User:Karmella Haynes/Notebook/miR Trigger/2009/12/21

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12/21/09

  • ✓ Order oligos for RT-PCR analysis of YFP(miRNA) vs. RFP expression
  • ✓ Split cells: U2OS plain (1x 1:10 flask); JDS33-47 U2OS (1x 1:10 flask, 2x 6-well ab-free 2.5x105/well, 1x 24-well ab-free)
  • ✓ Cultures for minipreps: KAH93, 94, 95/MV2 (2 each)

Order oligos
> For RT-PCR analysis of YFP(from transgene JDS33/MV2) and RFP(from tansgenes KAH93, 94, 95/MV2) expression; 100 bp amplicons
> Design MV2 vector primers to detect any DNA contamination (non-transcribed sequence) in cDNA samples

  1. YFP_RTPCR f1: 5'-GCGCGCCTGAGAACTTCAGG (B-globin exon 2)
  2. YFP_RTPCR r1: 5'-ctcgcccttgctcactctag (scar-YFP1)
  3. YFP_RTPCR f2: 5'-gcatggacgagctgtacaag (YFP1 end)
  4. YFP_RTPCR r2: 5'-tggccgtttacgtcgccgtc (YFP2 near start)
  5. RFP_RTPCR f1: 5'-gcctctaacattgagacagc (Gal4DB near end)
  6. RFP_RTPCR r1: 5'-cggaaattccggcgatacag (Gal4DB end)
  7. RFP_RTPCR f2: 5'-gcatggacgagctgtacaag (RFP1 near end)
  8. RFP_RTPCR r2: 5'-gagccctccatgtgcaccttg (RFP2 near start)
  9. MV2_RTPCR f: 5'-CGACGCTCAAGTCAGAGGTG (backbone, non-coding)
  10. MV2_RTPCR r: 5'-GCAGGGTCGGAACAGGAGAG (backbone, non-coding)