scholarly journals Serotype-Specific Identification of Polioviruses by PCR Using Primers Containing Mixed-Base or Deoxyinosine Residues at Positions of Codon Degeneracy

1998 ◽  
Vol 36 (2) ◽  
pp. 352-357 ◽  
Author(s):  
David R. Kilpatrick ◽  
Baldev Nottay ◽  
Chen-Fu Yang ◽  
Su-Ju Yang ◽  
Edson Da Silva ◽  
...  
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequences ◽  
High Rate ◽  
Rapid Screening ◽  
Reaction Conditions ◽  
Specific Primers ◽  
Specific Identification ◽  
Primer Sets ◽  
Reference Strains ◽  
Codon Degeneracy

We have developed a method for determining the serotypes of poliovirus isolates by PCR. Three sets of serotype-specific antisense PCR-initiating primers (primers seroPV1A, seroPV2A, and seroPV3A) were designed to pair with codons of VP1 amino acid sequences that are conserved within but that differ across serotypes. The sense polarity primers (primers seroPV1S, seroPV2S, and seroPV3S) matched codons of more conserved capsid sequences. The primers contain mixed-base and deoxyinosine residues to compensate for the high rate of degeneracy of the targeted codons. The serotypes of all polioviruses tested (48 vaccine-related isolates and 110 diverse wild isolates) were correctly identified by PCR with the serotype-specific primers. None of the genomic sequences of 49 nonpolio enterovirus reference strains were amplified under equivalent reaction conditions with any of the three primer sets. These primers are useful for the rapid screening of poliovirus isolates and for determining the compositions of cultures containing mixtures of poliovirus serotypes.

An efficient and eco-friendly method for the thiol-Michael addition in aqueous solutions using amino acid ionic liquids (AAILs) as organocatalysts

2020 ◽  
Vol 92 (1) ◽  
pp. 97-106
Author(s):  
Roberto Figueroa Guíñez ◽  
José G. Santos ◽  
Ricardo A. Tapia ◽  
Jackson J. Alcazar ◽  
Margarita E. Aliaga ◽  
...  
Keyword(s):  
Aqueous Solution ◽  
Ionic Liquids ◽  
Amino Acid ◽  
Efficient Method ◽  
Michael Addition ◽  
Rate Constants ◽  
Michael Reaction ◽  
High Rate ◽  
Reusable Catalyst ◽  
Reaction Conditions

AbstractA series of amino-acid based ionic liquids (Bmim[AA]s) have been synthesized and evaluated as catalysts, in aqueous solution. The results of a kinetic study of the thiol-Michael reaction of L-Cysteine with trans-β-nitrostyrene demonstrated the advantages of using (Bmim[AA]s) as organocatalysts. The benefits include high rate constants; mild reaction conditions; and, a reusable catalyst, which leads to a simple and efficient method for these important kinds of reactions.

scholarly journals Vascular endothelial growth factors encoded by Orf virus show surprising sequence variation but have a conserved, functionally relevant structure

2002 ◽  
Vol 83 (11) ◽  
pp. 2845-2855 ◽  
Author(s):  
A. A. Mercer ◽  
L. M. Wise ◽  
A. Scagliarini ◽  
C. J. McInnes ◽  
M. Büttner ◽  
...  
Keyword(s):  
Amino Acid ◽  
Sequence Variation ◽  
Amino Acid Identity ◽  
Amino Acid Sequences ◽  
High Rate ◽  
Orf Virus ◽  
Vegf Receptor ◽  
Vascular Endothelial ◽  
Vegf Gene ◽  
X Ray Crystallography

