scholarly journals Identifikasi Mutasi FecX Pada Gen BMP15 dan Pengaruhnya Terhadap Sifat Prolifik pada Kambing Lokal di Kabupaten Lombok Barat

2019 ◽  
Vol 1 (1) ◽  
pp. 1
Author(s):  
Rahmat Agus Hidayat ◽  
Sulaiman Ngongu Depamede ◽  
Maskur Maskur
Keyword(s):  
Litter Size ◽  
Mutant Allele ◽  
Secondary Data ◽  
Wild Type ◽  
Banding Patterns ◽  
Average Litter Size ◽  
Pcr Rflp ◽  
Pcr Products ◽  
Visually Based ◽  
Its Polymorphism

The aims of this study were to identify the mutations of FecX gene in the local goats and to analyze its polymorphism as well as its influence on the prolific nature of the local goats in West Lombok Regency, Indonesia. The study was conducted in the Immunobiology Laboratory of Mataram University, using DNA isolated from 100 blood samples of local female goats which have given birth of once to three times. The methods used were PCR-RFLP method and the PCR products were digested with HinfI restriction enzyme (G|ANTC) then analyzed visually based on DNA banding patterns on 2% agarose gels. The frequency of allele and genotype obtained, were then analyzed through a comparison with the secondary data of litter size obtained from the local goat keepers information. The results showed that the gene mutation of FecXG produced two alleles: "wild-type" (+) sized of 110 bp and 31 bp, and the mutant allele (G) of 141 bp with the allele frequency of 0,965 and 0,035 respectively. Combinations of alleles in the gene BMP15 produced two genotypes, namely (a) genotype ++ (110 bp/110 bp) with a frequency of 0.93, with the average litter size of 1.59 ± 0.319, and (b) genotype G + (141bp/110 bp), with a frequency of 0.07 and with the average litter size of 1.65 ± 0.202. The results of this study indicated that mutation occurred in BMP15 gene, i.e. FecXG gene, the gene responsible for the prolificacy of animals studied. Furthermore there was a correlation between polymorphism of FecXG gene and the prolific nature of the local goats, which was predicted to lead the divergence in litter size of each local goat genotype  

scholarly journals PCR-based genotyping assays to detect germline APC variant associated with hereditary gastrointestinal polyposis in Jack Russell terriers

2021 ◽  
Vol 17 (1) ◽  
Author(s):  
Kyoko Yoshizaki ◽  
Akihiro Hirata ◽  
Hiroyuki Matsushita ◽  
Naohito Nishii ◽  
Mifumi Kawabe ◽  
...  
Keyword(s):  
Mutant Allele ◽  
False Negative ◽  
Disease Onset ◽  
Pcr Amplification ◽  
Buccal Swab ◽  
Wild Type ◽  
Gastrointestinal Polyposis ◽  
Pcr Rflp ◽  
Formalin Fixed Paraffin Embedded ◽  
Rflp Assay

Abstract Background The prevalence of gastrointestinal (GI) neoplastic polyps in Jack Russell terriers (JRTs) has increased in Japan since the late 2000s. Recently, we demonstrated that JRTs with GI polyps harbor identical germline variant in the APC gene (c.[462_463delinsTT]) in the heterozygous state. Thus, this disease is an autosomal dominant hereditary disorder. Although the affected JRTs have distinct features, such as the development of multiple GI polyps and an early age of disease onset, genetic testing is indispensable for a definitive diagnosis. Here, polymerase chain reaction (PCR)-based assays capable of detecting germline APC variant were designed and validated using synthetic wild-type and mutant DNAs and genomic DNAs from carrier and non-carrier dogs. Result First, the PCR-restriction fragment length polymorphism (PCR-RFLP) assay was developed by taking advantage of the germline APC variant creating a new restriction site for MseI. In the PCR-RFLP assay, the 156-bp region containing the variant site was amplified by PCR and subsequently digested with MseI, yielding diagnostic 51 and 58 bp fragments from the mutant allele and allowing determination of the APC genotypes. It was possible to determine the genotypes using genomic DNA extracted from the peripheral blood, buccal swab, or formalin-fixed paraffin-embedded tissue. Next, a TaqMan duplex real-time PCR assay was developed, where a 78-bp region flanking the variant was amplified in the presence of wild-type allele- and mutant allele-specific fluorescent probes. Using blood-derived DNA, altogether 40 cycles of PCR amplification determined the APC genotypes of all examined samples by measuring the fluorescence intensities. Importantly, false-positive and false-negative errors were never detected in both assays. Conclusion In this study, we developed highly reliable genetic tests for hereditary GI polyposis in JRTs, providing accurate assessment of the presence of the causative germline APC variant. The genotyping assays could contribute to the diagnosis and prevention of hereditary GI polyposis in dogs.

