Detection of polymorphism of Calgranulin A gene in Indian Murrah Buffalo

2015 ◽  
Author(s):  
Sourabh Sulabh ◽  
Archana Verma ◽  
I. D. Gupta ◽  
S. Rajesh Kumar
Keyword(s):  
Immune System ◽  
Genetic Variants ◽  
Genomic Dna ◽  
Annealing Temperature ◽  
Rflp Analysis ◽  
Clinical Mastitis ◽  
Murrah Buffalo ◽  
Pcr Rflp ◽  
Pcr Products ◽  
Calgranulin A

Calgranulin A (S100A8) gene is one of the important candidate genes, which affects the host disease resistance by enhancing the immune system. Present study was undertaken with the objectives to identify polymorphism in Calgranulin A gene and to associate identified genetic variants with the incidence of clinical mastitis in Murrah buffalo. Genomic DNA was isolated from 100 randomly selected lactating Murrah. Two sets of primers were designed to amplify targeted regions of the gene. PCR products were obtained at annealing temperature of 63.8oC and were of 449 and 489 bp for respective primer set. PCR-RFLP analysis was carried out using HinfI for contig I and AluI and MboII restriction endonucleases for contig II. Both the contigs revealed monomorphism with frequency of the only prevailing A allele as 1.00. It was not feasible to analyze association with the incidence of mastitis, as no genetic variants were observed in the animals studied.

Polymorphism of the exon 3 of leptin gene in Malpura sheep

2016 ◽  
Author(s):  
A.S. Meena ◽  
A. Sahoo ◽  
R. S. Bhatt ◽  
A.S. Meena
Keyword(s):  
Genetic Variants ◽  
Genomic Dna ◽  
Extraction Method ◽  
Allelic Variation ◽  
Restriction Enzymes ◽  
Allelic Frequency ◽  
Adipose Tissues ◽  
Allelic Variants ◽  
Pcr Rflp ◽  
Pcr Products

Leptin (LEP) is primarily expressed in the adipose tissues. It regulates the feed intake, energy metabolism and body composition and plays a crucial role in regulating body weight and growth in mammals. The aim of the present study was to find out an allelic variation in leptin gene of Malpura sheep. A total of 112 Malpura sheep were selected and the genomic DNA was isolated by phenol-chloroform extraction method. PCR was carried out in order to amplify the 471 bp fragment of the exon 3 coding sequence of the leptin gene. The genotyping was done by PCR-RFLP technique. PCR products were digested with three (BcnI, SsiI and OliI) restriction enzymes. For detection of allelic variants, three non-synonymous SNPs were found in Malpura sheep. The A271G and A316C loci were found mono-morphic, while T387G locus was found polymorphic. Two genetic variants (G and T) and three genotypes (GG, GT and TT) were found in Malpura sheep. The allelic frequency of G and T allele at T387G locus was found 0.82 and 0.18, respectively.

scholarly journals Molecular Characterization of Pathogenicity Locus (PaLoc) and tcdC Genetic Diversity Among tcdA+B+ Clostridioides Difficile Clinical Isolates in Tehran, Iran

2020 ◽  
Author(s):  
Mansoor Kodori ◽  
Zohreh Ghalavand ◽  
Abbas Yadegar ◽  
Gita Eslami ◽  
Masoumeh Azimirad ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
Molecular Characterization ◽  
Disease Severity ◽  
Genetic Variants ◽  
Rflp Analysis ◽  
Clostridioides Difficile ◽  
Pcr Rflp ◽  
The Common ◽  
Healthcare Associated

Abstract Background: Clostridioides difficile is the main cause of healthcare-associated diarrhea worldwide. It is proposed that certain C. difficile toxinotypes with distinct pathogenicity locus (PaLoc) variants are associated with disease severity and outcomes. Additionally, few studies have described the common C. difficile toxinotypes, and also little is known about the tcdC variants in Iranian isolates. We characterized the toxinotypes and the tcdC genotypes from a collection of Iranian clinical C. difficile tcdA+B+ isolates with known ribotypes (RTs).Methods: Fifty C. difficile isolates with known RTs and carrying the tcdA and tcdB toxin genes were analyzed. Toxinotyping was carried out based on a PCR-RFLP analysis of a 19.6 kb region encompassing the PaLoc. Genetic diversity of the tcdC gene was determined by the sequencing of the gene.Results: Of the 50 C. difficile isolates investigated, five distinct toxinotypes were recognized. Toxinotypes 0 (33/50, 66%) and V (11/50, 22%) were the most frequently found. C. difficile isolates of the toxinotype 0 mostly belonged to RT 001 (12/33, 36.4%), whereas toxinotype V consisted of RT 126 (9/11, 81.8%). The tcdC sequencing showed six variants (35/50, 70%); tcdC-sc3 (24%), tcdC-A (22%), tcdC-sc9 (18%), tcdC-B (2%), tcdC-sc14 (2%), and tcdC-sc15 (2%). The remaining isolates were wild-types (15/50, 30%) in the tcdC gene.Conclusions: The present study demonstrates that the majority of clinical tcdA+B+ isolates of C. difficile frequently harbor tcdC genetic variants. We also found that the RT 001/ toxinotype 0 and the RT 126/ toxinotype V are the most common types among Iranian isolates. Further studies are needed to investigate the putative association of various tcdC genotypes with CDI severity and its recurrence.

