Team:Chiba/Calendar-Home/26 August 2008

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25 August 2008 <|> 27 August 2008

Contents

Laboratory work

Team:Input

no work

Team:Communication

(8/25)--->Gel Check
Chiba-0825-2.JPG
Sample No. 1~3
Sample DNA(μL) 30
Loading Dye(μL) 6
TOTAL(μL) 36


1, [http://partsregistry.org/Part:BBa_C0178 BBa_C0178]
Insert:609bp -> OK
Chiba-0825-3.JPG
2, [http://partsregistry.org/Part:BBa_C0170 BBa_C0170]
Insert:609bp -> OK
Chiba-0825-4.JPG
3, [http://partsregistry.org/Part:BBa_J04500 BBa_J04500](2007)
--> OK


--->Gel Extract


--->Zymo Clean
eluted with following amount of elution buffer
  1. 5μl
  2. 5μl
  3. 27μl


--->Gel Check
Chiba-0825-5.JPG
Sample No. 1~3
Sample DNA(μL) 2
Loading Dye(μL) 2
H2O(μL) 8
TOTAL(μL) 12
From left;
  1. insert:[http://partsregistry.org/Part:BBa_C0178 BBa_C0178] --> OK
  2. insert:[http://partsregistry.org/Part:BBa_C0170 BBa_C0170] --> OK?
  3. vector:[http://partsregistry.org/Part:BBa_J04500 BBa_J04500] --> OK


--->SAP
Sample No. 3
Sample DNA(μL) 26
SAP(μL) 1
SAP Buffer(μL) 3
TOTAL(μL) 30


--->Zymo Clean
Sample No.3 -->9μl


--->Gel Check
Chiba-0826.JPG
Sample No.3 1
Loading Dye(μL) 1
dH2O(μL) 4
TOTAL(μL) 6
Sample No.3 -> OK ・・・30ng?


--->Ligation
Sample No. (1)(2)(3)(4)(5)
insert(1)[http://partsregistry.org/Part:BBa_C0178 BBa_C0178](μL) 1--1-
insert(2)[http://partsregistry.org/Part:BBa_C0170 BBa_C0170](μL) -1--1
vector(3)[http://partsregistry.org/Part:BBa_J04500 BBa_J04500](μL) --222
ligase(μL) 11111
Buffer(μL) 22222
dH2O(μL) 66544
TOTAL(μL) 1010101010


--->Transformation
  1. insert(1)[http://partsregistry.org/Part:BBa_C0178 BBa_C0178]
  2. insert(2)[http://partsregistry.org/Part:BBa_C0170 BBa_C0170]
  3. vector(3)[http://partsregistry.org/Part:BBa_J04500 BBa_J04500]
  4. insert(1)[http://partsregistry.org/Part:BBa_C0178 BBa_C0178]+vector(3)[http://partsregistry.org/Part:BBa_J04500 BBa_J04500]
  5. insert(2)[http://partsregistry.org/Part:BBa_C0170 BBa_C0170]+vector(3)[http://partsregistry.org/Part:BBa_J04500 BBa_J04500]

Team:Output

Gel Check

  • [http://partsregistry.org/Part:BBa_F2620 BBa_F2620]
  • [http://partsregistry.org/Part:BBa_F2620 BBa_F2620]
  • [http://partsregistry.org/Part:BBa_J63001 BBa_J63001]
Sample No.
DNA tamplate 11111
Loading Dye 11111
dH2O 44444
TOTAL 6μl6μl6μl6μl6μl

Digestion

  • vector:[http://partsregistry.org/Part:BBa_F2620 BBa_F2620]①
Sample No.
DNA tamplate 100
SpeⅠ 2
PstⅠ 2
BSA(x10) 13
Buffer2 13
TOTAL 130μl
  • insert:[http://partsregistry.org/Part:BBa_J52008 BBa_J52008]①
[http://partsregistry.org/Part:BBa_E2030 BBa_E2030]②
Sample No.
DNA tamplate 3030
XbaⅠ 11
PstⅠ 11
DpnⅠ 11
Buffer3 44
BSA(x10) 44
TOTAL 40μl40μl


-->37°C 3hour
-->SAP
-->37°C 30min
-->65°C 15min
-->Zymo Clean

Ligation

  • vector:[http://partsregistry.org/Part:BBa_F2620 BBa_F2620] insert:[http://partsregistry.org/Part:BBa_J52008 BBa_J52008]①
  • vector:[http://partsregistry.org/Part:BBa_F2620 BBa_F2620] insert:[http://partsregistry.org/Part:BBa_E2030 BBa_E2030]②
  • Negative control:[http://partsregistry.org/Part:BBa_F2620 BBa_F2620]③
Sample No.
vector 555
inser 107.50
Ligase Buffer 444
Ligase 111
dH2O 02.510
TOTAL 20μl20μl20μl

-->RT 2hour

-->Transformation(XL10G)

-->37°C over night

-->colony PCR

-->Mini prep