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	<td>0.5</td>		<td>0.5</td>
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Revision as of 17:07, 11 September 2008

β班の実験ノート

Contents 1 Week 1 1.1 17 August, 2008 1.2 18 August, 2008 1.3 19 August, 2008 1.4 20 August, 2008 1.5 21 August, 2008 1.6 22 August, 2008 1.7 23 August, 2008 2 Week 2 2.1 24 August, 2008 2.2 25 August, 2008 2.3 26 August, 2008 2.4 27 August, 2008 2.5 28 August, 2008 2.6 29 August, 2008 3 Week 3 4 Week 4

Week 1

17 August, 2008

Transformation

• BBa_R0040(2008) [http://partsregistry.org/Part:BBa_R0040] -> No colonies on plates
• BBa_C0070(2008) [http://partsregistry.org/Part:BBa_C0070] -> No colonies on plates
• BBa_C0076(2008) [http://partsregistry.org/Part:BBa_C0076] -> No colonies on plates
• BBa_C0077(2008) [http://partsregistry.org/Part:BBa_C0077] -> No colonies on plates
• BBa_C0078(2008) [http://partsregistry.org/Part:BBa_C0078] -> No colonies on plates
• BBa_C0079(2008) [http://partsregistry.org/Part:BBa_C0079] -> No colonies on plates

18 August, 2008

PCR

• BBa_C0070(2008) [http://partsregistry.org/Part:BBa_C0070]
• BBa_C0076(2008) [http://partsregistry.org/Part:BBa_C0076]
• BBa_C0077(2008) [http://partsregistry.org/Part:BBa_C0077]
• BBa_C0078(2008) [http://partsregistry.org/Part:BBa_C0078]
• BBa_C0079(2008) [http://partsregistry.org/Part:BBa_C0079]
• BBa_F2620(2007) [http://partsregistry.org/Part:BBa_F2620]･･･control

DNA Template	1
dNTP mix	10
Foward Primer	5
Reverse Primer	5
DNA polymerase	1
Thermopol Buffer	10
dH2O	68
TOTAL	100μL

95℃,5min -> ( 95℃,1min -> 52℃,1min -> 72℃,1min )･･･25cycles -> 72℃,10min -> 6℃

--->Gel Check

Sample DNA 3 Loading Dye 2 dH2O 7 TOTAL 12μL From left; BBa_C0070(2008) [http://partsregistry.org/Part:BBa_C0070] ->None BBa_C0076(2008) [http://partsregistry.org/Part:BBa_C0076] ->None BBa_C0077(2008) [http://partsregistry.org/Part:BBa_C0077] ->None BBa_C0078(2008) [http://partsregistry.org/Part:BBa_C0078] ->None BBa_C0079(2008) [http://partsregistry.org/Part:BBa_C0079] ->None BBa_F2620(2007) [http://partsregistry.org/Part:BBa_F2620]･･･control ->OK

Transformation

compitent cell : XL10G

• BBa_C0070(2007) [http://partsregistry.org/Part:BBa_C0070]
• BBa_C0070(2006) [http://partsregistry.org/Part:BBa_C0070]
• BBa_C0076(2007) [http://partsregistry.org/Part:BBa_C0076]
• BBa_C0076(2006) [http://partsregistry.org/Part:BBa_C0076] -> No colonies on plates(19/8)
• BBa_C0078(2007) [http://partsregistry.org/Part:BBa_C0078]
• BBa_C0078(2006) [http://partsregistry.org/Part:BBa_C0078]
• BBa_C0170(2007) [http://partsregistry.org/Part:BBa_C0170]
• BBa_C0170(2006) [http://partsregistry.org/Part:BBa_C0170]
• BBa_C0178(2007) [http://partsregistry.org/Part:BBa_C0178]
• BBa_C0178(2006) [http://partsregistry.org/Part:BBa_C0178]

--->(20/8)Mini prep

19 August, 2008

PCR

• BBa_I9026(2007) [http://partsregistry.org/Part:BBa_I9026]
• BBa_I9026(2006) [http://partsregistry.org/Part:BBa_I9026]
• BBa_I9030(2007) [http://partsregistry.org/Part:BBa_I9030]
• BBa_I9030(2006) [http://partsregistry.org/Part:BBa_I9030]
• BBa_S3154(2007) [http://partsregistry.org/Part:BBa_S3154]
• BBa_S3154(2006) [http://partsregistry.org/Part:BBa_S3154]
• BBa_F2620(2007) [http://partsregistry.org/Part:BBa_F2620]･･･control