scholarly journals Two methods for genotyping a 4-base deletion in the canine ABCB1 gene

2019 ◽  
Vol 31 (6) ◽  
pp. 889-892
Author(s):  
Carolina A. Silvestro ◽  
Liliana A. Soria ◽  
Adriana Conte ◽  
Graciela Marrube
Keyword(s):  
Molecular Detection ◽  
High Resolution Melting ◽  
Fragment Length Polymorphism ◽  
Genetic Disorders ◽  
Chemotherapeutic Agents ◽  
Specific Method ◽  
Wild Type ◽  
Abcb1 Gene ◽  
Pcr Rflp ◽  
Pcr Products

A 4-bp deletion (c.230_233delATAG) of the ABCB1 gene, frequently found in various dog breeds, results in intolerance to certain drugs routinely used in veterinary medicine, including many chemotherapeutic agents and macrocyclic lactones. The use of rapid and reliable genetic testing is fundamental for early detection of the mutation and prevention of undesirable toxicoses. We developed and compared 2 genotyping tests: PCR–high-resolution melting (PCR-HRM) and PCR–restriction-fragment length polymorphism (PCR-RFLP) to identify the 4-bp deletion in the ABCB1 gene of canine breeds. Amplified PCR products were sequenced in order to confirm different genotypes. Both techniques were efficient in discriminating homozygous wild-type, homozygous mutated, and heterozygous ABCB1 genotypes, and proved to be reproducible and economical methods. The HRM analysis, a sensitive and specific method for the molecular detection of genetic disorders, does not require labeled probes, processing, or separations after PCR.

Genetic polymorphism in fecundity gene (FecG) among Indiansheep breeds Balangir, Shahabadi and Bonpala

2016 ◽  
Author(s):  
Jowel Debnath ◽  
Ran Vir Singh
Keyword(s):  
Litter Size ◽  
Transforming Growth Factor ◽  
Natural Habitat ◽  
Factor B ◽  
Sheep Breeds ◽  
Homozygous Genotype ◽  
Average Litter Size ◽  
Pcr Rflp ◽  
Selection Of ◽  
Gdf9 Gene

FecG (GDF9) is a member of the transforming growth factor-b (TGF- b) superfamily, have been shown to be essential for follicular growth and ovulation. Different mutations in FecG gene caused increased ovulation and infertility in sheep. The present study was designed for screening polymorphism of FecG gene in 250 selected ewes from different sheep flocks representing Balangir (100), Shahabadi (100) and Bonpala (50) by employing forced PCR-RFLP technique. Genomic DNA was extracted from the blood of Balangir, Shahabadi and Bonpala matured ewes with average litter size varying from 1.00± 0.00 to 1.14±0.02 at different parities. Digestion of FecG (GDF9) gene with DdeI restriction enzyme resulted into FecGHH homozygous genotype. In all three sheep breeds, genotypic frequencies of FecGHH were 100% and gene frequency of H allele was unity. This indicates that the FecG gene is fixed in the Balangir, Shahabadi and Bonpala population in the natural habitat. Litter size of Balangir and Bonpala sheep breeds were single but in Shahabadi sheep twin was recorded. In present study all the animals of three breeds were homozygous for FecG and there was no infertility observed in above mentioned breeds in field condition and organized farm, which is not in consonance with previous report. The observed effects could be caused by linkage disequilibrium with other nearby loci. The study revealed that FecG gene is not a reliable genetic marker for selection of high prolificacy in sheep.

scholarly journals Fingerprinting of fecB gene in five Egyptian sheep breeds

2009 ◽  
Vol 25 (3-4) ◽  
pp. 205-212 ◽  
Author(s):  
Hanafy el ◽  
Saadani el
Keyword(s):  
Point Mutation ◽  
Restriction Enzyme ◽  
Base Pair ◽  
Genomic Dna ◽  
Restriction Site ◽  
Wild Type ◽  
Specific Primer ◽  
Sheep Breeds ◽  
Pcr Rflp ◽  
Pcr Products

