs is affected by learning aptitude and memory capacity. While this trait has a heritable basis in canines, the specific genetic loci responsible for it remain unknown. Our previous results suggested that the BDNF, CCK and TAC1 genes are associated with learning and memory in canines. Experimental validation is crucial to confirm the effects of these candidate genes on trainability. Understanding the genetic foundation of this trait would offer insight into the inheritance pattern of complex behavioural characteristics. This study aimed to assess the genetic variations within candidate genes and explore their potential associations with behavioural phenotypes in dogs. The behavioural characteristics of 123 male Belgian Malinois dogs were assessed using a customised questionnaire. Target regions of candidate genes were screened for genetic variation by single-strand conformational polymorphism (SSCP). Following that, SSCP banding patterns were sequenced, and putative transcription factor binding sites were predicted using bioinformatics tools. Quantitative association analysis between identified genetic variants and behavioural trait scores was performed using the general linear model (GLM). Sequencing the coding and flanking regions revealed three mutations (c.-89C>T, c.-162G>C and c.*33T>A) in the dog TAC1 gene. Bioinformatics analysis predicted two single nucleotide polymorphisms (SNPs) were located within the putative TAC1 promoter and could disrupt transcription factor binding sites. Statistical tests revealed that the c.-89C>T was significantly associated with excitability (p < 0.01), while the c.-162G>C was significantly associated with trainability level (p < 0.05). In summary, we identified two regulatory SNPs in the 5'-UTR promoter region of the TAC1 gene that are associated with excitability and trainability in Belgian Malinois dogs. These genetic variations have the potential to alter the binding sites of transcription factors NRF1 and OTX1, consequently influencing TAC1 expression and related behavioural characteristics. Our findings implicate TAC1 polymorphisms as candidates influencing breed-specific behavioural characteristics in canines. Further studies on diverse breeds of dogs are necessary to validate these SNPs' effects.

Reproduction of Awassi and Hamdani Sheep Is Associated With a Novel Missense SNP (p.24Ile>Thr) of the GnIH Gene.

Litter size is a crucial economic factor in the sheep industry. Several factors and genes influence litter size, making the identification of genes or loci involved a genetic challenge. Gonadotropin-inhibitory hormone (GnIH) is one of several genes that influence sheep's reproductive traits. Thus, this study aimed to investigate whether variations in the GnIH gene affect the reproductive performance of Awassi and Hamdani ewes. DNA was extracted from 99 single-progeny ewes and 101 twin ewes. The polymerase chain reaction (PCR) produced amplicons of 262 bp, 275 bp, and 284 bp from exons 1, 2, and 3 of the GnIH gene. Single-strand conformational polymorphism (SSCP) technique was used for genotyping experiments. Sequencing and in silico analysis were performed on each set of SSCP-resolved bands. Two genotypes of 262 bp amplicons were found: TT and TC. Sequence analysis revealed a novel missense mutation in the TC genotype at position c.122T>C. Five in silico tools were used to assess the impact of this mutation on GnIH protein structure, function, and stability, all of them demonstrated a deleterious effect. An analysis of statistical data revealed a strong correlation between the c.122T>C single-nucleotide polymorphism (SNP) and reproductive performance. Ewes with the SNP 122T>C exhibited a significant increase in litter size, twinning rates, lambing rates, and days to lambing when compared with ewes with the TT genotype. A lower number of lambs were born to ewes with the TT genotype than those with the TC genotype. These results concluded that the c.122T>C SNP variant positively influences the reproductive performance of Awassi and Hamdani sheep. Sheep that carry the c.122T>C SNP show higher litter size and increased productivity.

