Characterization of Exon 3 of PROP1 gene and Screening of H173R polymorphism in Karan Fries bulls

2016 ◽  
Author(s):  
M. R. Vineeth ◽  
I. D. Gupta ◽  
Archana Verma ◽  
Ankit Magotra ◽  
Rakesh Kumar ◽  
...  
Keyword(s):  
Bos Taurus ◽  
Pcr Amplification ◽  
Multiple Alignment ◽  
Reference Sequence ◽  
Target Region ◽  
Region Sequence ◽  
Blast Analysis ◽  
Prop1 Gene ◽  
Karan Fries

Present study was done in thirty Karan Fries bulls to characterize the Exon 3 of PROP1 gene and to screen for the H173R polymorphism as well as other variations including reported and novel SNPs in the targeted region. The exon 3 was characterized by sequencing the amplicons obtained after PCR amplification using custom designed primers. The BLAST analysis of the obtained sequence yielded 100% and 99% homology with sequences of Bos taurus and bison respectively. The multiple alignment of the target region sequence with Bos taurus reference sequence revealed that the bulls under the study were free of H173R mutations. No variations were observed thus giving the targeted region a highly conserved one.

Sequence Characterization of Forebrain Embryonic Zinc Finger-like (FEZL) Gene in Indian Zebu (Bos indicus) Cattle and their Crossbreds

2021 ◽  
Author(s):  
S. Rajesh Kumar ◽  
I.D. Gupta ◽  
S. Goyal ◽  
Kathiravan Periasamy ◽  
A. Verma ◽  
...  
Keyword(s):  
Zinc Finger ◽  
Neuronal Development ◽  
Bos Taurus ◽  
Bos Indicus ◽  
Complete Sequence ◽  
Gene Markers ◽  
Sequence Characterization ◽  
Exon 1 ◽  
Karan Fries

Background: Forebrain embryonic zinc finger-like (FEZL) gene is an important candidate associated with mastitis resistance in dairy cattle. FEZL is involved in transcriptional regulation of neuronal development and there exists a crosstalk between neuronal development and immunity via downstream cytokine expression. A single glycine insertion into glycine stretch of FEZL gene has large effect on downstream cytokine pathway making the cows susceptible to mastitis. The present study was aimed to sequence characterize FEZL gene in Sahiwal (Bos indicus) and Karan Fries (Bos inidcus X Bos taurus) cattle.Methods: Sequence characterization of bovine FEZL gene was carried out by primer walking method. Ten sets of oligonucleotide primers were designed to synthesize overlapping fragments and generate the complete sequence of about 3.7 kb covering all exons and 5’ upstream regulatory and flanking regions.Result: A total of eight nucleotide variations including three INDELS and five substitution mutations were observed among FEZL gene sequences of Bos taurus, Bos indicus (Sahiwal) and Bos taurus X Bos indicus (Karan Fries) cattle. The conceptualized amino acid sequence of bovine FEZL gene in Sahiwal and Karan Fries cattle was found to have 13 tandem Glycine residues and a serine to proline change within exon 1 region. The percent identity of FEZL gene of Sahiwal and Karan Fries cattle was 99% with that of Bos taurus, 95% with dog, horse and pig, 94% with human, 93% with rabbit, 92% with marmoset, 89% with rat and 79% with chicken. Sequence characterization of ~0.7 kb 5’ flanking region showed that it is highly conserved among bovines and resulted in prediction of six putative sites for binding of transcription factors (including Elk-1, Oct-1, HNF4, Lmo2 complex, GATA-3 and Nkx2-5). Elucidation of Bos indicus FEZL gene will further form the basis to identify candidate gene markers for association with mastitis resistance/susceptibility in cattle.

Characterization of a cDNA for Rhesus Monkey Protein C Inhibitor – Evidence for N-Terminal Involvement in Heparin Stimulation

1995 ◽  
Vol 74 (04) ◽  
pp. 1079-1087 ◽  
Author(s):  
Klaus-P Radtke ◽  
José A Fernández ◽  
Bruno O Villoutreix ◽  
Judith S Greengard ◽  
John H Griffin
Keyword(s):  
Rhesus Monkey ◽  
Protein C ◽  
Activated Protein C ◽  
Pcr Amplification ◽  
Amino Acid Sequences ◽  
Heparin Binding ◽  
Reactive Center ◽  
Protein C Inhibitor ◽  
Polymerase Chain