scholarly journals Phylogenetic Relationships of Local Durian Species based on Morphological Characteristics and PCR-RFLP Analysis of the Ribosomal Internal Transcribed Spacer (ITS) DNA

2016 ◽  
Vol 8 (3) ◽  
pp. 362
Author(s):  
Fidia Fibriana ◽  
Lutfia Nur Hadiyanti
Keyword(s):  
Phylogenetic Relationships ◽  
Internal Transcribed Spacer ◽  
Morphological Characteristics ◽  
Biology Education ◽  
Rflp Analysis ◽  
Restriction Pattern ◽  
Internal Transcribed Spacers ◽  
Central Java ◽  
Pcr Rflp ◽  
Pcr Products

<p>In this study, twenty local durian accessions obtained from Central Java in situ collection were characterized using the morphological characteristics and the restriction patterns which generated from the region spanning the internal transcribed spacers ITS LEU and ITS 4. Morphological characteristics of durian leaf, stem, tree, and fruit showed variations for the different accessions, whereas polymerase chain reaction (PCR) products of ribosomal DNA region showed a low length of variation. The size of the PCR products and the restriction analyses with the restriction endonucleases Bsp1431yielded a restriction pattern for each accessions. The results of this study can be utilized by local durian farmers as a preliminary reference for durian propagation. The data obtained need to be supported by further research using the other molecular markers to obtain more accurate data. The clear identity of durian species can help the management of propagation systems by farmers to get superior local durian.</p><p><strong>How to Cite</strong></p><p>Fibriana, F., &amp; Hadiyanti, L. N. (2016). Phylogenetic Relationships of Local Durian Species based on Morphological Characteristics and PCR-RFLP Analysis of the Ribosomal Internal Transcribed Spacer (ITS) DNA. <em>Biosaintifika: Journal of Biology &amp; Biology Education</em>, 8(3), 362-370. </p>

scholarly journals Analysis of Slovak Spotted breed for bovine beta casein A1 variant as risk factor for human health.

1970 ◽  
Vol 60 (4) ◽  
Author(s):  
Martina Miluchová ◽  
Michal Gábor ◽  
Anna Trakovická
Keyword(s):  
Diabetes Mellitus ◽  
Genomic Dna ◽  
Total Population ◽  
Human Consumption ◽  
Effective Number ◽  
Immune Systems ◽  
Pcr Rflp ◽  
Pcr Products ◽  
Beta Casein ◽  
Commercial Kit

The goal of work was identification A1 variant of bovine beta casein which involves ischemic heart disease and diabetes mellitus in human. The digestion of A1beta casein can result in the production of bioactive beta casomorphin-7 (BCM-7); this is not the case with A2. This bioactive peptide has been linked to physiological traits that may elicit effects on components of the vascular and immune systems. The material involved 111 Slovak Spotted breed. Bovine genomic DNA was extracted from whole blood by using commercial kit, and used in order to estimate beta-casein genotypes by means of PCR-RFLP method. The PCR products were digested with DdeI restriction enzyme. In the population included in the study were detected all three genotypes, homozygote genotype A1A1 (14 animals), heterozygote genotype A1A2 (37 animals) and homozygote genotype A2A2 (60 animals). In the total population of cattle homozygotes A2A2-0.5405 were the most frequent, while homozygotes A1A1-0.1261 were the least frequent ones. This suggests a superiority of allele A2 (0.7072) which does not produce BCM-7, and thus is safe for human consumption. The expected homozygosity for gene CSN2 is in the population stated a slight increase in homozygosity (0.5858). This caused a slight decrease in the level of possible variability realization (41.80%), which corresponds to the effective number of alleles (1.7071).