DNA Template	1
dNTP mix	10
Foward Primer	5
Reverse Primer	5
DNA polymerase VENT	1
Thermopol Buffer	5
dH2O	28
TOTAL	50μL

95℃,5min -> ( 95℃,1min -> 52℃,1min -> 72℃,1min )･･･25cycles -> 72℃,10min -> 6℃

--->Gel Check

Sample DNA 3 Loading Dye 2 dH2O 7 TOTAL 12μL From left; BBa_I9026(2007) [http://partsregistry.org/Part:BBa_I9026] ->None BBa_I9026(2006) [http://partsregistry.org/Part:BBa_I9026] ->None BBa_I9030(2007) [http://partsregistry.org/Part:BBa_I9030] ->None BBa_I9030(2006) [http://partsregistry.org/Part:BBa_I9030] ->None BBa_S3154(2007) [http://partsregistry.org/Part:BBa_S3154] ->None BBa_S3154(2006) [http://partsregistry.org/Part:BBa_S3154] ->None BBa_F2620(2007) [http://partsregistry.org/Part:BBa_F2620]･･･control ->OK

20 August, 2008

(18/8)--->Mini prep

• BBa_C0070(2007) [http://partsregistry.org/Part:BBa_C0070]
• BBa_C0070(2006) [http://partsregistry.org/Part:BBa_C0070]
• BBa_C0076(2007) [http://partsregistry.org/Part:BBa_C0076]
• BBa_C0078(2007) [http://partsregistry.org/Part:BBa_C0078]
• BBa_C0078(2006) [http://partsregistry.org/Part:BBa_C0078]
• BBa_C0170(2007) [http://partsregistry.org/Part:BBa_C0170]
• BBa_C0170(2006) [http://partsregistry.org/Part:BBa_C0170]
• BBa_C0178(2007) [http://partsregistry.org/Part:BBa_C0178]
• BBa_C0178(2006) [http://partsregistry.org/Part:BBa_C0178]

--->Gel Check

Sample DNA 3 Loading Dye 2 dH2O 7 TOTAL 12μL From left; BBa_C0076(2007) [http://partsregistry.org/Part:BBa_C0076] -> None BBa_C0078(2007) [http://partsregistry.org/Part:BBa_C0078] -> None BBa_C0078(2006) [http://partsregistry.org/Part:BBa_C0078] -> None

Sample DNA 10 Loading Dye 4 dH2O 10 TOTAL 24μL From left; BBa_C0078(2007) [http://partsregistry.org/Part:BBa_C0078] -> None BBa_C0078(2006) [http://partsregistry.org/Part:BBa_C0078] -> OK BBa_C0076(2007) [http://partsregistry.org/Part:BBa_C0076] -> None BBa_C0070(2007) [http://partsregistry.org/Part:BBa_C0070] -> OK BBa_C0070(2006) [http://partsregistry.org/Part:BBa_C0070] -> OK

Sample DNA 3 Loading Dye 2 dH2O 7 TOTAL 12μL From left; BBa_C0170(2007) [http://partsregistry.org/Part:BBa_C0170] -> OK BBa_C0170(2006) [http://partsregistry.org/Part:BBa_C0170] -> OK BBa_C0178(2007) [http://partsregistry.org/Part:BBa_C0178] -> OK BBa_C0178(2006) [http://partsregistry.org/Part:BBa_C0178] -> OK

--->[Team:Chiba/protocol/digestion|Digestion test]]

• BBa_C0170(2007) [http://partsregistry.org/Part:BBa_C0170]
• BBa_C0170(2006) [http://partsregistry.org/Part:BBa_C0170]
• BBa_C0178(2007) [http://partsregistry.org/Part:BBa_C0178]
• BBa_C0178(2006) [http://partsregistry.org/Part:BBa_C0178]