Correlation analysis between polymorphism of leptin and IGFI genes and body measurements in Barki and Farafra sheep

BackgroundGrowth performance, which can be evaluated using various body measurements, is crucial for providing red meat for human consumption. This study aims to improve the growth performance of sheep by identifying the different genotypes of leptin and IGFI genes associated with higher body measurements.MethodsIn this study, we collected blood samples from Barki (n = 30) and Farafra (n = 30) sheep, and measured their body height, width, length, and weight, and tail width and length to assess their growth performance. We used single strand conformation polymorphism polymerase chain reaction (SSCP-PCR) and sequencing analysis to identify single nucleotide polymorphisms (SNPs) and different genotypes of the leptin and insulin-like growth factor receptor (IGFI) genes associated with sheep body measurements.ResultsThe leptin gene (209 bp) and IGFR gene (345 bp) were found to have three (TT, TG, and GG) and (TT, TC, and CC) genotypes, respectively, by SSCP-PCR. Sequence analysis revealed a SNP in the leptin gene (200G > T) with two alleles (G allele: OR058867 and T allele: OR058868). This SNP resulted in the substitution of the amino acid valine with leucine, which increased the width, length, and weight of the body and tail width in the heterozygous form (TG). The IGFI gene had three SNPs: 305 G > T in TT genotype (OR260670), 184C > T, 265T > C, and 305G > T in TC genotype (OR260669), and 265T > C in CC genotype (OR260671). The TC genotype of the IGFR gene was correlated with higher values of body measurements. Barki sheep were found to have the highest values for body weight, width, length, and tail width compared to Farafra sheep.ConclusionWe recommend the use of entire Barki sheep in inbreeding programs to improve growth performance in Egyptian sheep.

In Silico Analysis of Functional SNPs in Genes of Complete Androgen Insensitivity Syndrome (CAIS): A Retrospective, Case-Control Study.

Complete androgen insensitivity syndrome (CAIS) is one of the categories of androgen insensitivity syndrome (AIS) described as complete failure of the cell to react to androgens with external genitalia of a normal female. People with AIS condition are genetically male, with XY karyotype in each cell, but their bodies are unable to respond to male sex hormones (called androgens). It is associated with infertility as well as developing cancerous conditions. The genetic association of CAIS involves polymorphism of genes such as NR5A1, SOX9, SRD5A2, CBX2, GATA4, and SRY. Their mutation and participation in genetics of CAIS can be studied by Single Nucleotide polymorphism (SNP) analysis which is a way to detect genetic variations. SNP in coding region leads to synonymous and non-synonymous mutations. Hence, this study highlights analysis of SNPs associated with CAIS. Our aim is to study SNP analysis of NR5A1, SOX9, SRD5A2, CBX2, GATA4, SRY genes in Complete Androgen Insensitivity Syndrome. SIFT and Polyphen analysis was performed for all the genes and samples were subjected for PCR-SSCP technique. SNPs were analyzed for the genes associated with CAIS. Benign and damaging SNPs were identified. DNA Samples were amplified using PCR technique and they will be analyzed using Single-strand conformation polymorphism (SSCP). As SNPs have decreased stability, damaging and benign character, they can be used as candidate hallmarks in study of Complete Androgen Insensitivity Syndrome.

Regional bias of tumor suppressor gene mutations of STARD8 and WNK2 in colon cancers

StAR-related lipid transfer domain protein 8 (STARD8), encoding a Rho-GTPase-activating protein, and WNK2, encoding a serine/threonine kinase are candidate tumor suppressor genes (TSGs) in human cancers. Inactivation of these genes that would promote cancer pathogenesis is largely unknown in colon cancer (CC). Our study addressed to address whether STARD8 and WNK2 genes are mutated in CC. STARD8 and WNK2 genes possess mononucleotide repeats in their exons, which could be the targets for frameshift mutations in cancers with high microsatellite instability (MSI-H). By single-strand conformation polymorphism (SSCP) analysis, we analyzed the repeated sequences in 140 CCs (95 CCs with MSI-H and 45 CCs with stable MSI (MSS)). By DNA sequencing, we found that five MSI-H CCs (5/95: 5.3%) harbored the frameshift mutations, whereas MSS CCs (0/45) did not. In addition, we detected regional heterogeneous frameshift mutations of these genes in four (25%) of 16 MSI-H CCs. In immunohistochemistry for WNK2, WNK2 expression in the MSI-H CCs was significantly lower than that in the MSS CCs. Our results for the mutation and expression indicate that STARD8 and WNK2 genes are altered at various levels (frameshift mutation, expression, and regional heterogeneity) in MSI-H CCs, which might play a role in the pathogenesis by inactivating their TSG functions.

