Team:HokkaidoU Japan/Notebook/asB0034/asB0034,32-1C3-construction

From 2014.igem.org

Notebook
Lab Documents

Anti-Sense B0034,B0032 on 1C3 Construction

  • Start

  • Digestion

    Cut vector with EcoRI, PstI (using 10×H buffer)
    Cut insert with EcoRI, PstI (using 10×M buffer)

  • Digestion

    Cut vector with EcoRI, PstI (using 10×H buffer)

  • Gel extraction

    digested vector insert

  • Ethanol Precipitation

    digested vector insert

  • Ligation

    Ligate vector with insert

  • Transformation

    ligation products
    5.0µL to DH5α

  • Colony PCR

    R0010-stem-asB0034-stem-B0015(on pSB1C3) 100up_F Tm:62.6℃ & asB0034_R Tm:68.7℃
    R0010-stem-asB0032-stem-B0015(on pSB1C3) 100up_F Tm:62.6℃ & asB0032_R Tm:70.8℃

  • Culture

    R0010-stem-asB0034-stem-B0015(on pSB1C3)
    R0010-stem-asB0032-stem-B0015(on pSB1C3)

  • Mini-Prep

    R0010-stem-asB0034-stem-B0015(on pSB1C3)
    R0010-stem-asB0032-stem-B0015(on pSB1C3)

  • Sequencing

    R0010-stem-asB0034-stem-B0015(on pSB1C3) with 100up_F Tm:62.8℃ & 200dn_R Tm:66.6℃
    R0010-stem-asB0032-stem-B0015(on pSB1C3) with 100up_F Tm:62.6℃ & 200dn_R Tm:66.6℃

  • Complete!

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