Sample No	1	2	3	4	5	6	7	8
BBa_C0170(2006) [http://partsregistry.org/Part:BBa_C0170]	3	-	-	-	3	-	-	-
BBa_C0170(2007) [http://partsregistry.org/Part:BBa_C0170]	-	3	-	-	-	3	-	-
BBa_C0178(2006) [http://partsregistry.org/Part:BBa_C0178]	-	-	3	-	-	-	3	-
BBa_C0178(2007) [http://partsregistry.org/Part:BBa_C0178]	-	-	-	3	-	-	-	3
EcoRⅠ	0.1	0.1	0.1	0.1	0.1	0.1	0.1	0.1
PstⅠ	-	-	-	-	0.1	0.1	0.1	0.1
Buffer EcoRⅠ	1	1	1	1	1	1	1	1
BSA	1	1	1	1	1	1	1	1
dH2O	4.9	4.9	4.9	4.9	4.8	4.8	4.8	4.8
TOTAL	10	10	10	10	10	10	10	10

--->Gel Check

Sample DNA 3 Loading Dye 2 dH2O 7 TOTAL 12μL From left; (Single Digestion 1~4) BBa_C0170(2006) [http://partsregistry.org/Part:BBa_C0170] -> OK BBa_C0170(2007) [http://partsregistry.org/Part:BBa_C0170] -> OK BBa_C0178(2006) [http://partsregistry.org/Part:BBa_C0178] -> OK BBa_C0178(2007) [http://partsregistry.org/Part:BBa_C0178] -> OK (Double Digestion 1~4) BBa_C0170(2006) [http://partsregistry.org/Part:BBa_C0170] -> OK BBa_C0170(2007) [http://partsregistry.org/Part:BBa_C0170] -> OK BBa_C0178(2006) [http://partsregistry.org/Part:BBa_C0178] -> OK BBa_C0178(2007) [http://partsregistry.org/Part:BBa_C0178] -> OK

21 August, 2008

(20/8)--->PCR

• BBa_C0070(2007) [http://partsregistry.org/Part:BBa_C0070]
• BBa_C0070(2006) [http://partsregistry.org/Part:BBa_C0070]
• BBa_C0076(2007) [http://partsregistry.org/Part:BBa_C0076]
• BBa_C0078(2007) [http://partsregistry.org/Part:BBa_C0078]
• BBa_C0078(2006) [http://partsregistry.org/Part:BBa_C0078]

DNA Template	1
dNTP mix	10
Foward Primer	5
Reverse Primer	5
DNA polymerase TAQ	1
Thermopol Buffer	5
dH2O	28
TOTAL	50μL

95℃,5min -> ( 95℃,1min -> 52℃,1min -> 72℃,1min )･･･25cycles -> 72℃,10min -> 6℃

--->Gel Check

Sample DNA 3 Loading Dye 2 dH2O 7 TOTAL 12μL From left; BBa_C0078(2007) [http://partsregistry.org/Part:BBa_C0078] -> None BBa_C0078(2006) [http://partsregistry.org/Part:BBa_C0078] -> None BBa_C0076(2007) [http://partsregistry.org/Part:BBa_C0076] -> None BBa_C0070(2007) [http://partsregistry.org/Part:BBa_C0070] -> OK BBa_C0070(2006) [http://partsregistry.org/Part:BBa_C0070] -> OK

Transformation

• BBa_S03154(2007) [http://partsregistry.org/Part:BBa_S03154]
• BBa_S03154(2006) [http://partsregistry.org/Part:BBa_S03154]
• BBa_I9026(2007) [http://partsregistry.org/Part:BBa_I9026]
• BBa_I9026(2006) [http://partsregistry.org/Part:BBa_I9026]
• BBa_I9030(2007) [http://partsregistry.org/Part:BBa_I9030]
• BBa_I9030(2006) [http://partsregistry.org/Part:BBa_I9030]

Compitentcell : XL10G

--->(23/8)Digestion

22 August, 2008

Transformation

• BBa_J13002(2007) [http://partsregistry.org/Part:BBa_J13002]
• BBa_J13002(2006) [http://partsregistry.org/Part:BBa_J13002]
• BBa_J04500(2007) [http://partsregistry.org/Part:BBa_J04500]
• BBa_J04500(2006) [http://partsregistry.org/Part:BBa_J04500]

(Locate & 2 Punch ~iyama ver.~)

