Construction of an Internal Control to Quantitate Multiple Porcine Cytokine mRNAs by RT-PCR
| Autor: | N.R. Jayagopala Reddy, Bruce N. Wilkie, Bonnie A. Mallard |
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| Jazyk: | angličtina |
| Rok vydání: | 1996 |
| Předmět: | |
| Zdroj: | BioTechniques, Vol 21, Iss 5, Pp 868-875 (1996) |
| Druh dokumentu: | article |
| ISSN: | 1940-9818 0736-6205 |
| DOI: | 10.2144/96215st05 |
| Popis: | A multiple internal control was constructed to be used as an exogenously added control in reverse transcription polymerase chain reaction (RT-PCR) for pig cytokines. It consists of 5′ and 3′ primer sequences in the order of β2 microglobulin (β2-m), IL-1, IL-4, IL-6, IL-8, IL-2, IL-10, TNF-α, TNF-β and IFN-γ. Construction was accomplished by overlapping and extension PCR (OEPCR) utilizing short oligonucleotides. The primers were designed to give two products of different sizes on co-amplification of control and target RNAs by RT-PCR in a single tube. This permits analysis of message for several cytokines using a single exogenously added competitive template. Incorporated endonuclease sites allow construct modification by oligonucleotide addition. |
| Databáze: | Directory of Open Access Journals |
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