Team:HokkaidoU Japan/Notebook/aggregation Week 6

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August 6th

Ethanol precipitation

Ethanol precipitation for digestion and gel extraction product.

  1. Added 5 ul of NaoAc, 1.5 ul of glycogen and 125ul of 100% ethanol.
  2. Centrifuged in 15000 rpm, 10 min at 4C.
  3. Remove supernatant and added 220ul of 70% ethanol.
  4. Centrifuged in 15000 rpm, 10 min at 4C.
  5. Remove supernatant and air drying in room temperature then added 10 ul of DW.

Ligation

We ligated pT7-RBS on pSB1K3 (digested Spe1) as vector and Ag43-dT (digested Spe1 and Xba1 and HindIII) as insert.

pT7-RBS 1 ul
Ag43-dT 2 ul
DW 2 ul
Ligation Mighty Mix(TAKARA) 5 ul
Total 10 ul


Ligation reaction time was written below.

Degree Minute
16 30
65 10
4 Hold