The first report of a vascular endothelial growth factor (VEGF)-like gene in Orf virus included the surprising observation that the genes from two isolates (NZ2 and NZ7) shared only 41·1% amino acid sequence identity. We have examined this sequence disparity by determining the VEGF gene sequence of 21 isolates of Orf virus derived from diverse sources. Most isolates carried NZ2-like VEGF genes but their predicted amino acid sequences varied by up to 30·8% with an average amino acid identity between pairs of NZ2-like sequences of 86·1%. This high rate of sequence variation is more similar to interspecies than intraspecies variability. In contrast, only three isolates carried an NZ7-like VEGF gene and these varied from the NZ7 sequence by no more than a single nucleotide. The VEGF family are ligands for a set of tyrosine kinase receptors. The viral VEGFs are unique among the family in that they recognize VEGF receptor 2 (VEGFR-2) but not VEGFR-1 or VEGFR-3. Comparisons of the viral VEGFs with other family members revealed some correlations between conserved residues and the ability to recognize specific VEGF receptors. Despite the sequence variations, structural predictions for the viral VEGFs were very similar to each other and to the structure determined by X-ray crystallography for human VEGF-A. Structural modelling also revealed that a groove seen in the VEGF-A homodimer and believed to play a role in its binding to VEGFR-1 is blocked in the viral VEGFs. This may contribute to the inability of the viral VEGFs to bind VEGFR-1.

scholarly journals Comparação da seqüência de nucleotídeos do gene da proteína capsidial de estirpes severas e premunizantes do Papaya ringspot virus

2003 ◽  
Vol 28 (6) ◽  
pp. 678-681 ◽  
Author(s):  
Marilia G. S. Della Vecchia ◽  
Luis E. A. Camargo ◽  
Jorge A. M. Rezende
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Core Region ◽  
Amino Acid Sequences ◽  
Ringspot Virus ◽  
Papaya Ringspot Virus ◽  
Specific Primers ◽  
Sequence Comparisons ◽  
The Core ◽  
Cp Gene

This study compared three mild and three severe strains of Papaya ringspot virus - type W (PRSV-W), based on nucleotide and amino acid sequences of the capsid protein (CP) gene. The CP nucleotide sequences of the mild strains shared 98% to 100% identity. When compared to the severe strains the identity ranged from 93% to 95%, except in the case of PRSV-W-2R, which resulted from reversion of the mild strains PRSV-W-2. The CP sequence of the reverting strain showed 100% identity with the sequence of its parental strain. An insertion of six nucleotides in the core region of the CP gene, which reflected the addition of two amino acids (Asn and Asp) in the deduced sequence of the protein, was found in all mild strains. These sequence comparisons were used to design strain-specific primers that were used to specifically amplify regions for either the mild or severe strains.

scholarly journals Genomic organization of three novel toxins from the scorpion Buthus martensi Karsch that are active on potassium channels

2000 ◽  
Vol 346 (3) ◽  
pp. 805-809 ◽  
Author(s):  
Li DAI ◽  
Jing-Jiang WU ◽  
Yi-Hua GU ◽  
Zheng-Dao LAN ◽  
Min-Hua LING ◽  
...  
Keyword(s):  
Amino Acid ◽  
Genomic Dna ◽  
Amino Acid Sequences ◽  
Signal Peptides ◽  
Amino Acid Residues ◽  
Specific Primers ◽  
Cdna Sequences ◽  
C Terminus ◽  
Partial Cdna ◽  
Rapid Amplification

The cDNA and genomic DNA of three novel toxins from the scorpion Buthus martensi Karsch that are active on K+ channels, designated BmKTX (where KTX is kaliotoxin), BmTX1 and BmTX2, were cloned and sequenced. On the basis of their known amino acid sequences, gene-specific primers for 3ʹ and 5ʹ rapid amplification of cDNA ends (RACE) were designed and synthesized. By overlapping the two partial cDNA sequences obtained by 3ʹ and 5ʹ RACE, their full-length cDNA sequences were completed. BmKTX encodes a signal peptide of 22 amino acid residues and a mature toxin of 38 residues, whereas BmTX1 and BmTX2 encode signal peptides of 20 and 21 residues respectively and a mature toxin of 38 residues for each. Their cDNA-deduced amino acid sequences were totally consistent with those determined except that the C-terminus of BmKTX had an additional Gly residue, which was removed during post-translational processing and was indispensable for the amidation of its C-terminal Lys residue. In addition, the first deduced amino acid for both BmTX1 and BmTX2 is Gln instead of pyro-Glu in the reported toxins, which obviously also undergoes post-translational processing. The genomic DNA species of these three toxins were also amplified by PCR, then cloned and sequenced. They all consisted of two exons disrupted by a small single intron. All of these introns were inserted within the signal peptides at position -6 for BmKTX and at position -5 for both BmTX1 and BmTX2 upstream of the mature toxins, and consisted of 87, 87 and 80 bp respectively.

