Template:Kyoto/Notebook/Aug 17
From 2013.igem.org
(Difference between revisions)
(→Colony PCR) |
(→Colony PCR) |
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|8/15 BBa_K112806(lysis2) -(1)||828 | |8/15 BBa_K112806(lysis2) -(1)||828 | ||
|- | |- | ||
- | | | + | |NC||-- |
|} | |} | ||
{| class="wikitable" | {| class="wikitable" | ||
Line 18: | Line 18: | ||
|5min||30s||30s||1min||30cycles | |5min||30s||30s||1min||30cycles | ||
|} | |} | ||
- | |||
{| class="wikitable" | {| class="wikitable" | ||
!Sample||base pair | !Sample||base pair | ||
Line 24: | Line 23: | ||
|8/10 BBa_Q0400 -(1)||3995 | |8/10 BBa_Q0400 -(1)||3995 | ||
|- | |- | ||
- | | | + | |NC||-- |
|} | |} | ||
{| class="wikitable" | {| class="wikitable" | ||
Line 33: | Line 32: | ||
|5min||30s||30s||30s||30cycles | |5min||30s||30s||30s||30cycles | ||
|} | |} | ||
- | [[File: | + | </div> |
+ | |||
+ | ===Electrophoresis=== | ||
+ | <div class="experiment"> | ||
+ | <span class="author">No name</span> | ||
+ | {| class="wikitable" | ||
+ | !Lane||Sample | ||
+ | |- | ||
+ | |1||1kbp ladder | ||
+ | |- | ||
+ | |2||K112806(lysis2) -1 | ||
+ | |- | ||
+ | |3||NC | ||
+ | |- | ||
+ | |4||Q04400(tetR) -1 | ||
+ | |- | ||
+ | |5||NC | ||
+ | |- | ||
+ | |6||1kbp ladder | ||
+ | |} | ||
+ | [[File:Igku Aug17colonyPCR.jpg]]<br> | ||
</div> | </div> | ||
Line 46: | Line 65: | ||
|8/15 BBa_Q04400(tetR) -(1)||Plusgrow medium(+Amp) | |8/15 BBa_Q04400(tetR) -(1)||Plusgrow medium(+Amp) | ||
|- | |- | ||
+ | |8/16 pBR322 -(1)||Plusgrow medium(+Amp) | ||
+ | |} | ||
+ | </div> | ||
+ | |||
+ | ===Master Plate=== | ||
+ | <div class="experiment"> | ||
+ | <span class="author">Nakamoto</span> | ||
+ | {| class="wikitable" | ||
+ | !Number||Use LB plate (+CP) | ||
+ | |- | ||
+ | |1||BBa_K112806(lysis2) -(1) | ||
+ | |} | ||
+ | </div> | ||
+ | <div class="experiment"> | ||
+ | {| class="wikitable" | ||
+ | !Number||Use LB plate (+Kana) | ||
+ | |- | ||
+ | |1||BBa_Q04400(tetR) -(1) | ||
+ | |} | ||
+ | </div> | ||
+ | ===Miniprep=== | ||
+ | <div class="experiment"> | ||
+ | <span class="author">Nakamoto and Murakami</span> | ||
+ | {|class="wikitable" | ||
+ | !DNA||concentration[µg/mL]||260/280||260/230 | ||
+ | |- | ||
+ | |K748002(lysis3) -(1)||330||1.66||2.08 | ||
+ | |- | ||
+ | |B0015(DT) -(1)||188||1.73||1.82 | ||
+ | |- | ||
+ | |R0061(Plux) -(1)||76||1.64||1.30 | ||
+ | |- | ||
+ | |K117000(lysis1) -(1)||286||1.71||2.01 | ||
+ | |- | ||
+ | |K117000(lysis1) -(2)||248||1.69||1.78 | ||
