Detection of polymorphisms within the human IL10 receptor cDNA gene sequence by RT-PCR RFLP

1997 ◽  
Vol 46 (5) ◽  
pp. 439-441 ◽  
Author(s):  
Yosuke Tanaka ◽  
H. Nakashima ◽  
Takeshi Otsuka ◽  
Yoshiaki Nemoto ◽  
Hiroaki Niiro ◽  
...  
Keyword(s):  
Gene Sequence ◽  
Rt Pcr ◽  
Pcr Rflp ◽  
Receptor Cdna ◽  
Human Il10

Parathyroid Ca2+-conducting currents are modulated by muscarinic receptor agonists and antagonists

1997 ◽  
Vol 273 (5) ◽  
pp. E880-E890 ◽  
Author(s):  
Wenhan Chang ◽  
Tsui-Hua Chen ◽  
Stacy A. Pratt ◽  
Benedict Yen ◽  
Michael Fu ◽  
...  
Keyword(s):  
Muscarinic Receptors ◽  
Inositol Phosphate ◽  
Receptor Protein ◽  
Dependent Manner ◽  
Rt Pcr ◽  
Pipette Solution ◽  
Pcr Products ◽  
Inositol Phosphate Accumulation ◽  
Receptor Cdna ◽  
Dose Dependent

Parathyroid cells express Ca2+-conducting cation currents, which are activated by raising the extracellular Ca2+ concentration ([Ca2+]o) and blocked by dihydropyridines. We found that acetylcholine (ACh) inhibited these currents in a reversible, dose-dependent manner (50% inhibitory concentration ≈10−8 M). The inhibitory effects could be mimicked by the agonist (+)-muscarine. The effects of ACh were blunted by the antagonist atropine and reversed by removing ATP from the pipette solution. (+)-Muscarine enhanced the adenosine 3′,5′-cyclic monophosphate (cAMP) production by 30% but had no effect on inositol phosphate accumulation in parathyroid cells. Oligonucleotide primers, based on sequences of known muscarinic receptors (M1-M5), were used in reverse transcriptase-polymerase chain reaction (RT-PCR) to amplify receptor cDNA from parathyroid poly (A)+ RNA. RT-PCR products displayed >90% nucleotide sequence identity to human M2- and M4-receptor cDNAs. Expression of M2-receptor protein was further confirmed by immunoblotting and immunocytochemistry. Thus parathyroid cells express muscarinic receptors of M2 and possibly M4 subtypes. These receptors may couple to dihydropyridine-sensitive, cation-selective currents through the activation of adenylate cyclase and ATP-dependent pathways in these cells.

RT-PCR-RFLP analysis for evaluating cross protection by an attenuated isolate of Cucumber mosaic virus

2005 ◽  
Vol 71 (3) ◽  
pp. 243-246 ◽  
Author(s):  
Eiko Nakazono-Nagaoka ◽  
Masako Suzuki ◽  
Yoshitaka Kosaka ◽  
Tomohide Natsuaki
Keyword(s):  
Cucumber Mosaic Virus ◽  
Mosaic Virus ◽  
Rflp Analysis ◽  
Cross Protection ◽  
Rt Pcr ◽  
Pcr Rflp ◽  
Attenuated Isolate

scholarly journals Identification, Characterization, and Molecular Detection of Alfalfa mosaic virus in Potato

2006 ◽  
Vol 96 (11) ◽  
pp. 1237-1242 ◽  
Author(s):  
H. Xu ◽  
J. Nie
Keyword(s):  
Mosaic Virus ◽  
Gene Sequence ◽  
Alfalfa Mosaic Virus ◽  
Enzyme Linked Immunosorbent Assay ◽  
Polymorphism Analysis ◽  
Rt Pcr ◽  
Potato Leaf ◽  
Simple Method ◽  
Cp Gene ◽  
Nucleotide Sequence Alignment

Alfalfa mosaic virus (AMV) was detected in potato fields in several provinces in Canada and characterized by bioassay, enzyme-linked immunosorbent assay, and reverse-transcription polymerase chain reaction (RT-PCR). The identity of eight Canadian potato AMV isolates was confirmed by sequence analysis of their coat protein (CP) gene. Sequence and phylogenetic analysis indicated that these eight AMV potato isolates fell into one strain group, whereas a slight difference between Ca175 and the other Canadian AMV isolates was revealed. The Canadian AMV isolates, except Ca175, clustered together among other strains based on alignment of the CP gene sequence. To detect the virus, a pair of primers, AMV-F and AMV-R, specific to the AMV CP gene, was designed based on the nucleotide sequence alignment of known AMV strains. Evaluations showed that RT-PCR using this primer set was specific and sensitive for detecting AMV in potato leaf and tuber samples. AMV RNAs were easily detected in composite samples of 400 to 800 potato leaves or 200 to 400 tubers. Restriction analysis of PCR amplicons with SacI was a simple method for the confirmation of PCR tests. Thus, RT-PCR followed by restriction fragment length polymorphism analysis may be a useful approach for screening potato samples on a large scale for the presence of AMV.

scholarly journals Isolado do vírus do mosaico do pepino obtido de bananeira no Estado de São Paulo pertence ao subgrupo Ia