scholarly journals Molecular Characterization of Chromosomal Class C β-Lactamase and Its Regulatory Gene in Ochrobactrum anthropi

2001 ◽  
Vol 45 (8) ◽  
pp. 2324-2330 ◽  
Author(s):  
David Nadjar ◽  
Roger Labia ◽  
Claude Cerceau ◽  
Chantal Bizet ◽  
Alain Philippon ◽  
...  
Keyword(s):  
Amino Acid ◽  
Regulatory Gene ◽  
Amino Acid Sequences ◽  
Clinical Strain ◽  
Kinetic Properties ◽  
Ochrobactrum Anthropi ◽  
Amino Terminal ◽  
Class C ◽  
Human Infections ◽  
Reference Strains

ABSTRACT Ochrobactrum anthropi, formerly known as CDC group Vd, is an oxidase-producing, gram-negative, obligately aerobic, non-lactose-fermenting bacillus of low virulence that occasionally causes human infections. It is highly resistant to all β-lactams except imipenem. A clinical isolate, SLO74, and six reference strains were tested. MICs of penicillins, aztreonam, and most cephalosporins tested, including cefotaxime and ceftazidime, were >128 μg/ml and of cefepime were 64 to >128 μg/ml. Clavulanic acid was ineffective and tazobactam had a weak effect in association with piperacillin. Two genes, ampR and ampC, were cloned by inserting restriction fragments of genomic DNA from the clinical strain O. anthropi SLO74 into pBK-CMV to give the recombinant plasmid pBK-OA1. The pattern of resistance to β-lactams of this clone was similar to that of the parental strain, except for its resistance to cefepime (MIC, 0.5 μg/ml). The deduced amino acid sequence of the AmpC β-lactamase (pI, 8.9) was only 41 to 52% identical to the sequence of other chromosomally encoded and plasmid-encoded class C β-lactamases. The kinetic properties of this β-lactamase were typical for this class of β-lactamases. Upstream from the ampC gene, the ampR gene encodes a protein with a sequence that is 46 to 62% identical to those of other AmpR proteins and with an amino-terminal DNA-binding domain typical of transcriptional activators of the Lys-R family. The deduced amino acid sequences of theampC genes of the six reference strains were 96 to 99% identical to the sequence of the clinical strain. The β-lactamase characterized from strain SLO74 was named OCH-1 (gene, bla OCH-I).

scholarly journals The Evolutionary History of Daphniid α-Carbonic Anhydrase within Animalia

2015 ◽  
Vol 2015 ◽  
pp. 1-11 ◽  
Author(s):  
Billy W. Culver ◽  
Philip K. Morton
Keyword(s):  
Amino Acid ◽  
Evolutionary History ◽  
Homo Sapiens ◽  
Aquatic Organism ◽  
Amino Acid Sequences ◽  
High Rate ◽  
Ancestral State ◽  
Carbonic Anhydrases ◽  
Acid Base ◽  
History Of

Understanding the mechanisms that drive acid-base regulation in organisms is important, especially for organisms in aquatic habitats that experience rapidly fluctuating pH conditions. Previous studies have shown that carbonic anhydrases (CAs), a family of zinc metalloenzymes, are responsible for acid-base regulation in many organisms. Through the use of phylogenetic tools, this present study attempts to elucidate the evolutionary history of the α-CA superfamily, with particular interest in the emerging model aquatic organism Daphnia pulex. We provide one of the most extensive phylogenies of the evolution of α-CAs, with the inclusion of 261 amino acid sequences across taxa ranging from Cnidarians to Homo sapiens. While the phylogeny supports most of our previous understanding on the relationship of how α-CAs have evolved, we find that, contrary to expectations, amino acid conservation with bacterial α-CAs supports the supposition that extracellular α-CAs are the ancestral state of animal α-CAs. Furthermore, we show that two cytosolic and one GPI-anchored α-CA in Daphnia genus have homologs in sister taxa that are possible candidate genes to study for acid-base regulation. In addition, we provide further support for previous findings of a high rate of gene duplication within Daphnia genus, as compared with other organisms.