SummarycDNAs for protein C inhibitor (PCI) were cloned from human and rhesus monkey 1 liver RNAs by reverse transcription and polymerase chain reaction (PCR) amplification. Sequencing showed that rhesus monkey and human PCI cDNAs were 93% identical. Predicted amino acid sequences differed at 26 of 387 residues. Pour of these differences (T352M, N359S, R362K, L3631) were in the reactive center loop that is important for inhibitory specificity, and two were in the N-terminal helix (M8T, E13K) that is implicated in glycosaminoglycan binding. PCI in human or rhesus monkey plasma showed comparable inhibitory activity towards human activated protein C in the presence of 10 U/ml heparin. However, maximal acceleration of the inhibition of activated protein C required 5-fold lower heparin concentration for rhesus monkey than for human plasma, consistent with the interpretation that the additional positive charge (E13K) in a putative-heparin binding region increased the affinity for heparin.

scholarly journals Isolation and characterization of two virulent Aeromonads associated with haemorrhagic septicaemia and tail-rot disease in farmed climbing perch Anabas testudineus

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Abhishek Mazumder ◽  
Hrishikesh Choudhury ◽  
Abhinit Dey ◽  
Dandadhar Sarma
Keyword(s):  
Natural Infection ◽  
Winter Season ◽  
Pcr Amplification ◽  
Body Parts ◽  
Anabas Testudineus ◽  
Climbing Perch ◽  
Isolation And Characterization ◽  
Biochemical Characterisation ◽  
Rot Disease

AbstractDiseased Anabas testudineus exhibiting signs of tail-rot and ulcerations on body were collected from a fish farm in Assam, India during the winter season (November 2018 to January 2019). Swabs from the infected body parts were streaked on sterilized nutrient agar. Two dominant bacterial colonies were obtained, which were then isolated and labelled as AM-31 and AM-05. Standard biochemical characterisation and 16S rRNA and rpoB gene sequencing identified AM-31 isolate as Aeromonas hydrophila and AM-05 as Aeromonas jandaei. Symptoms similar to that of natural infection were observed on re-infecting both bacteria to disease-free A. testudineus, which confirmed their virulence. LC50 was determined at 1.3 × 104 (A. hydrophila) and 2.5 × 104 (A. jandaei) CFU per fish in intraperitoneal injection. Further, PCR amplification of specific genes responsible for virulence (aerolysin and enterotoxin) confirmed pathogenicity of both bacteria. Histopathology of kidney and liver in the experimentally-infected fishes revealed haemorrhage, tubular degeneration and vacuolation. Antibiotic profiles were also assessed for both bacteria. To the best of our knowledge, the present work is a first report on the mortality of farmed climbing perch naturally-infected by A. hydrophila as well as A. jandaei, with no records of pathogenicity of the latter in this fish.

scholarly journals Characterization of the New Metallo-β-Lactamase VIM-13 and Its Integron-Borne Gene from a Pseudomonas aeruginosa Clinical Isolate in Spain

2008 ◽  
Vol 52 (10) ◽  
pp. 3589-3596 ◽  
Author(s):  
Carlos Juan ◽  
Alejandro Beceiro ◽  
Olivia Gutiérrez ◽  
Sebastián Albertí ◽  
Margalida Garau ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Pcr Amplification ◽  
Class 1 Integron ◽  
Negative Results ◽  
New Variant ◽  
Class B ◽  
Class 1 ◽  
Encoding Gene ◽  
Good Agreement

ABSTRACT During a survey conducted to evaluate the incidence of class B carbapenemase (metallo-β-lactamase [MBL])-producing Pseudomonas aeruginosa strains from hospitals in Majorca, Spain, five clinical isolates showed a positive Etest MBL screening test result. In one of them, strain PA-SL2, the presence of a new bla VIM derivative (bla VIM-13) was detected by PCR amplification with bla VIM-1-specific primers followed by sequencing. The bla VIM-13-producing isolate showed resistance to all β-lactams (except aztreonam), gentamicin, tobramycin, and ciprofloxacin. VIM-13 exhibited 93% and 88% amino acid sequence identities with VIM-1 and VIM-2, respectively. bla VIM-13 was cloned in parallel with bla VIM-1, and the resistance profile conferred was analyzed both in Escherichia coli and in P. aeruginosa backgrounds. Compared to VIM-1, VIM-13 conferred slightly higher levels of resistance to piperacillin and lower levels of resistance to ceftazidime and cefepime. VIM-13 and VIM-1 were purified in parallel as well, and their kinetic parameters were compared. The k cat/K m ratios for the antibiotics mentioned above were in good agreement with the MIC data. Furthermore, EDTA inhibited the activity of VIM-13 approximately 25 times less than it inhibited the activity of VIM-1. VIM-13 was harbored in a class 1 integron, along with a new variant (Ala108Thr) of the aminoglycoside-modifying enzyme encoding gene aacA4, which confers resistance to gentamicin and tobramycin. Finally, the VIM-13 integron was apparently located in the chromosome, since transformation and conjugation experiments consistently yielded negative results and the bla VIM-13 probe hybridized only with the genomic DNA.

