Double-nested polymerase chain reaction for detection of caprine arthritis-encephalitis virus proviral DNA in blood, milk, and tissues of infected goats

Barlough, Jeffrey E.; East, Nancy; Rowe, Joan D; oosear, Karen Van; DeRock, E.; Bigornia, Luisa; Rimstad, Espen

doi:10.1016/0166-0934(94)90167-8

Cited by 64 publications

(41 citation statements)

References 15 publications

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“…The cells were washed twice in minimum essential medium (MEM) and once in sterile phosphate-buffered saline (PBS), diluted in 200 p1 lysis buffer 110 mM Tris hydrochloride (pH 8.3), 0.45% NP-40. 0.45% Tween 20, 50 pg proteinase K ml-'1, and incubated in a 56°C water bath for 3 h. The proteinase K was subsequently inactivated by incubating the samples at 97°C for 15 min (Barlough et al 1994).…”

Section: Methodsmentioning

confidence: 99%

“…Portions of the following tissues were obtained: skin, gill, liver, heart, spleen, fore kidney, hind kidney, stomach, lower intestine, pyloric cecae, gonad, and brain. The samples were processed for PCR by phenolchloroform extraction, essentially as described (Barlough et al 1993(Barlough et al , 1994. The DNA concentration was assessed by absorbance at 260 nm, and the working concentration was adjusted to 0.2-1 pg DNA for each PCR reaction.…”

Section: Methodsmentioning

confidence: 99%

“…Appropriate internal Non-radioactive Southern blotting. The oligonucleotide probe ES-R was used to verify the identity of the 407 bp PCR I1 product by Southern blotting, essentially as described (Barlough et al 1994(Barlough et al , 1995. A nonradioactive, digoxigenin (DIG)-based system was employed (GeniusT"; Boehringer Mannheim, Indianapolis, IN) (Martin et al 1990).…”

Section: Methodsmentioning

confidence: 99%

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Nested polymerase chain reaction for detection of Enterocytozoon salmonis genomic DNA in chinook salmon Oncorhynchus tshawytscha

et al. 1995

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A nested polymerase chain reaction (PCR) was developed for detection of the microsporidian parasite Enterocytozoon saln~onis in blolog~cal samples (blood buffy-coat cells, feces, tissues, lymphocyte cultures) of chinook salmon Oncorhynchus tshaulj/tscha. A major second-round PCR product of 407 bp was readily identifiable in ethidium bromide-stained agarose minigels An internal probe was used to verify the identity of the amplified product by non-radioactive (digoxigeninbased) Southern blotting; final confirmation was made by DNA sequence analys~s. A dilution study using infected lymphocytes from in vitro cultures indicated that a single round of PCR (35 cycles) was able to detect E. salrnonis DNA from approximately 1000 infected cells. Sensitivity was increased with the full nested PCR (35 additional cycles), which detected parasite DNA from 110 infected lymphocytes. The specificity of the PCR was assessed with a panel of microsporidian and myxosporean DNAs. In an experimental infection study, E. salmonis DNA was detected in blood, feces, and tissues of infected chinook salmon but not in uninfected control fish.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Nested polymerase chain reaction for detection of Enterocytozoon salmonis genomic DNA in chinook salmon Oncorhynchus tshawytscha

et al. 1995

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“…O cDNA foi amplificado inicialmente com um par de iniciadores externos (primers 1 -5'-caagcagcaggagggagaagctg-3' e 2 -5'tcctacccccataatttgatccac-3'), resultando na amplificação de um fragmento de DNA de 296pb. O produto desta amplificação foi, então, amplificado novamente com iniciadores internos (primers 3 -5´-gttccagcaactgcaaacagtagcaatg-3´ e 4 -5´-acctttctgcttcttcatttaatttccc -3'), descritos por Barlough et al, (1994), resultando em um fragmento de 187pb.…”

Section: Coleta De Materialsunclassified

“…As condições de amplificação são realizadas segundo metodologia descrita por Barlough et al (1994), com modificações (Andrioli et al, 2006).…”

Section: Coleta De Materialsunclassified

Detecção do vírus da Artrite Encefalite Caprina por nested PCR e nested RT-PCR em ovócitos e fluido uterino

et al. 2013

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Caprine arthritis-encephalitis (CAE) is aninfectious disease caused by the caprine arthritis-encephalitis virus (CAEV), belonging to the lentivirus genus. The presence of the virus has been observed in the nervous system, respiratory tract and mammary gland, and also in the male and female genital tract. The objective of this study is to identify the virus in oocyte and uterine fluid of infected goats by molecular diagnostic techniques, in order to assess the possibility of CAEV transmission with reproduction. Thirteen infected goats were selected and submitted to euthanasia for the collection of the reproductive system, aspiration of the uterine fluid and dissection of ovaries for oocyte collection. In order to identify the CAEV in the collected material, in the protovirus and free forms, it was submitted to the nRT-PCR and nPCR techniques, respectively. As a result, it was observed that 53.8% of oocytes were positive to nRT-PCR, while only 9.1% were positive to nPCR. The nRT-PCR also identified the virus in the uterine fluid of 46.1% of the tested females. Even though the 13 goats had CAEV, 30.8% presented negative results in nPCR and nRT-PCR in all of the analyzed samples (oocyte and uterine fluid). This work concludes that nRT-PCR and nPCR can be used in the diagnosis of CAE for the analysis with oocytes and uterine fluid, and that the presence of CAEV in these materials points out to the risk of CAEV transmission through reproductive technologies used in females.

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PCR-Based Methods — A Promising Tool for Detection and Identification of Fungi in Soil

Lübeck

1996

Developments in Plant Pathology

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Double-nested polymerase chain reaction for detection of caprine arthritis-encephalitis virus proviral DNA in blood, milk, and tissues of infected goats

Cited by 64 publications

References 15 publications

Nested polymerase chain reaction for detection of Enterocytozoon salmonis genomic DNA in chinook salmon Oncorhynchus tshawytscha

Nested polymerase chain reaction for detection of Enterocytozoon salmonis genomic DNA in chinook salmon Oncorhynchus tshawytscha

Detecção do vírus da Artrite Encefalite Caprina por nested PCR e nested RT-PCR em ovócitos e fluido uterino

PCR-Based Methods — A Promising Tool for Detection and Identification of Fungi in Soil

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