2001 ◽  
Vol 26 (1) ◽  
pp. 53-59 ◽  
Author(s):  
MARCELO EIRAS ◽  
ADDOLORATA COLARICCIO ◽  
ALEXANDRE L.R. CHAVES
Keyword(s):  
Mosaic Virus ◽  
Vigna Unguiculata ◽  
Sao Paulo ◽  
São Paulo ◽  
Rt Pcr ◽  
Nicotiana Glutinosa ◽  
Musa Sp ◽  
De Se ◽  
Pcr Rflp ◽  
Das Elisa

Em 1996, foi feita a caracterização parcial de um isolado do vírus do mosaico do pepino (Cucumis mosaic virus, CMV) obtido de bananeira (Musa sp.) proveniente do município de Miracatu, SP. Com o objetivo de se determinar o subgrupo do isolado de CMV, recorreu-se às técnicas de ELISA, RT-PCR, RFLP e seqüenciamento de fragmentos de RNA genômico. Amostras de folhas infetadas, desidratadas com cloreto de cálcio e armazenadas à -20 °C desde 1994 na viroteca do Laboratório de Fitovirologia e Fisiopatologia, foram inoculadas em plantas de Nicotiana glutinosa. Dez dias após a inoculação, folhas apresentando mosaico foram utilizadas para DAS-ELISA e extração de RNAs totais. Em ELISA, houve reação apenas contra o anti-soro específico para CMV subgrupo I. Através de RT-PCR com primers desenhados para anelar em regiões conservadas da porção terminal 3' do gene da capa protéica, foi amplificado um fragmento de DNA com 486 pares de bases. O produto obtido via RT-PCR foi submetido à digestão com as enzimas EcoRI, HindIII, BamHI e MspI, obtendo-se um padrão de restrição esperado para o subgrupo I. Estes resultados foram confirmados através do seqüenciamento do produto de PCR, o qual apresentou homologia de 96% a 98% com os isolados do CMV pertencentes ao subgrupo I. Pelos sintomas observados na hospedeira diferencial Vigna unguiculata, o isolado foi confirmado como sendo do subgrupo Ia.

Detection and differentiation of serologically cross-reacting tobamoviruses of economical importance by RT-PCR and RT-PCR-RFLP

2002 ◽  
Vol 106 (1) ◽  
pp. 1-10 ◽  
Author(s):  
Bettina Letschert ◽  
Günter Adam ◽  
Dietrich-Eckhardt Lesemann ◽  
Peter Willingmann ◽  
Cornelia Heinze
Keyword(s):  
Rt Pcr ◽  
Pcr Rflp

Carotenoid analysis of sweetpotato Ipomoea batatas and functional identification of its lycopene β- and ε-cyclase genes

2016 ◽  
Vol 71 (9-10) ◽  
pp. 313-322 ◽  
Author(s):  
Muhammad Zubair Khan ◽  
Miho Takemura ◽  
Takashi Maoka ◽  
Motoyasu Otani ◽  
Norihiko Misawa
Keyword(s):  
Ipomoea Batatas ◽  
Hplc Analysis ◽  
Gene Sequence ◽  
Efficient Production ◽  
Functional Identification ◽  
Rt Pcr ◽  
Pantoea Ananatis ◽  
E Coli ◽  
White Star ◽  
Β Carotene

Abstract Sweetpotato Ipomoea batatas is known as a hexaploid species. Here, we analyzed carotenoids contained in the leaves and tubers of sweetpotato cultivars ‘White Star’ (WS) and W71. These cultivars were found to contain several carotenoids unique to sweetpotato tubers such as β-carotene-5,6,5′,8′-diepoxide and β-carotene-5,8-epoxide. Next, we isolated two kinds of carotene cyclase genes that encode lycopene β- and ε-cyclases from the WS and W71 leaves, by RT-PCR and subsequent RACE. Two and three lycopene β-cyclase gene sequences were, respectively, isolated from WS, named IbLCYb1, 2, and from W71, IbLCYb3, 4, 5. Meanwhile, only a single lycopene ε-cyclase gene sequence, designated IbLCYe, was isolated from both WS and W71. These genes were separately introduced into a lycopene-synthesizing Escherichia coli transformed with the Pantoea ananatis crtE, crtB and crtI genes, followed by HPLC analysis. β-Carotene was detected in E. coli cells that carried IbLCYb1-4, indicating that the IbLCYb1-4 genes encode lycopene β-cyclase. Meanwhile, the introduction of IbLCYe into the lycopene-synthesizing E. coli led to efficient production of δ-carotene with a monocyclic ε-ring, providing evidence that the IbLCYe gene codes for lycopene ε-(mono)cyclase. Expression of the β- and ε-cyclase genes was analyzed as well.

Detection of morbillivirus infection by RT-PCR RFLP analysis in cetaceans and carnivores

2017 ◽  
Vol 247 ◽  
pp. 22-27 ◽  
Author(s):  
Federica Verna ◽  
Federica Giorda ◽  
Ilaria Miceli ◽  
Giovanna Rizzo ◽  
Alessandra Pautasso ◽  
...  
Keyword(s):  
Rflp Analysis ◽  
Rt Pcr ◽  
Pcr Rflp
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