• BBa_C0076(2008) [http://partsregistry.org/Part:BBa_C0076] -> No colonies on plates
• BBa_C0077(2008) [http://partsregistry.org/Part:BBa_C0077] -> No colonies on plates
• BBa_C0078(2008) [http://partsregistry.org/Part:BBa_C0078] -> No colonies on plates
• BBa_J04500(2008) [http://partsregistry.org/Part:BBa_J04500] -> No colonies on plates

Compitentcell : XL10G

--->(24/8)Mini prep

23 August, 2008

(21/8)--->Mini prep

• BBa_S03154(2007) [http://partsregistry.org/Part:BBa_S03154]
• BBa_S03154(2006) [http://partsregistry.org/Part:BBa_S03154]
• BBa_I9026(2007) [http://partsregistry.org/Part:BBa_I9026]
• BBa_I9026(2006) [http://partsregistry.org/Part:BBa_I9026]
• BBa_I9030(2007) [http://partsregistry.org/Part:BBa_I9030]
• BBa_I9030(2006) [http://partsregistry.org/Part:BBa_I9030]

--->Digestion test

• BBa_S03154(2007) [http://partsregistry.org/Part:BBa_S03154]
• BBa_S03154(2006) [http://partsregistry.org/Part:BBa_S03154]
• BBa_I9026(2007) [http://partsregistry.org/Part:BBa_I9026]
• BBa_I9026(2006) [http://partsregistry.org/Part:BBa_I9026]
• BBa_I9030(2007) [http://partsregistry.org/Part:BBa_I9030]
• BBa_I9030(2006) [http://partsregistry.org/Part:BBa_I9030]
• BBa_J04500(2007) [http://partsregistry.org/Part:BBa_J04500]

Sample No	1	2	3	4	5	6	7	8	9	10	11	12	13	14	15	16	17	18	19	20	21
BBa_I9030(2007) [http://partsregistry.org/Part:BBa_I9030]	3	-	-	-	-	-	-	3	-	-	-	-	-	-	3	-	-	-	-	-	-
BBa_I9030(2006) [http://partsregistry.org/Part:BBa_I9030]	-	3	-	-	-	-	-	-	3	-	-	-	-	-	-	3	-	-	-	-	-
BBa_S03154(2007) [http://partsregistry.org/Part:BBa_S03154]	-	-	3	-	-	-	-	-	-	3	-	-	-	-	-	-	3	-	-	-	-
BBa_S03154(2006) [http://partsregistry.org/Part:BBa_S03154]	-	-	-	3	-	-	-	-	-	-	3	-	-	-	-	-	-	3	-	-	-
BBa_I9026(2007) [http://partsregistry.org/Part:BBa_I9026]	-	-	-	-	3	-	-	-	-	-	-	3	-	-	-	-	-	-	3	-	-
BBa_I9026(2007) [http://partsregistry.org/Part:BBa_I9026]	-	-	-	-	-	3	-	-	-	-	-	-	3	-	-	-	-	-	-	3	-
BBa_J04500(2007) [http://partsregistry.org/Part:BBa_J04500]	-	-	-	-	-	-	1	-	-	-	-	-	-	3	-	-	-	-	-	-	3
EcoRⅠ	-	-	-	-	-	-	-	0.1	0.1	0.1	0.1	0.1	0.1	0.1	0.1	0.1	0.1	0.1	0.1	0.1	0.1
PstⅠ	-	-	-	-	-	-	-	-	-	-	-	-	-	-	0.1	0.1	0.1	0.1	0.1	0.1	0.1
Buffer EcoRⅠ	-	-	-	-	-	-	-	0.9	0.9	0.9	0.9	0.9	0.9	0.9	0.8	0.8	0.8	0.8	0.8	0.8	0.8
BSA	-	-	-	-	-	-	-	1	1	1	1	1	1	1	1	1	1	1	1	1	1
dH2O	-	-	-	-	-	-	-	5	5	5	5	5	5	5	5	5	5	5	5	5	5
TOTAL	3	3	3	3	3	3	1	10	10	10	10	10	10	10	10	10	10	10	10	10	10