+ | |- | ||
+ | |I732020(RBS-lacZα-DT) -(1)||198||1.71||1.93 | ||
+ | |- | ||
+ | |I13504(RBS-GFP-DT) -(1)||248||1.69||2.03 | ||
+ | |- | ||
+ | |I13504(RBS-GFP-DT) -(2)||246||1.55||1.73 | ||
+ | |- | ||
+ | |I0462(RBS-luxR-DT) -(1)||326||1.65||1.74 | ||
+ | |} | ||
+ | </div> | ||
+ | |||
+ | ===Restriction Enzyme Digestion=== | ||
+ | <div class="experiment"> | ||
+ | <span class="author">Murata, Murakami, Nakamoto, Ashida, Matsumoto and Yoshida</span> | ||
+ | {| class="wikitable" | ||
+ | ! ||8/11 J23100 -(2)||SpeI||PstI||Buffer||MilliQ||total | ||
+ | |- | ||
+ | |2 cuts||4.0µL||1.0µL||1.0µL||3.0µL||21µL||30µL | ||
+ | |- | ||
+ | |1 cut||1.3µL||0.2µL||--||1.0µL||7.5µL||10µL | ||
+ | |- | ||
+ | |1 cut||1.3µL||--||0.2µL||1.0µL||7.5µL||10µL | ||
+ | |- | ||
+ | |NC||1.3µL||--||--||1.0µL||7.7µL||10µL | ||
+ | |} | ||
+ | {| class="wikitable" | ||
+ | ! ||8/15 B0034(RBS) -(2)||SpeI||PstI||Buffer||MilliQ||total | ||
+ | |- | ||
+ | |2 cuts||6.0µL||1.0µL||1.0µL||3.0µL||19µL||30µL | ||
+ | |- | ||
+ | |1 cut||2.0µL||0.2µL||--||1.0µL||6.8µL||10µL | ||
+ | |- | ||
+ | |1 cut||2.0µL||--||0.2µL||1.0µL||6.8µL||10µL | ||
+ | |- | ||
+ | |NC||2.0µL||--||--||1.0µL||7.0µL||10µL | ||
+ | |} | ||
+ | {| class="wikitable" | ||
+ | ! ||8/17 K117000(lysis1) -(1)||XbaI||PstI||BSA||Buffer||MilliQ||total | ||
+ | |- | ||
+ | |2 cuts||2.0µL||1.0µL||1.0µL||3.0µL||3.0µL||20µL||30µL | ||
+ | |- | ||
+ | |1 cut||1.0µL||0.2µL||--||1.0µL||1.0µL||6.8µL||10µL | ||
+ | |- | ||
+ | |1 cut||1.0µL||--||0.2µL||1.0µL||1.0µL||6.8µL||10µL | ||
+ | |- | ||
+ | |NC||1.0µL||--||--||1.0µL||1.0µL||7.0µL||10µL | ||
+ | |} | ||
+ | {| class="wikitable" | ||
+ | ! ||8/17 K748002(lysis3) -(1)||XbaI||PstI||BSA||Buffer||MilliQ||total | ||
+ | |- | ||
+ | |2 cuts||2.0µL||1.0µL||1.0µL||3.0µL||3.0µL||20µL||30µL | ||
+ | |- | ||
+ | |1 cut||1.0µL||0.2µL||--||1.0µL||1.0µL||6.8µL||10µL | ||
+ | |- | ||
+ | |1 cut||1.0µL||--||0.2µL||1.0µL||1.0µL||6.8µL||10µL | ||
+ | |- | ||
+ | |NC||1.0µL||--||--||1.0µL||1.0µL||7.0µL||10µL | ||
+ | |} | ||
+ | {| class="wikitable" | ||
+ | ! ||8/17 I13504(RBS-GFP-DT) -(1)||XbaI||PstI||BSA||Buffer||MilliQ||total | ||
+ | |- | ||
+ | |2 cuts||2.0µL||1.0µL||1.0µL||3.0µL||3.0µL||20µL||30µL | ||
+ | |- | ||
+ | |1 cut||1.0µL||0.2µL||--||1.0µL||1.0µL||6.8µL||10µL | ||
+ | |- | ||
+ | |1 cut||1.0µL||--||0.2µL||1.0µL||1.0µL||6.8µL||10µL | ||
+ | |- | ||
+ | |NC||1.0µL||--||--||1.0µL||1.0µL||7.0µL||10µL | ||
+ | |} | ||
+ | {| class="wikitable" | ||