scholarly journals Differentiation of Borrelia burgdorferiSensu Lato on the Basis of RNA Polymerase Gene (rpoB) Sequences

2000 ◽  
Vol 38 (7) ◽  
pp. 2557-2562 ◽  
Author(s):  
Seung-Hyun Lee ◽  
Bum-Joon Kim ◽  
Jong-Hyun Kim ◽  
Kyung-Hee Park ◽  
Seo-Jeong Kim ◽  
...  
Keyword(s):  
Amino Acid ◽  
Restriction Analysis ◽  
Direct Sequencing ◽  
Amino Acid Sequences ◽  
Amino Acid Residues ◽  
Borrelia Lusitaniae ◽  
Primer Sets ◽  
Species Specific ◽  
Identification And Characterization

We determined the nucleotide sequences (329 bp) of therpoB DNAs from 22 reference strains ofBorrelia. No insertions or deletions were observed. Deduced amino acid sequences of amplified rpoB DNA comprised 109 amino acid residues (N450 to M558[Escherichia coli numbering]). All amino acid sequences were identical with the exception of those of Borrelia lusitaniae PotiB2 (T461→A) and B. bissettii DN127 (I498→V). Each species of B. burgdorferi sensu lato was differentiated as a distinct entity in the phylogenetic tree constructed by a neighbor-joining method. B. burgdorferi sensu lato could be distinguished from B. turicatae and B. hermsii, which are associated with relapsing fever. Seventeen Korean isolates could be identified by PCR-linked direct sequencing and restriction analysis of the rpoB DNA. These results suggest that rpoB DNA is useful for identification and characterization of Borrelia. In addition, we developed the rapid species identification method using the species-specific primer sets based on rpoB gene sequences.

Diversity of Primary Structures of the Carboxy-terminal Regions of Mammalian Fibrinogen Aα-Chains

1993 ◽  
Vol 69 (04) ◽  
pp. 351-360 ◽  
Author(s):  
Masahiro Murakawa ◽  
Takashi Okamura ◽  
Takumi Kamura ◽  
Tsunefumi Shibuya ◽  
Mine Harada ◽  
...  
Keyword(s):  
Amino Acid ◽  
Rhesus Monkey ◽  
Syrian Hamster ◽  
Tandem Repeats ◽  
Mammalian Species ◽  
Amino Acid Sequences ◽  
Point Of View ◽  
Cross Linking ◽  
Amino Terminal ◽  
Rgd Sequence

SummaryThe partial amino acid sequences of fibrinogen Aα-chains from five mammalian species have been inferred by means of the polymerase chain reaction (PCR). From the genomic DNA of the rhesus monkey, pig, dog, mouse and Syrian hamster, the DNA fragments coding for α-C domains in the Aα-chains were amplified and sequenced. In all species examined, four cysteine residues were always conserved at the homologous positions. The carboxy- and amino-terminal portions of the α-C domains showed a considerable homology among the species. However, the sizes of the middle portions, which corresponded to the internal repeat structures, showed an apparent variability because of several insertions and/or deletions. In the rhesus monkey, pig, mouse and Syrian hamster, 13 amino acid tandem repeats fundamentally similar to those in humans and the rat were identified. In the dog, however, tandem repeats were found to consist of 18 amino acids, suggesting an independent multiplication of the canine repeats. The sites of the α-chain cross-linking acceptor and α2-plasmin inhibitor cross-linking donor were not always evolutionally conserved. The arginyl-glycyl-aspartic acid (RGD) sequence was not found in the amplified region of either the rhesus monkey or the pig. In the canine α-C domain, two RGD sequences were identified at the homologous positions to both rat and human RGD S. In the Syrian hamster, a single RGD sequence was found at the same position to that of the rat. Triplication of the RGD sequences was seen in the murine fibrinogen α-C domain around the homologous site to the rat RGDS sequence. These findings are of some interest from the point of view of structure-function and evolutionary relationships in the mammalian fibrinogen Aα-chains.

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