--->Gel Check

Sample No 1~6 7 Sample DNA 3 1 Loading Dye 1 1 dH2O 2 4 TOTAL 6 6 From left; Sanple No. 1~7 BBa_S03154(2007) [http://partsregistry.org/Part:BBa_S03154] BBa_S03154(2006) [http://partsregistry.org/Part:BBa_S03154] BBa_I9026(2007) [http://partsregistry.org/Part:BBa_I9026] BBa_I9026(2006) [http://partsregistry.org/Part:BBa_I9026] BBa_I9030(2007) [http://partsregistry.org/Part:BBa_I9030] BBa_I9030(2006) [http://partsregistry.org/Part:BBa_I9030] BBa_J04500(2007) [http://partsregistry.org/Part:BBa_J04500]

Sample No 8~21 Sample DNA 10 Loading Dye 2 TOTAL 12 From left; (Single Digestion) Sanple No. 8~14 BBa_S03154(2007) [http://partsregistry.org/Part:BBa_S03154] -> OK BBa_S03154(2006) [http://partsregistry.org/Part:BBa_S03154] -> OK BBa_I9026(2007) [http://partsregistry.org/Part:BBa_I9026] -> OK BBa_I9026(2006) [http://partsregistry.org/Part:BBa_I9026] -> OK BBa_I9030(2007) [http://partsregistry.org/Part:BBa_I9030] -> OK BBa_I9030(2006) [http://partsregistry.org/Part:BBa_I9030] -> OK BBa_J04500(2007) [http://partsregistry.org/Part:BBa_J04500] -> OK (Double Digestion) Sample No. 15~21 BBa_S03154(2007) [http://partsregistry.org/Part:BBa_S03154] -> OK BBa_S03154(2006) [http://partsregistry.org/Part:BBa_S03154] -> OK BBa_I9026(2007) [http://partsregistry.org/Part:BBa_I9026] -> OK BBa_I9026(2006) [http://partsregistry.org/Part:BBa_I9026] -> OK BBa_I9030(2007) [http://partsregistry.org/Part:BBa_I9030] -> OK BBa_I9030(2006) [http://partsregistry.org/Part:BBa_I9030] -> OK BBa_J04500(2007) [http://partsregistry.org/Part:BBa_J04500] -> OK?

Digestion test

BBa_J04500(2007:old-sample)[http://partsregistry.org/Part:BBa_J04500]

Sample No	1	2	3
Sample DNA	3	3	3
EcoRⅠ	-	0.1	0.1
SpeⅠ	-	-	0.1
EcoRⅠ Buffer	-	0.9	0.8
BSA	-	1	1
dH2O	-	5	5
TOTAL	3	10	10

--->Gel Check

Sample No 1 2 3 Sample DNA 3 10 10 Loading Dye 1 2 2 dH2O 2 - - TOTAL 6 12 12 From left; BBa_J04500(2007) [http://partsregistry.org/Part:BBa_J04500] -> OK BBa_J04500(2007) [http://partsregistry.org/Part:BBa_J04500]Single Digestion -> OK BBa_J04500(2007) [http://partsregistry.org/Part:BBa_J04500]Double Digestion -> Bad

Week 2

24 August, 2008

(22/8)--->Mini prep

1. BBa_J04500[http://partsregistry.org/Part:BBa_J04500](2007)
2. BBa_J04500[http://partsregistry.org/Part:BBa_J04500](2006)
3. BBa_J13002[http://partsregistry.org/Part:BBa_J13002](2007)
4. BBa_J13002[http://partsregistry.org/Part:BBa_J13002](2006)

--->Digestion test

1. BBa_J04500[http://partsregistry.org/Part:BBa_J04500](2007)
2. BBa_J04500[http://partsregistry.org/Part:BBa_J04500](2006)
3. BBa_J13002[http://partsregistry.org/Part:BBa_J13002](2007)
4. BBa_J13002[http://partsregistry.org/Part:BBa_J13002](2006)

Sample No	5	6	7	8
BBa_J04500[http://partsregistry.org/Part:BBa_J04500](2007)	1	-	-	-
BBa_J04500[http://partsregistry.org/Part:BBa_J04500](2006)	-	1	-	-
BBa_J13002[http://partsregistry.org/Part:BBa_J13002](2007)	-	-	1	-
BBa_J13002[http://partsregistry.org/Part:BBa_J13002](2007)	-	-	-	1
EcoRⅠ	0.1	0.1	0.1	0.1
Buffer 4	0.9	0.9	0.9	0.9
dH2O	8	8	8	8
TOTAL	10	10	10	10

