Selected article for: "additional file and nr database"
Author: Peña, Andrea A; Bols, Niels C; Marshall, Sergio H
Title: An evaluation of potential reference genes for stability of expression in two salmonid cell lines after infection with either Piscirickettsia salmonis or IPNV
  • Document date: 2010_4_14
    • ID: 0xn6mqh6_9
    Snippet: Five reference genes (ACTB, UBQ, EF1A, GAPDH and TUBA), belonging to different functional classes, were selected to reduce the chance of their co-regulation ( Table 1 ). All primers were designed on conserved regions so that they could amplify each gene in both species under study: O. mykiss and O. tschawitcha. Primers were evaluated with the OligoCalc application [23] to check annealing temperatures and self-complementarity. The specificity of t.....
    Document: Five reference genes (ACTB, UBQ, EF1A, GAPDH and TUBA), belonging to different functional classes, were selected to reduce the chance of their co-regulation ( Table 1 ). All primers were designed on conserved regions so that they could amplify each gene in both species under study: O. mykiss and O. tschawitcha. Primers were evaluated with the OligoCalc application [23] to check annealing temperatures and self-complementarity. The specificity of the primers was tested using BLAST analysis against the nr NCBI database. Primer specifications are summarised on Additional file 1. The desired amplicon length (182 -204 base pairs) was chosen to be similar among all genes to avoid significant differences in PCR efficiencies due to amplicon length. PCR products were cloned into TOPO vector (pCR 2.1, Invitrogen) and submitted to sequencing for verification. Partial O. tschawitcha sequences were deposited into GenBank accession numbers: FJ890356 (EF1A), FJ890357 (ACTB), FJ890359 (UBQ) and FJ890358 (TUBA).

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