Recently, many aspects of FecB gene, including reproductive endocrinology, organs development and body mass have been studied. FecB has an additive effect on litter size and ovulation. The present investigation was carried out to study polymorphism by forced PCR-RFLP of FecB gene in five Egyptian local sheep breeds and its comparison with other foreign sheep breeds. Genomic DNA was isolated from a total of 100 animals of Egyptian sheep breeds namely Rahmani, Ossimi, Awassi, Barki and Awassi x Barki crossbred. Forced PCR of the FecB gene 190 base pair (bp) was amplified using specific primer designed to introduc a point mutation in the resulting PCR products with FecB carrier sheep containing an AvaII restriction site (G|GACC), whereas products from noncarriers lacked (of) this site. Digestion of FecB gene 190 base pair with AvaII restriction enzyme resulted in non carrier 190 bp band (wild type) in all the animals belonging to the five Egyptian breeds studied revealing absence of this restriction site in those five breeds. .

scholarly journals PCR-based Genotyping Assays to Detect Germline APC mutations Associated with Hereditary Gastrointestinal Polyposis in Jack Russell Terriers

2020 ◽  
Author(s):  
Kyoko Yoshizaki ◽  
Akihiro Hirata ◽  
Hiroyuki Matsushita ◽  
Naohito Nishii ◽  
Mifumi Kawabe ◽  
...  
Keyword(s):  
Mutant Allele ◽  
False Negative ◽  
Disease Onset ◽  
Pcr Amplification ◽  
Buccal Swab ◽  
Wild Type ◽  
Mutation Site ◽  
Gastrointestinal Polyposis ◽  
Pcr Rflp ◽  
Rflp Assay

Abstract Background: The prevalence of gastrointestinal (GI) neoplastic polyps in Jack Russell Terriers (JRTs) has increased in Japan since the late 2000s. Recently, we demonstrated that JRTs with GI polyps harbor identical germline mutations in the APC gene (c.[462A>T; 463A>T]) in the heterozygous state. Thus, this disease is an autosomal dominant hereditary disorder. Although the affected JRTs have distinct features, such as the development of multiple GI polyps and an early age of disease onset, genetic testing is indispensable for a definitive diagnosis. Here, polymerase chain reaction (PCR)-based assays capable of detecting germline APC mutations were designed and validated using synthetic wild-type and mutant DNAs and genomic DNAs from carrier and non-carrier dogs.Result: First, the PCR-restriction fragment length polymorphism (PCR-RFLP) assay was developed by taking advantage of the germline APC mutations creating a new restriction site for MseI. In the PCR-RFLP assay, the 156-bp region containing the mutation site was amplified by PCR and subsequently digested with MseI, yielding diagnostic 51 and 58 bp fragments from the mutant allele and allowing determination of the APC genotypes. It was possible to determine the genotypes using genomic DNA extracted from the peripheral blood, buccal swab, or formalin-fixed paraffin-embedded tissue. Next, a TaqMan duplex real-time PCR assay was developed, where a 78-bp region flanking the mutation was amplified in the presence of wild-type allele- and mutant allele-specific fluorescent probes. Using blood-derived DNA, altogether 40 cycles of PCR amplification determined the APC genotypes of all examined samples by measuring the fluorescence intensities. Importantly, false-positive and false-negative errors were never detected in both assays. Conclusion: In this study, we developed highly reliable genetic tests for hereditary GI polyposis in JRTs, providing accurate assessment of the presence of the causative germline APC mutations. The genotyping assays could contribute to the diagnosis and prevention of hereditary GI polyposis in dogs.

scholarly journals Molecular Typing of Methicillin ResistantStaphylococcus aureusClinical Isolates on the Basis of Protein A and Coagulase Gene Polymorphisms

2014 ◽  
Vol 2014 ◽  
pp. 1-11 ◽  
Author(s):  
Nancy Younis Omar ◽  
Hala Abdel Salam Ali ◽  
Reem Abdel Hameed Harfoush ◽  
Engy Hamdy El Khayat
Keyword(s):  
Molecular Typing ◽  
Gene Polymorphisms ◽  
Protein A ◽  
Rflp Analysis ◽  
Restriction Digestion ◽  
Banding Patterns ◽  
Pcr Rflp ◽  
Pcr Products ◽  
Mrsa Strains