--->Gel Check

Sample No 1~8 Sample DNA 1 Loading Dye 1 dH2O 4 TOTAL 6 From left; BBa_J04500[http://partsregistry.org/Part:BBa_J04500](2007) -> ? BBa_J04500[http://partsregistry.org/Part:BBa_J04500](2006) -> OK BBa_J13002[http://partsregistry.org/Part:BBa_J13002](2007) -> OK BBa_J13002[http://partsregistry.org/Part:BBa_J13002](2006) -> OK （↓single digestuion） BBa_J04500[http://partsregistry.org/Part:BBa_J04500](2007) -> OK BBa_J04500[http://partsregistry.org/Part:BBa_J04500](2006) -> OK BBa_J13002[http://partsregistry.org/Part:BBa_J13002](2007) -> OK BBa_J13002[http://partsregistry.org/Part:BBa_J13002](2006) -> None?

Sample No 5~8 Sample DNA 9 Loading Dye 2 dH2O 1 TOTAL 12 From the left; （single digestuion） 5, BBa_J04500[http://partsregistry.org/Part:BBa_J04500](2007) -> OK 6, BBa_J04500[http://partsregistry.org/Part:BBa_J04500](2006) -> OK 7, BBa_J13002[http://partsregistry.org/Part:BBa_J13002](2007) -> 3400bp:Bad, 2200bp:OK 8, BBa_J13002[http://partsregistry.org/Part:BBa_J13002](2006) -> OK

25 August, 2008

(24/8)--->Digestion

1. Double Digestion:BBa_J04500[http://partsregistry.org/Part:BBa_J04500](2007)
2. Double Digesiton:BBa_R0010[http://partsregistry.org/Part:BBa_R0010](2007)
3. Single Digestion:BBa_R0010[http://partsregistry.org/Part:BBa_R0010](2007)
4. BBa_R0010[http://partsregistry.org/Part:BBa_R0010](2007)

Sample No	1	2	3	4
Sample DNA	5	5	1	1
EcoRⅠ	0.5	o.5	-	-
SpeⅠ	0.5	0.5	-	-
Buffer 1	1	1	-	-
Buffer 3	-	-	1	-
BSA	1	1	-	-
dH2O	2	2	7.5	-
TOTAL	10	10	10	1

--->Gel Check

Sample No 1~3 4 Sample DNA 10 1 Loading Dye 2 1 dH2O - 4 TOTAL 12 6 From right; Double Digestion:BBa_J04500[http://partsregistry.org/Part:BBa_J04500](2007) -> OK Double Digesiton:BBa_R0010[http://partsregistry.org/Part:BBa_R0010](2007) -> ?(low) Single Digestion:BBa_R0010[http://partsregistry.org/Part:BBa_R0010](2007) -> ?(low) BBa_R0010[http://partsregistry.org/Part:BBa_R0010](2007) -> OK(low)

--->Digestion (Double Digestion)

1. BBa_C0178[http://partsregistry.org/Part:BBa_C0178]
2. BBa_C0170[http://partsregistry.org/Part:BBa_C0170]
3. BBa_J04500[http://partsregistry.org/Part:BBa_J04500](2007)

]

</tr>

Sample No	1	2	3
Sample DNA	33ng/μl×60μl	33ng/μl×60μl	100ng/μl×20μl
PstⅠ	2	2	2
XbaⅠ	2	2	-
SpeⅠ	-	-	2
Buffer 2	-	-	5
Buffer 3	10	10	-
BSA	10	10	5
dH2O	16	16	16
TOTAL	100	100	52

26 August, 2008

(8/25)--->Gel Check

Sample No 1~3 Sample DNA 30 Loading Dye 6 TOTAL 36 1, BBa_C0178[http://partsregistry.org/Part:BBa_C0178] Insert:609bp -> OK

2, BBa_C0170[http://partsregistry.org/Part:BBa_C0170] Insert:609bp -> OK

3, BBa_J04500[http://partsregistry.org/Part:BBa_J04500](2007) -> OK

--->Gel Extract

--->Zymo Clean

1. 5μl
2. 5μl
3. 27μl

--->Gel Check

Sample No 1~3 Sample DNA 2 Loading Dye 2 H2O 8 TOTAL 12 From left; insert:C0178 -> OK insert:C0170 -> OK? vector:J04500 -> OK

--->SAP

Sample No	3
Sample DNA	26
SAP	1
SAP Buffer	3
TOTAL	30μL

--->Zymo Clean

Sample No.3->9μl

--->Gel Check

Sample No.3 1 Loading Dye 1 dH2O 4 TOTAL 6 Sample No.3 -> OK ･･･30ng?