scholarly journals Genomic characterization of 99 viruses from the bunyavirus families Nairoviridae, Peribunyaviridae, and Phenuiviridae, including 35 previously unsequenced viruses

2021 ◽  
Vol 17 (3) ◽  
pp. e1009315
Author(s):  
Marylee L. Kapuscinski ◽  
Nicholas A. Bergren ◽  
Brandy J. Russell ◽  
Justin S. Lee ◽  
Erin M. Borland ◽  
...  
Keyword(s):  
Plant Pathogens ◽  
Reference Sequence ◽  
Large Sample Size ◽  
Emerging Pathogens ◽  
Genome Sequences ◽  
Pathogenic Potential ◽  
Genomic Characterization ◽  
Response Decisions ◽  
Rapid Characterization

Bunyaviruses (Negarnaviricota: Bunyavirales) are a large and diverse group of viruses that include important human, veterinary, and plant pathogens. The rapid characterization of known and new emerging pathogens depends on the availability of comprehensive reference sequence databases that can be used to match unknowns, infer evolutionary and pathogenic potential, and make response decisions in an evidence-based manner. In this study, we determined the coding-complete genome sequences of 99 bunyaviruses in the Centers for Disease Control and Prevention’s Arbovirus Reference Collection, focusing on orthonairoviruses (family Nairoviridae), orthobunyaviruses (Peribunyaviridae), and phleboviruses (Phenuiviridae) that either completely or partially lacked genome sequences. These viruses had been collected over 66 years from 27 countries from vertebrates and arthropods representing 37 genera. Many of the viruses had been characterized serologically and through experimental infection of animals but were isolated in the pre-sequencing era. We took advantage of our unusually large sample size to systematically evaluate genomic characteristics of these viruses, including reassortment, and co-infection. We corroborated our findings using several independent molecular and virologic approaches, including Sanger sequencing of 197 genome segments, and plaque isolation of viruses from putative co-infected virus stocks. This study contributes to the described genetic diversity of bunyaviruses and will enhance the capacity to characterize emerging human pathogenic bunyaviruses.

scholarly journals Analysis of The Open Reading Frame (ORF) 29-TrnC (GCA) Sequence to Detect Indica and Japonica Sub Species on Upland Rice of Situ Bagendit and Inbred Rice of Ciherang

2018 ◽  
Vol 10 (1) ◽  
pp. 153-159
Author(s):  
Rohma Istiana ◽  
Hermin Pancasakti Kusumaningrum ◽  
Rejeki Siti Ferniah
Keyword(s):  
Plant Breeding ◽  
Upland Rice ◽  
Pcr Amplification ◽  
Breeding Program ◽  
Abiotic Factor ◽  
Reading Frame ◽  
Plant Breeding Program ◽  
Pcr Products

The identification and the characterization of genetic diversity of rice was the first step in the rice plant breeding program. This study aimed to detect indica or japonica sub-species on upland rice Situ Bagendit and inbred rice Ciherang using molecular markers ORF 29-TrnC (GCA) on the chloroplast genome. Rice was included to the indica sub-species if the 32 bp insertion on ORF 29-TrnC (GCA) sequence was found, on the contrary, if the deletion 32 bp on ORF 29-TrnC (GCA) was found then it was included to the japonica sub-species. DNA isolation was examined from the leaves of the rice plants, and then it tested quantitatively to determine the transparency and DNA concentration from the isolation results. PCR amplification was performed using a pair of primers CP2 and it was followed by agarose gel electrophoresis. The visualization of the DNA bands used the gel documentation. Sequencing of PCR products produced a long base 390 bp in Situ Bagendit rice and 390 bp in Ciherang rice. Analysis of the sequences showed that the insertions occurred throughout the 32 bp in Situ Bagendit rice and the insertions occurred throughout the 32 bp in Ciherang rice. The results showed that upland rice Situ Bagendit and inbred rice Ciherang were included in the indica sub-species. The knowledge of variety of genetics of rice can be used as bio-information in the plant breeding program. Further, the knowledge can be used to protect in genetic power source, the selection and the composing of superior varieties of rice which is tolerant with kinds of biotic and abiotic factor.

scholarly journals Characterization of a Putative Founder Mutation that Accounts for the High Incidence of Cystinosis in Brittany