Genetic variations of α-casein and K-casein genes associated with milk composition in goats

The quality of animal milk varies between individuals of the same breed. The present study aimed to detect the favorable SNPs for α-casein (CSN1S1) and k-casein (CSN3) genes associated with high-quality milk traits. Blood and milk samples of Zaraibi and Damascus goats were collected from Sakha farm in Kafr El-Shiekh. The solid non-fat, lactose, protein, and fat contents, were determined in milk using the biochemical method, and amino acids were determined using HPLC. To identify genotypes and nucleotide variations (SNPs), single-strand conformation polymorphism (SSCP) and sequencing methods were used. One-way ANOVA test was used to determine the effect of different genotypes on milk traits. The SSCP-PCR results in Zaraibi and Damascus goats showed that the CSN1S1 gene has 3 genotypes (AA, AB, and BB), and also the CSN3 gene has 3 genotypes (AA, AG, and GG). The results of sequencing showed three SNPs at positions 41G˂T, 69A˂G, and 80C˂T in the CSN1S1 gene, and three SNPs at positions 49C˂T, 111A˂G, and 340C˂T in the CSN3 gene. The milk results showed that the high protein, fat, and lactose content were associated with the AB genotype in the CSN1S1 gene while they were associated with the AG genotype in the CSN3 gene. Likewise, essential and branched-chain amino acids were increased in goat milk with the AB and AG genotypes of CSN1S1 and CSN3 genes, respectively. The essential amino acids for protein synthase were higher in Zaraibi than in Damascus goats. Therefore, the Zaraibi goats may more possess genotypes in the CSN1S1 and CSN3 genes, which affect the fat and protein level in milk.

Ovine KRTAP36-2: A New Keratin-Associated Protein Gene Related to Variation in Wool Yield.

Keratin-associated proteins (KAPs) are structural components of wool fibres. High-glycine/tyrosine (HGT)-KAPs are a subset of the KAP family, and their abundance in fibres varies. In this study, we report the discovery of an ovine HGT-KAP gene to which we assigned the name KRTAP36-2. Polymerase chain reaction and single-strand conformation polymorphism (PCR-SSCP) analyses revealed four variants of this gene in a screening population of 170 sheep from a variety of breeds. The DNA sequencing of the variants revealed four single-nucleotide polymorphisms (SNPs) and a dinucleotide deletion. Three of these SNPs were in the coding region, and one of these was non-synonymous and potentially led to the amino acid substitution p.Cys27Gly near the middle of the protein. The remaining SNP was located near the putative TATA box, and the di-nucleotide deletion was near the putative transcription initiation site. The effect of this variation in KRTAP36-2 was investigated in 274 Southdown × Merino lambs that were the progeny of five sires. Variation was only found to be associated with wool yield, that is, the proportion of the greasy fleece that remained as clean fleece upon scouring (expressed as a percentage). This may have some value in increasing wool production.