--->Ligation

]

</tr>

Sample No	(1)	(2)	(3)	(4)	(5)
insert①C0178	1	-	-	1	-
insert②C0170	-	1	-	-	1
vector③J04500	-	-	2	2	2
ligase	1	1	1	1	1
Buffer	2	2	2	2	2
dH2O	6	6	5	4	4
TOTAL	10	10	10	10	10

--->Transformation

1. insert①C0178
2. insert②C0170
3. vector③J04500
4. insert①C0178+vector③J04500
5. insert②C0170+vector③J04500

27 August, 2008

Digestion Test

Sample DNA : BBa_R0010[http://partsregistry.org/Part:BBa_R0010]

Sample No	1	2	3
Sample DNA	1	1	1
PstⅠ	-	-	0.5
EcoRⅠ	-	0.5	0.5
Buffer 3	-	1	1
BSA	-	-	1
dH2O	-	7.5	2
TOTAL	1	10	10

--->Gel Check

Sample No 1 2 3 Sample DNA 1 5 5 Loading Dye 1 1 1 H2O 4 - - TOTAL 6 6 6 From left; BBa_R0010 Single Digestion Double Digestion

(26/8)--->Colony-PCR

Colony PCR of 5 colonies from ligation plates (26/8-(4),(5)) and one from control plate(BBa_F2620[http://partsregistry.org/Part:BBa_F2620](2007)).

]

</tr>

Sample No	(4)-1~5	(5)-1~5	(F2620)
insert①C0178	1	-	-
insert②C0170	-	1	-
vector③J04500	-	-	2
ligase	1	1	1
Buffer	2	2	2
dH2O	6	6	5
TOTAL	10	10	10

--->Gel Check

Sample DNA 10 Loading Dye 2 TOTAL 12 From left; 1~5 insert:C0170 + vector:J04500 -> OK 6 F2620･･Positive Control -> OK 7~11 insert:C0178 + vector:J04500 -> OK

--->(28/8)Miniprep

Transformation comptent cell : XL10G

• BBa_C0062(2007) [http://partsregistry.org/Part:BBa_C0062]
• BBa_C0062(2006) [http://partsregistry.org/Part:BBa_C0062] -> no colony on plates
• BBa_C0179(2007) [http://partsregistry.org/Part:BBa_C0179]
• BBa_C0179(2006) [http://partsregistry.org/Part:BBa_C0179]
• BBa_S03156(2007) [http://partsregistry.org/Part:BBa_S03156]
• BBa_S03156(2006) [http://partsregistry.org/Part:BBa_S03156]
• BBa_S03158(2007) [http://partsregistry.org/Part:BBa_S03158]
• BBa_S03158(2006) [http://partsregistry.org/Part:BBa_S03158]
• BBa_S03160(2007) [http://partsregistry.org/Part:BBa_S03160]
• BBa_S0316(2006) [http://partsregistry.org/Part:BBa_S03160]

Compitentcell : XL10G

--->Mini prep

--->(29/8)Gel Check

28 August, 2008

(27/8)--->Mini prep

1. insert:C0170 + vector:J04500
2. insert:C0178 + vector:J04500

--->Digestion Test

1. insert:C0170 + vector:J04500
2. insert:C0178 + vector:J04500

Sample No	3	4	5	6
insert:C0170 + vector:J04500	1	-	1	-
insert:C0178 + vector:J04500	-	1	-	1
EcoRⅠ	0.1	0.1	0.1	0.1
SpeⅠ	-	-	0.1	0.1
Buffer 3	1	1	1	1
BSA	-	-	1	1
dH2O	7.8	7.8	6.8	6.8
TOTAL	10	10	10	10