2001 ◽  
Vol 12 (10) ◽  
pp. 2170-2174
Author(s):  
VASILIKI KALATZIS ◽  
STÉPHANIE CHERQUI ◽  
GENEVIÈVE JEAN ◽  
BÉATRICE CORDIER ◽  
PIERRE COCHAT ◽  
...  
Keyword(s):  
Autosomal Recessive ◽  
Founder Mutation ◽  
Pcr Amplification ◽  
Splice Site Mutation ◽  
Autosomal Recessive Disorder ◽  
Site Mutation ◽  
Live Births ◽  
Reverse Transcription Pcr ◽  
High Incidence

Abstract. Cystinosis is an autosomal recessive disorder, characterized by an accumulation of intralysosomal cystine, with an incidence of 1 in 100,000 to 200,000 live births. A higher incidence of cystinosis, 1 in 26,000 live births, has been reported in the western French province of Brittany. PCR amplification and sequencing has identified a 27-bp deletion starting 3 bp before the end of exon 8 and continuing into intron 8, 898-900+24del27, which has only been detected in families from this region. Reverse transcription—PCR amplification of RNA from an affected individual has shown that this mutation is indeed a splice-site mutation and results in the production of aberrant transcripts. These transcripts are predicted to either severely truncate cystinosin or alter its topology, thus accounting for the severe phenotype of these individuals. The mutation 898-900+24del27 has been identified in 7 of 18 alleles studied. This mutation is likely to be a founder mutation and would account for the higher incidence of cystinosis in Brittany.1

scholarly journals Identification and Evolutionary Characterization of Papillomavirus Sequences in New World Monkeys (Genera Sapajus and Alouatta) from Argentina

2021 ◽  
Author(s):  
Candelaria Sanchez Fernandez ◽  
Elisa M Bolatti ◽  
Andres C.A. Culasso ◽  
Diego Chouhy ◽  
Martin M Kowalewski ◽  
...  
Keyword(s):  
Pcr Amplification ◽  
Pcr Primers ◽  
Host Tree ◽  
New World Monkeys ◽  
Alouatta Caraya ◽  
Captive Populations ◽  
L1 Protein ◽  
Viral Sequences ◽  
Northeastern Argentina

Abstract Objective: In this study, we investigated the occurrence of papillomavirus (PV) infection in non-human primates (NHP, Platyrrhine) of northeastern Argentina by using broad-spectrum PCR primers at the L1 gene. In addition, we conducted a phylogenetic and coalescence analysis of viral sequences to explore their evolutionary history and evaluate the co-speciation hypothesis in the context of primate evolution. Methods: We obtained samples of 57 individuals from wild and captive populations of Alouatta caraya, Sapajus nigritus and Sapajus cay. We assessed PV infection by PCR amplification with the CUT primer system and sequencing of 337 bp (112 amino acids) of the L1 protein. The viral sequences were analyzed by phylogenetic and Bayesian coalescence methods to estimate the age of the most common recent ancestor (tMCRA) with BEAST, v1.4.8 software. We evaluated viral/host tree congruence with TreeMap v3.0. Results: We identified two novel putative PV sequences of the genus Gamma- PV in Sapajus sp and Alouatta caraya (SPV1 and AcPV1, respectively). The tMRCA of SPV1 was estimated at 11,941,682 years before present (ybp) and that of AcPV1 at 46,638,071 ybp, both predating the coalescence times of their hosts: 6.4 million years (MYA) and 6.8 MYA, respectively. Based on the comparison of primate and viral phylogenies, we could not reject the null hypothesis that the PV tree is no more congruent with the host tree than a random tree would be (P>0.05). Thus, a model of virus-host coevolution was rejected. Conclusion: This study presents the first report of PV infection in Platyrrhine species from Argentina, expands the range of described hosts for these viruses, and proposes new scenarios for their origin and dispersal.

Characterization of nematode mitochondrial genomes.

2021 ◽  
pp. 250-264
Author(s):  
Danny A. Humphreys-Pereira ◽  
Taeho Kim ◽  
Joong-Ki Park
Keyword(s):  
Polymerase Chain Reaction ◽  
Mitochondrial Genome ◽  
Genome Annotation ◽  
Pcr Amplification ◽  
Gene Identification ◽  
Mitochondrial Genomes ◽  
Pcr Cloning ◽  
Chain Reaction ◽  
Polymerase Chain

Abstract This chapter presents procedures on polymerase chain reaction (PCR) amplification, protocols for PCR, cloning and sequencing, and mitochondrial genome annotation and gene identification for the characterization of nematodes.

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