Team:Tokyo Metropolitan/Notebook/Fiber/2010/10/23

E.coli Fiber Project Notebook

2010/10/23 Saturday (Naoto)

member

naoto and watachin

Experiment:Sequence

material

For PCR

• bcsA (No.9)
• Big Dye
• primer
• DW

For Ethanol precipitation

• ethanol
• EDTA
• Hi-Di solution

procedure

1. mix materials(DNA<50ng)
2. PCR
3. Add EDTA and Ethanol and put at room temperature　(15min)
4. centrifuge (8000rpm,30min) and throw away supernatant
5. add ethanol again
6. centrifuge (8000rpm,15min) and throw away supernatant
7. add Hi-Di solution
8. heat at 95℃
9. transfer these sample to plate for sequence
10. read sequence

result bcsA wasn't inserted into pSB1C3...

Experiment:Colony PCR

material

• colony of E.coli
  • bcsB:1~32
  • bcsC:33~64
  • bcsD:65~96

other materials were same as protocol3

procedure

see protocol3

result

From the length of bands

bcsC→No.54 and 64

bcsD→No.75

seem to be correct inserts