--->(29/8)Gel Check

Sample No 1,2 3~6 Sample DNA 1 10 Loading Dye 1 2 H2O 4 - TOTAL 6 12 From left; 　-> OK 　-> OK 　-> OK 　-> OK 　-> OK 　-> OK

29 August, 2008

--->Digestion Test

1. BBa_R0010(2007) [http://partsregistry.org/Part:BBa_R0010]
2. BBa_C0062(2007) [http://partsregistry.org/Part:BBa_C0062]
3. BBa_C0179(2007) [http://partsregistry.org/Part:BBa_C0179]
4. BBa_C0179(2006) [http://partsregistry.org/Part:BBa_C0179]
5. BBa_S03156(2007) [http://partsregistry.org/Part:BBa_S03156]
6. BBa_S03156(2006) [http://partsregistry.org/Part:BBa_S03156]
7. BBa_S03158(2007) [http://partsregistry.org/Part:BBa_S03158]
8. BBa_S03158(2006) [http://partsregistry.org/Part:BBa_S03158]
9. BBa_S03160(2007) [http://partsregistry.org/Part:BBa_S03160]
10. BBa_S03160(2006) [http://partsregistry.org/Part:BBa_S03160]

Sample No	S1~10	ｗ1	ｗ2~10
Sample DNA	1	4	2
PstⅠ	-	0.075	0.1
EcoRⅠ	0.1	0.075	0.1
Buffer 3	1	0.75	1
BSA	-	0.75	1
dH2O	7.9	4.35	5.8
TOTAL	10	10	10

--->Gel Check

width="257">Sample No. 1~10 S1~S10,W1~W10 Sample DNA 1 10 Loading Dye 1 2 TOTAL 6 12 From left; 1~5,S1,S5,W1~W5

From left; 6~10,S6~S10,W6~W10

(27/8)--->Gel Check

Sample DNA 0.5 Loading Dye 1 dH2O 4.5 TOTAL 6μL R0010 -> OK

Digestion

①BBa_I9026(2007) [http://partsregistry.org/Part:BBa_I9026]

②BBa_I9030(2007) [http://partsregistry.org/Part:BBa_I9030]

③BBa_S03154(2007) [http://partsregistry.org/Part:BBa_S03154]

④BBa_R0010(2006) [http://partsregistry.org/Part:BBa_R0010]

Sample No	S①~③	④
Sample DNA	100ng/μL×20	200ng/μL×30
PstⅠ	2	2
XbaⅠ	2	-
SpeⅠ	-	2
Buffer 2	-	5
Buffer 3	5	-
BSA	5	5
dH2O	16	6
TOTAL	50	50

--->Gel Check

Sample DNA	10	3
Loading Dye	2	2
dH2O	-	7
TOTAL	12

From left;

From left;

From left;

From left;

)--->Gel Extract

--->Zymo Clean

=>1.5μL/2colum

--->SAP

 ④

--->Zymo Clean

---> miss!

• 30 August, 2008

Transformation comptent cell : XL10G

• BBa_I9026(2007) [http://partsregistry.org/Part:BBa_I9026]
• BBa_I9030(2006) [http://partsregistry.org/Part:BBa_I9030]
• BBa_S03154(2007) [http://partsregistry.org/Part:BBa_S03154]
• BBa_S03156(2007) [http://partsregistry.org/Part:BBa_S03156]
• BBa_S03158(2007) [http://partsregistry.org/Part:BBa_S03158]
• BBa_S03160(2007) [http://partsregistry.org/Part:BBa_S03160]
• BBa_C0062(2007) [http://partsregistry.org/Part:BBa_C0062]
• BBa_C0179(2007) [http://partsregistry.org/Part:BBa_C0179]

--->(31/8)Mini prep

(29/8)--->Mini prep

①insert:C0170 + vector:J04500

②insert:C0178 + vector:J04500

--->Digestion Test

--->Gel Check

From left;

From left;

From left;

Week 3

• 31 August, 2008

• 1 September, 2008

• 2 September, 2008

• 3 September, 2008

• 4 September, 2008

• 5 September, 2008

• 6 September, 2008

Week 4

• 7 September, 2008

• 8 September, 2008

• 9 September, 2008

• 10 September, 2008

ホーム	メンバー紹介	プロジェクト紹介	Parts Submitted to the Registry	モデリング	ノート

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