Increased frequency of methicillin-resistantStaphylococcus aureus(MRSA) in hospitalized patients requires rapid and reliable characterization of isolates for control of MRSA spread in hospitals. This study evaluated polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) as a molecular typing technique for MRSA strains on the basis of protein A (spa) and coagulase (coa) gene polymorphisms to verify their ability in assessing the relatedness of isolates. Seventy-five MRSA isolates, from different ICUs of Alexandria University Main Hospital, were characterized using antibiotyping and PCR-RFLP analysis ofcoaandspagenes. Thirty-two antibiotypes were identified.coagene PCR generated 3 types and 10 subtypes of band patterns.HaeIIIrestriction digestion of amplifiedcoagene products produced 5 major banding patterns and 12 subtypes.spagene PCR products generated 4 major and 11 minor types, and theirHaeIIrestriction digestion showed 5 major and 12 minor banding patterns. The combinedcoaandspaRFLP patterns generated 22 combined R types. Typing usingcoaPCR and PCR-RFLP had the same discriminatory index (DI) value (0.64), which was comparable to that of bothspaPCR and PCR-RFLP techniques (0.68). The combined grouping increased the DI value to 0.836. The current study revealed that testing for multiple gene polymorphisms is more useful for local epidemiologic purposes.

scholarly journals Polymorphisms of TLR4 Asp299Gly and TNF-α -308G/A in Leptospirosis

2016 ◽  
Vol 2 (1) ◽  
pp. 17
Author(s):  
Nur Farhanah ◽  
Muhammad Hussein Gasem ◽  
Sultana MH Faradz
Keyword(s):  
Mutant Allele ◽  
Fragment Length Polymorphism ◽  
Wild Type ◽  
Chain Reaction ◽  
Healthy Group ◽  
Tnf Α ◽  
Pcr Rflp ◽  
Polymerase Chain ◽  
Severe Leptospirosis ◽  
Increased Susceptibility

AbstractBackground : TLR4 Asp299Gly and TNF-α -308G/A polymorphisms have been shown to be associated with increased susceptibility and severity of infection. TLR4 Asp299Gly polymorphism could affect the host’s ability to respond to leptospira sp. TNF-α -308G/A polymorphism, is associated with the high producer of TNF-α.Methods : Total of 36 leptospirosis patients (IgM anti leptospira and MAT positive) and healthy individual with the equal number were included. The polymorphisms were determined  by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) using site spesific restriction enzyme.Results : Distribution of  homozygous wild-type TLR4 Asp299Gly polymorphism was higher in both of groups ( 94.5:97.2%.) and homozygous mutant allele was absent. There was not significantly difference of  TLR4 Asp299Gly in leptospirosis patients and healthy group ( ρ=1.00; OR 0.5; 95%CI, 0.04-5.6) and between mild and severe leptospirosis (ρ=0.54; OR 1.54 ; 95% CI, 1.20-1.98). The presence of homozygous wild-type TNF-α -308G/A polymorphism was higher between leptospirosis patients and healthy group (100:94.5%) andhomozygous mutant allele was not found in both of the groups. No significantly different of TNF-α -308G>A polymorphism between leptospirosis patient and healthy group (ρ=0.49).Conclusions : In this study, the polymorphisms of TLR4 Asp299Gly and TNF-α -308G/A were not associated with the susceptibility and severity of leptospirosis. Keywords : Leptospirosis, TLR4 Asp299Gly polymorphism, TNF-α -308G/A polymorphism

Association Study of Fecundity Gene BMPR1B with Prolificacy in Surti Goats under Farm and Field Condition

2019 ◽  
Vol 14 (04) ◽  
Author(s):  
N. S. Dangar ◽  
G. M. Pandya ◽  
U. V. Ramani ◽  
Y. D. Padheriya ◽  
T. Sangma ◽  
...  
Keyword(s):  
Litter Size ◽  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Pcr Primers ◽  
Dual Purpose ◽  
Protein Receptor ◽  
Morphogenetic Protein ◽  
Pcr Rflp ◽  
Base Size ◽  
Mutation Information