Association of polymorphism at intron 2 of FABP3 Gene with milk production traits in Sahiwal and Karan Fries cattle

2018 ◽  
Author(s):  
Alok Kumar Yadav ◽  
Anupama Mukherjee ◽  
Suchit Kumar
Keyword(s):  
Milk Production ◽  
Rflp Analysis ◽  
Production Traits ◽  
Milk Production Traits ◽  
Pcr Rflp ◽  
Sahiwal Cattle ◽  
Pcr Products ◽  
The Mean ◽  
Intron 2 ◽  
Karan Fries

PCR-RFLP analysis of PCR products were carried out using Aci I / SSi I for 100 Sahiwal and 115 Karan Fries cattle. In Sahiwal cattle and Karan Fries cattle, 438bp has three genotypes AA (438), AB (438+299+139 bp) and BB (299+139 bp). In Sahiwal cattle these genotypes are highly significant for FL305DPY but in Karan Fries cattle these genotypes are highly significant for FL305DMY, FLTMY, FL305DFY and FL305DPY. In Sahiwal cattle, mean ± SE of AA genotype for FL305DMY, FLTMY, FL305DFY, FL305DSNFY, FL305DPY were 1809.90 ± 15.7 kg, 2029.4 ± 15.6 kg, 99.90 ± 0.66 kg, 154.87 ± 0.17 kg and 44.81 ± 0.06 kg, respectively and for AB genotype were 1800.76 ± 9.48 kg, 1993.99 ± 9.42 kg, 100.54 ± 0.39 kg, 154.79 ± 0.10 kg, 43.99 ± 0.04 kg, respectively and for BB genotype were 1830.0 ± 14.10 kg, 2032.80 ± 14.0 kg, 100.24 ± 0.59 kg, 155.11 ± 0.15 kg, 42.98 ± 0.05 kg, respectively. Heterozygous AB genotype was found to be superior for, FL305DFY trait. AA genotype was significantly superior for FL305DPY traits whereas BB genotype was found to be superior for FL305DMY, FLTMY, FL305DSNFY. In Karan Fries cattle, the mean ± SE of AA genotype for FL305DMY, FLTMY, FL305DFY, FL305DSNFY, FL305DPY were 3442.17 ± 8.39 kg, 4461.93 ± 8.39 kg, 124.96 ± 7.20 kg, 277.35 ± 0.08 kg and 112.51 ± 0.08 kg, respectively and for AB genotype were 3572.69 ± 5.93 kg, 4592.45 ± 5.93 kg, 140.17 ± 5.09 kg, 278.60 ± 0.06 kg, 113.91 ± 0.05 kg, respectively and for BB genotypes were 3502.41 ± 9.19 kg, 4522.17 ± 9.19 kg, 136.91 ± 7.89 kg, 277.93 ± 0.09 kg, 113.19 ± 0.08 kg, respectively.

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 Molecular Analysis of Vitex Species Using Candidate DNA Barcoding and PCR-RFLP of the matK Gene for Authentication of Vitex glabrata

2013 ◽  
Vol 8 (1) ◽  
pp. 1934578X1300800 ◽  
Author(s):  
Waranyoo Phoolcharoen ◽  
Suchada Sukrong
Keyword(s):  
Folk Medicine ◽  
Intergenic Spacer ◽  
Rflp Analysis ◽  
Size Difference ◽  
Dna Barcodes ◽  
Pcr Rflp ◽  
Matk Gene ◽  
Pcr Products ◽  
The Difference ◽  
Vitex Species

Several species of the Vitex genus, family Lamiaceae, are used in folk medicine for a variety of remedies. V. glabrata is unique among Vitex species because its main effect is sexual enhancement. However, crude drugs derived from different Vitex species might not be easily distinguishable, which could lead to their misidentification and misuse. Therefore, the accurate authentication of V. glabrata is critical for its effective medicinal use. In this study, the mat K gene and the psbA- trnH intergenic spacer candidate DNA barcodes were sequenced and analyzed to identify five different Vitex species that are medicinally used in Thailand: V. negundo, V. trifolia, V. rotundifolia, V. limonifolia, and V. glabrata. Each region was successfully amplified from the leaves of the five species using a single set of primers, and the sequences determined. The size difference in PCR products of psb A- trn H and PCR restriction fragment length polymorphism (PCR-RFLP) of the matK gene sequences were used to differentiate V. glabrata from other Vitex species. These results indicate both the matK gene and the psbA-trnH intergenic spacer as candidate DNA barcodes of Vitex species and suggest that the difference of psbA-trnH PCR products and PCR-RFLP analysis based on the mat K gene are effective for the authentication of V. glabrata.

scholarly journals Determination of Echinococcus granulosus genotypes in livestock slaughtered in Shush County, Southwest Iran using PCR-RFLP

2019 ◽  
Vol 56 (3) ◽  
pp. 196-201 ◽  
Author(s):  
S. Fallahizadeh ◽  
R. Arjmand ◽  
A. Jelowdar ◽  
A. Rafiei ◽  
F. Kazemi
Keyword(s):  
Rflp Analysis ◽  
Sensu Stricto ◽  
Enzymatic Reactions ◽  
Hydatid Cysts ◽  
Larval Stages ◽  
Pcr Rflp ◽  
Digestion Pattern ◽  
Pcr Products ◽  
Southwestern Iran ◽  
Southwest Iran

