Team:Tokyo Metropolitan/Notebook/Fiber/2010/09/15

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E.coli Fiber Project Notebook

August 2010
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Contents

2010/9/15 Wednesday (Naoto)

Experiment:Electrophoresis

member

naoto and watachin

material

  • PCR production (pSB1C3)

If you want to know other materials, see the protocol4


procedure

see the protocol4


result

bands could be seen slightly

Experiment:subculture of A.xylinum

member

naoto and watachin

material

see protocol1 A.xylinum(sub-cultured at 8/24 and 9/1)

procedure

see protocol1 we made 3 OWW plates and 3 tubes of broth

Experiment:Extraction of DNA from agarose gel with QIAGEN

member

naoto and watachin

material

  • pSB1C3 in agarose gel

If you want to know other materials, see the protocol5


procedure

see the protocol5


Experiment:PCR

member

naoto and watachin

material

  • pSB1C3,bcsB(A.xylinum)
  • a culture of E.coli

If you want to know other materials, see the protocol3

procedure

  1. add materials to PCR tubes
  2. PCR reaction (see below)
  • nitialization :95°C 3min
  • denaturation:96°C 1min
  • annealing :51°C 1min
  • elongation:72°C 3min

(30cycles from 96°C 1min to 72°C 3min)

  • reaction stop :10°C ∞

Experiment:Electrophoresis

member

naoto and watachin

material

  • PCR production (see above PCR)

If you want to know other materials, see the protocol4

procedure

see the protocol4

result

a band of pSB1C3 could be seen slightly bands of bcsB(A.xylinum),bcsC(E.coli) appeared clearly

Experiment:Digestion

member naoto and watachin

material

  • pSB1C3,bcsA(A.xylinum) and bcsC(E.coli)
  • 10×Mbuffer
  • BSA
  • XbaI
  • SpeI
composition
pBS1C3bcsAbcsC
solution of bcs(μl) 505050
10×Mbuffer(μl) 5 5 5
BSA(μl) 5 5 5
XbaI(μl) 1 1 1
SpeI(ul) 1 1 1

procedure

  1. add "material" to PCR tubes(show below)
  2. incubation at 37℃,14h

Experiment:Extraction of DNA from agarose gel

member

naoto and watachin

material

  • pSB1C3,bcsB(A.xylinum) and bcsC(E.coli) in agarose gel

If you want to know other materials, see the protocol5

procedure

see the protocol5