The Surti is a dual purpose goat breed of Gujarat. The bone morphogenetic protein receptor type 1B (BMPR1B) gene of transforming growth factor beta (TGF-β) superfamily ligands is playing a role in ovulation as well as litter size. Mutation in Exon-6 region of BMPR1B gene with base size 190 bp reported increasing litter size. Based on the known mutation information in goat and sheep, PCR primers were designed to screen polymorphism in total 100 Surti goats, 50 Surti goats from University Farm, Navsari and 50 Surti goats from field units of Southern part of Gujarat. During PCR-RFLP study no polymorphic sites were found for Exon-6 region of BMPR1B on Surti goats. Moreover, the twinning rate was 10% in first parity and higher in subsequent second (62.5%) and third (76.8%) parties.

scholarly journals Mutator-Suppressible Alleles of rough sheath1 and liguleless3 in Maize Reveal Multiple Mechanisms for Suppression

Genetics ◽  
2000 ◽  
Vol 154 (1) ◽  
pp. 437-446 ◽  
Author(s):  
Lisa Girard ◽  
Michael Freeling
Keyword(s):  
Untranslated Region ◽  
Mutant Allele ◽  
Negative Regulation ◽  
Wild Type ◽  
Knox Gene ◽  
Transcript Accumulation ◽  
Mu Suppression ◽  
Different Types ◽  
Rna Blot Analysis

Abstract Insertions of Mutator transposons into maize genes can generate suppressible alleles. Mu suppression is when, in the absence of Mu activity, the phenotype of a mutant allele reverts to that of its progenitor. Here we present the characterization of five dominant Mu-suppressible alleles of the knox (knotted1-like homeobox) genes liguleless3 and rough sheath1, which exhibit neomorphic phenotypes in the leaves. RNA blot analysis suggests that Mu suppression affects only the neomorphic aspect of the allele, not the wild-type aspect. Additionally, Mu suppression appears to be exerting its effects at the level of transcription or transcript accumulation. We show that truncated transcripts are produced by three alleles, implying a mechanism for Mu suppression of 5′ untranslated region insertion alleles distinct from that which has been described previously. Additionally, it is found that Mu suppression can be caused by at least three different types of Mutator elements. Evidence presented here suggests that whether an allele is suppressible or not may depend upon the site of insertion. We cite previous work on the knox gene kn1, and discuss our results in the context of interactions between Mu-encoded products and the inherently negative regulation of neomorphic liguleless3 and rough sheath1 transcription.

scholarly journals Differential Suppression of priA2::kan Phenotypes in Escherichia coli K-12 by Mutations in priA, lexA, and dnaC

Genetics ◽  
1996 ◽  
Vol 143 (1) ◽  
pp. 5-13 ◽  
Author(s):  
Steven J Sandler ◽  
Hardeep S Samra ◽  
Alvin J Clark
Keyword(s):  
Escherichia Coli ◽  
Dna Replication ◽  
Mutant Allele ◽  
Wild Type ◽  
Suppressor Mutations ◽  
Dnab Helicase ◽  
K 12 ◽  
Extragenic Suppressors ◽  
Assembly Activity

Abstract First identified as an essential component of the ϕX174 in vitro DNA replication system, PriA has ATPase, helicase, translocase, and primosome-assembly activities. priA1::kan strains of Escherichia coli are sensitive to UV irradiation, deficient in homologous recombination following transduction, and filamentous. priA2::kan strains have eightfold higher levels of uninduced SOS expression than wild type. We show that (1) priA1::kan strains have eightfold higher levels of uninduced SOS expression, (2) priA2::kan strains are UVS and Rec−, (3) lexA3 suppresses the high basal levels of SOS expression of a priA2::kan strain, and (4) plasmid-encoded priA300 (K230R), a mutant allele retaining only the primosome-assembly activity of priA+, restores both UVR and Rec+ phenotypes to a priA2::kan strain. Finally, we have isolated 17 independent UVR Rec+ revertants of priA2::kan strains that carry extragenic suppressors. All 17 map in the C-terminal half of the dnaC gene. DnaC loads the DnaB helicase onto DNA as a prelude for primosome assembly and DNA replication. We conclude that priA's primosome-assembly activity is essential for DNA repair and recombination and that the dnaC suppressor mutations allow these processes to occur in the absence of priA.

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