+ | ! ||8/17 I0462(RBS-luxR-DT) -(1)||XbaI||PstI||BSA||Buffer||MilliQ||total | ||
+ | |- | ||
+ | |2 cuts||2.0µL||1.0µL||1.0µL||3.0µL||3.0µL||20µL||30µL | ||
+ | |- | ||
+ | |1 cut||1.0µL||0.2µL||--||1.0µL||1.0µL||6.8µL||10µL | ||
+ | |- | ||
+ | |1 cut||1.0µL||--||0.2µL||1.0µL||1.0µL||6.8µL||10µL | ||
+ | |- | ||
+ | |NC||1.0µL||--||--||1.0µL||1.0µL||7.0µL||10µL | ||
+ | |} | ||
+ | {| class="wikitable" | ||
+ | ! ||8/17 I732020(RBS-lacZα-DT) -(1)||XbaI||PstI||BSA||Buffer||MilliQ||total | ||
+ | |- | ||
+ | |2 cuts||2.0µL||1.0µL||1.0µL||3.0µL||3.0µL||20µL||30µL | ||
+ | |- | ||
+ | |1 cut||1.0µL||0.2µL||--||1.0µL||1.0µL||6.8µL||10µL | ||
+ | |- | ||
+ | |1 cut||1.0µL||--||0.2µL||1.0µL||1.0µL||6.8µL||10µL | ||
+ | |- | ||
+ | |NC||1.0µL||--||--||1.0µL||1.0µL||7.0µL||10µL | ||
+ | |} | ||
+ | {| class="wikitable" | ||
+ | ! ||8/11 R6061 -(1)||SpeI||PstI||Buffer||MilliQ||total | ||
+ | |- | ||
+ | |2 cuts||8.0µL||1.0µL||1.0µL||3.0µL||17µL||30µL | ||
+ | |- | ||
+ | |1 cut||3.0µL||0.2µL||--||1.0µL||5.8µL||10µL | ||
+ | |- | ||
+ | |1 cut||3.0µL||--||0.2µL||1.0µL||5.8µL||10µL | ||
+ | |- | ||
+ | |NC||3.0µL||--||--||1.0µL||6.0µL||10µL | ||
+ | |} | ||
+ | *incubate at 37 °C, for 1 h | ||
+ | </div> | ||
+ | |||
+ | ===Electrophoresis=== | ||
+ | <div class="experiment"> | ||
+ | <span class="author">No name</span> | ||
+ | {| class="wikitable" | ||
+ | !Lane||Sample||Enzyme1||Enzyme2 | ||
+ | |- | ||
+ | |1||J23100 -②||SpeI||PstI | ||
+ | |- | ||
+ | |2||J23100 -②||SpeI||-- | ||
+ | |- | ||
+ | |3||J23100 -②||--||PstI | ||
+ | |- | ||
+ | |4||J23100 -②||--||-- | ||
+ | |- | ||
+ | |5||B0034(RBS) -(2)||SpeI||PstI | ||
+ | |- | ||
+ | |6||B0034(RBS) -(2)||SpeI||-- | ||
+ | |- | ||
+ | |7||B0034(RBS) -(2)||--||PstI | ||
+ | |- | ||
+ | |8||B0034(RBS) -(2)||--||-- | ||
+ | |- | ||
+ | |9||1kb ladder||--||-- | ||
+ | |- | ||
+ | |10||K11700(lysis1) -(1)||XbaI||PstI | ||
+ | |- | ||
+ | |11||K11700(lysis1) -(1)||XbaI||-- | ||
+ | |- | ||
+ | |12||K11700(lysis1) -(1)||--||PstI | ||
+ | |- | ||
+ | |13||K11700(lysis1) -(1)||--||-- | ||
+ | |- | ||
+ | |14||K748002(lysis3) -(1)||XbaI||PstI | ||
+ | |- | ||
+ | |15||K748002(lysis3) -(1)||XbaI||-- | ||
+ | |- | ||
+ | |16||K748002(lysis3) -(1)||--||PstI | ||
+ | |- | ||
+ | |17||K748002(lysis3) -(1)||--||-- | ||
+ | |} | ||
+ | [[File:Igku Aug17 Electrophoresis(1).jpg]]<br> | ||
+ | {| class="wikitable" | ||
+ | !Lane||Sample||Enzyme1||Enzyme2 | ||
+ | |- | ||
+ | |18||I13504(RBS-GFP-DT) -(1)||XbaI||PstI | ||
+ | |- | ||