Transmission of isolates of Citrus tristeza virus by single aphids (Toxoptera citricida)

Citrus tristeza virus (CTV) can be transmitted by several aphid species in a semi-persistent mode, with Toxoptera citricida being the most efficient vector. In Brazil, mild CTV isolates are used for pre-immunization of citrus trees against severe isolates. We aimed to determine the capacity and efficiency of T. citricida in separating the viral complex into haplotypes from three well-characterized CTV isolates (PIAC, CS1, and Forte Rolândia). Single-aphid transmission assays were conducted to determine CTV transmission efficiency. The results showed that T. citricida transmitted only haplotypes from the PIAC and CS1 isolates, with efficiencies of 8 and 4%, respectively. Both isolates caused mild CTV symptoms in Brazil. However, isolate Forte Rolândia, which causes severe symptoms in citrus trees, was not transmitted by T. citricida. The detection of CTV haplotypes from PIAC and CS1 isolates in sweet orange (Citrus sinensis) plants after a single aphid transmission occurred at different time points. The first haplotype observed was from PIAC 150 days after citrus was challenged by a single aphid, followed by CS1 at day 210 after transmission. In addition, differences in the single-strand conformation polymorphism patterns between the CTV isolates and CTV haplotypes were determined, suggesting that an aphid can acquire and transmit only one CTV haplotype to citrus plants during phloem feeding. The study of the mechanism of transmission by the vector can increase our knowledge of the interactions among hosts, vectors, and pathogens, which are often neglected.

Analysis of genetic polymorphism in genes encoding cathelicidins from Vietnam indigenous yellow cattle breeds

Cathelicidins include antimicrobial peptides (AMPs) and are involved in the innate immune defense against infections in mammals. Polymorphisms in the DNA sequence of cathelicidin genes could be relevant to inherited variations of host innate immunity, adaptation, and pathogen resistance. This study aims to investigate the sequence polymorphism of cathelicidin genes including CATHL2 and CATHL4 from local indigenous yellow cattle breeds of Vietnam. Genomic DNA samples were extracted from 52 individuals collected from different cattle populations in Vietnam including Ha Giang, Thanh Hoa, Nghe An and Phu Yen. The CATHL2 and CATHL4 genes were amplified by PCR, following sequencing for identification of the single nucleotide polymorphisms (SNPs) and/or insertion-deletion (indel) DNA sequence variations. The copy number variations (CNVs) of the CATHL4 gene were determined by cloning, single strand conformation polymorphism (SSCP) and real-time PCR. The comparative analysis results showed that there are 13 SNPs detected in all sequences of CATHL2 gene including 9 SNPs in intron 2 and intron 3, and 4 missense substitutions in exon 1, 2, and 3. These SNPs do not affect the predicted tertiary structure of Bac5 encoded by this gene. For CATHL4, the results revealed 15 SNPs, 3 indels and 1 repeat variation as motif (TG)n(G)n/(AC)n(C)n. Among these variations, the deletion 46delC in exon 1 of CATHL4 was detected in a sample of the Ha Giang breed causing a truncated polypeptide as predicted. The average copy number of CATHL4 of Vietnam indigenous cattle breeds using GADPH as a reference gene showed in range from 2.53 to 2.89 copies.

Monitoring Drought Tolerance in Oil Palm: Choline Monooxygenase as a Novel Molecular Marker.

Water scarcity negatively impacts oil palm production, necessitating the development of drought-tolerant varieties. This study aimed to develop molecular markers for oil palm breeding programs focused on drought tolerance. Genes associated with drought tolerance were selected, and single nucleotide polymorphism (SNP)-based markers were developed. Genomic DNA was successfully extracted from 17 oil palm varieties, and 20 primers out of 44 were effectively amplified. Screening with single-strand conformation polymorphism (SSCP) revealed an informative SNP marker from the choline monooxygenase (CMO) gene, exhibiting CC, CT, and TT genotypes. Notably, the oil palm variety La Mé showed the CT genotype, while Surat Thani 2 (Deli × La Mé) exhibited the CT and CC genotypes in a 1:1 ratio. Gene expression analysis confirmed the association of the CMO gene with drought tolerance in commercial oil palm varieties. The full-length CMO gene was 1308 bp long and shared sequence similarities with other plant species. However, amino acid sequence variations were observed compared with existing databases. These findings highlight the potential utility of the CMO marker for drought tolerance selection, specifically within the La Mé parent of oil palm Surat Thani 2 varieties, and strongly confirm the La Mé S5 population and Surat Thani 2 as drought-tolerant varieties.