SummaryEchinococosis is a zoonotic disease caused by the larval stages of Echinococcus spp. that occurs in most parts of the world. Herein, we aimed to evaluate the genotypes of isolated hydatid cysts from slaughtered animals in Shush county, southwestern Iran. Totally, 96 hydatid cysts were collected, including 11 buffaloes, 13 cattle, 12 goat and 60 sheep. The PCR was done by a primer pair (BDI and 4s) to amplify ITS1 fragment. Four restriction endonucleases including AluI, HpaII, RsaI, and TaqI were used for RFLP products and enzymatic reactions were electrophoresed. Finally, twenty PCR products were sent for sequencing and phylogenetic tree was drawn with MEGA6. Molecular identification of 96 hydatid cysts demonstrated a distinctive 1000 bp fragment in all samples from four animal hosts. RFLP analysis showed similar digestion patterns in all samples. AluI digestion yielded 800 bp and 200 bp fragments, HpaII digestion made 700 bp and 300 bp fragments and RsaI digestion entailed 655 and 345segments. Moreover, TaqI rendered no digestion pattern on rDNA-ITS1 region. Additionally, E. granulosus sensu stricto (G1-3 complex) was the prevailing genotype in all livestock samples, according to PCR-RFLP and sequencing analyses.

scholarly journals Molecular-Phylogenetic Research of the Genus Hypericum L. in Flora of Azerbaijan

2021 ◽  
Vol 7 (11) ◽  
pp. 22-27
Keyword(s):  
Experimental Study ◽  
Chloroplast Dna ◽  
Genomic Dna ◽  
Native Species ◽  
Leaf Tissue ◽  
Rflp Analysis ◽  
Flowering Plant ◽  
The Family ◽  
Pcr Rflp ◽  
Molecular Phylogenetic

Hypericum is one of the 100 largest flowering plant genera forming the family Hypericaceae Juss., which belongs to the clusioid clade of the Malpighiales. Hypericum is represented in Azerbaijan flora by 19 native species and 1 subspecies belonging to 7 taxonomic sections. The chloroplast DNA of 8 species from the genus was studied by PCR-RFLP analysis. Total genomic DNA was extracted from leaf tissue using the DNeasyPlantMini kit. (Qiagen Inc.; Valencia, CA, USA) following the supplied protocol and quanti field using a Nanodrop (Nanodrop Technologies; Wilmington, DE, USA) spectrophotometer. The article is part of an experimental study that comprises molecular-phylogenetic research of this genus in the flora of Azerbaijan.

scholarly journals Detection and Resolution of Cryptosporidium Species and Species Mixtures by Genus-Specific Nested PCR-Restriction Fragment Length Polymorphism Analysis, Direct Sequencing, and Cloning

2011 ◽  
Vol 77 (12) ◽  
pp. 3998-4007 ◽  
Author(s):  
Norma J. Ruecker ◽  
Rebecca M. Hoffman ◽  
Rachel M. Chalmers ◽  
Norman F. Neumann
Keyword(s):  
Restriction Fragment Length Polymorphism ◽  
Length Polymorphism ◽  
Fragment Length ◽  
Nested Pcr ◽  
Fragment Length Polymorphism ◽  
Rflp Analysis ◽  
Content Type ◽  
Pcr Rflp ◽  
Pcr Products ◽  
Restriction Fragment Length

ABSTRACTMolecular methods incorporating nested PCR-restriction fragment length polymorphism (RFLP) analysis of the 18S rRNA gene ofCryptosporidiumspecies were validated to assess performance based on limit of detection (LoD) and for detecting and resolving mixtures of species and genotypes within a single sample. The 95% LoD was determined for seven species (Cryptosporidium hominis,C. parvum,C. felis,C. meleagridis,C. ubiquitum,C. muris, andC. andersoni) and ranged from 7 to 11 plasmid template copies with overlapping 95% confidence limits. The LoD values for genomic DNA from oocysts on microscope slides were 7 and 10 template copies forC. andersoniandC. parvum, respectively. The repetitive nested PCR-RFLP slide protocol had an LoD of 4 oocysts per slide. When templates of two species were mixed in equal ratios in the nested PCR-RFLP reaction mixture, there was no amplification bias toward one species over another. At high ratios of template mixtures (>1:10), there was a reduction or loss of detection of the less abundant species by RFLP analysis, most likely due to heteroduplex formation in the later cycles of the PCR. Replicate nested PCR was successful at resolving many mixtures ofCryptosporidiumat template concentrations near or below the LoD. The cloning of nested PCR products resulted in 17% of the cloned sequences being recombinants of the two original templates. Limiting-dilution nested PCR followed by the sequencing of PCR products resulted in no sequence anomalies, suggesting that this method is an effective and accurate way to study the species diversity ofCryptosporidium, particularly for environmental water samples, in which mixtures of parasites are common.

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