Hosuk:LabNotes/2013-12-30

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Revision as of 18:21, 10 January 2014 by >Hosuki78 (Created page with "*LabNote ===RT primer annealing specific position in mRNA=== ====Primer Design==== *Target gene : RAB7A *RAB7A mRNA target sequence: CAGAACTTGGACCTTCTCGCT...")
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RT primer annealing specific position in mRNA

Primer Design

  • Target gene : RAB7A
  • RAB7A mRNA target sequence: CAGAACTTGGACCTTCTCGCTTCTGTCCTCCGTTTAGTCTCCTC
    • This is base 125-177
  • Choose primers that are 10bp long while minimizing GC content and ~200bp, ~300bp, ~500bp, and ~1000bp from TSS


  • mRNA target of RAB7A 220b distance from TSS : CGCGTTTGAA
  • mRNA target of RAB7A 300b distance from TSS : ACTCATGAAC
  • mRNA target of RAB7A 510b distance from TSS : CAACACATTC
  • mRNA target of RAB7A 1000b distance from TSS: TTACACCCCA


  • RT_RAB7A_220b: [/5Phos/TCTCGGGAACGCTGAAGA] + TTCAAACGCG (--> D220)
  • RT_RAB7A_300b: [/5Phos/TCTCGGGAACGCTGAAGA] + GTTCATGAGT (--> D300)
  • RT_RAB7A_510b: [/5Phos/TCTCGGGAACGCTGAAGA] + GAATGTGTTG (--> D510)
  • RT_RAB7A_1000b: [/5Phos/TCTCGGGAACGCTGAAGA] + TGGGGTGTAA(--> D1000)


Rolony Fab.

  • Use 96 well plate
    • Control : Random Hexamer

File:PGP1F 96well RTpositionTest Cells 2.png