Association Between of the Growth Hormone Gene GH-1 with Growth Indicators of Common Carp Cyprinus carpio L. in Three Different Ecosystems

This study was conducted in the central of health laboratory/Animal and Fish Resource Research Center, Agricultural Research Department, Ministry of Science and Technology, for the period from 15/8/2021 to 15/11/2021 in order to investigate the polymorphism of the growth hormone gene GH-1 and its relationship to the productive performance (body weight, total length, age and case factor) of common carp Cyprinus carpio, in 90 fish representing three different ecosystems (30 fish for each site) including floating cages, ponds, and the environment of the Tigris River, south of Baghdad, with an average weight of (975-1517±150) g. Some physical and chemical measurements of water were measured, including temperature, dissolved oxygen concentration, pH, and salinity. These fish were transferred to the laboratory, and the weight, total length, and age of each fish were measured, as well as DNA extraction for the purpose of amplifying the size of a segment (459pb) of the growth hormone gene that includes exon 4, intron 4, and exon 5 by polymerase chain reaction PCR using primers designed for this gene. The genotypes of the growth hormone gene were investigated using the Single-Stranded Conformation Polymorphism (SSCP) technique, and the produced bands were compared to distinguish different genotypes of the gene by the electrophoresis method. The results showed that the water temperature was close to the environmental systems under study and ranged between (25.4-27 ± 0.50), the rates of dissolved oxygen concentration were (6.5 ± 0.09) mg/l, pH (7.0 ± 0.02) and salinity 0.8 g/l higher in ponds compared to the lowest value of them in the river. The analysis of the SSCP technique revealed the presence of six different genotypes of this hormone, including A, B, C, E, F, and H with a frequency of 24.4, 5.6, 15.6, 15.6, 16.4, and 22.4, respectively, for the fish community in the studied sites. The H genotype was significantly (P< 0.05) superior in the floating cages in the average body weight (1600 ± 0 g) and the relative case coefficient (2.5 ± 0). significant differences were observed in the average total body length and age, according to the multiplicity of genotypes. We conclude from this study an improvement in the growth and health status of common carp in cages and ponds compared to the river at the probability level (P<0.05) and the possibility of focusing on the polygenic genotypes of the growth hormone gene and adopting the H genotype as a guide for selecting fish mothers in genetic improvement programs for artificial propagation and achieving the largest economic return from fish breeding projects.

Screening for causative mutations in ovine BMPR1B and BMP15 genes and their homologous fragments in human.

The BMPR1B and BMP15 genes are well known for their considerable associations with prolificacy in sheep. These genes may also affect fertility or prolificacy in other species, including human. This study was conducted to investigate possible causative mutations in BMPR1B and BMP15 genes in human and an indigenous breed of sheep. Blood samples were collected from 83 singleton- and prolific Mehraban ewes and 81 infertile, singleton- and twin-bearing women. A 190-bp fragment, containing the FecB mutation in ovine BMPR1B, a 380-bp fragment in ovine BMP15 gene and their homologous fragments in human were amplified and then investigated by single-stranded conformation polymorphism and DNA sequencing methods. The FecB mutation of BMPR1B (g.159A>G) was detected in the sheep population, but no polymorphic loci were found in the homologous fragment in studied human samples. The studied fragments of BMP15 were monomorphic in both sheep and human samples. A total of nine and 69 point-differences in the studied fragments of BMPR1B and BMP15 genes were detected between the species, respectively. In sheep, the G allele of BMPR1B had a positive effect on litter size (p<0.05), whereby all AG or GG ewes were prolific. The FecB mutation for the first time was detected in Mehraban sheep and therefore could be considered for marker-assisted selection in this breed. The studied fragments of BMPR1B and BMP15 genes are not responsible for reproduction variation in human. More studies on other genes, associated with fertility in human, are necessary in the future.