+ | |19||I13504(RBS-GFP-DT) -(1)||XbaI||-- | ||
+ | |- | ||
+ | |20||I13504(RBS-GFP-DT) -(1)||--||PstI | ||
+ | |- | ||
+ | |21||I13504(RBS-GFP-DT) -(1)||--||-- | ||
+ | |- | ||
+ | |22||I0462(RBS-luxR-DT) -(1)||XbaI||PstI | ||
+ | |- | ||
+ | |23||I0462(RBS-luxR-DT) -(1)||XbaI||-- | ||
+ | |- | ||
+ | |24||I0462(RBS-luxR-DT) -(1)||--||PstI | ||
+ | |- | ||
+ | |25||I0462(RBS-luxR-DT) -(1)||--||-- | ||
+ | |- | ||
+ | |26||1kb ladder||--||-- | ||
+ | |- | ||
+ | |27||I732020(RBS-lacZα-DT) -(1)||XbaI||PstI | ||
+ | |- | ||
+ | |28||I732020(RBS-lacZα-DT) -(1)||XbaI||-- | ||
+ | |- | ||
+ | |29||I732020(RBS-lacZα-DT) -(1)||--||PstI | ||
+ | |- | ||
+ | |30||I732020(RBS-lacZα-DT) -(1)||--||-- | ||
+ | |- | ||
+ | |31||R0061(Plux) -(1)||SpeI||PstI | ||
+ | |- | ||
+ | |32||R0061(Plux) -(1)||SpeI||-- | ||
+ | |- | ||
+ | |33||R0061(Plux) -(1)||--||PstI | ||
+ | |- | ||
+ | |34||R0061(Plux) -(1)||--||-- | ||
+ | |} | ||
+ | [[File:Igku Aug17 Electrophoresis(2).jpg]]<br> | ||
+ | </div> | ||
+ | |||
+ | ===Miniprep=== | ||
+ | <div class="experiment"> | ||
+ | <span class="author">Murata</span> | ||
+ | {|class="wikitable" | ||
+ | !DNA||concentration[µg/mL]||260/280||260/230 | ||
+ | |- | ||
+ | |8/17 PBR322(TaKaRa) -(1)||47||1.68||1.55 | ||
+ | |- | ||
+ | |8/17 K112806(lysis2) -(1)||64||1.67||1.70 | ||
|} | |} | ||
</div> | </div> |
Latest revision as of 07:41, 27 September 2013
Contents |
Aug 17
Colony PCR
Sample | base pair |
---|---|
8/15 BBa_K112806(lysis2) -(1) | 828 |
NC | -- |
PreDenature | Denature | Annealing | Extension | cycle |
---|---|---|---|---|
94°C | 94°C | 55°C | 68°C | -- |
5min | 30s | 30s | 1min | 30cycles |
Sample | base pair |
---|---|
8/10 BBa_Q0400 -(1) | 3995 |
NC | -- |
PreDenature | Denature | Annealing | Extension | cycle |
---|---|---|---|---|
94°C | 94°C | 55°C | 68°C | -- |
5min | 30s | 30s | 30s | 30cycles |
Electrophoresis
Liquid Culture
Sample | medium |
---|---|
8/15 BBa_K112806(lysis2) -(1) | Plusgrow medium(+CP) |
8/15 BBa_Q04400(tetR) -(1) | Plusgrow medium(+Amp) |
8/16 pBR322 -(1) | Plusgrow medium(+Amp) |
Master Plate
Number | Use LB plate (+CP) |
---|---|
1 | BBa_K112806(lysis2) -(1) |
Number | Use LB plate (+Kana) |
---|---|
1 | BBa_Q04400(tetR) -(1) |
Miniprep
DNA | concentration[µg/mL] | 260/280 | 260/230 |
---|---|---|---|
K748002(lysis3) -(1) | 330 | 1.66 | 2.08 |
B0015(DT) -(1) | 188 | 1.73 | 1.82 |
R0061(Plux) -(1) | 76 | 1.64 | 1.30 |
K117000(lysis1) -(1) | 286 | 1.71 | 2.01 |
K117000(lysis1) -(2) | 248 | 1.69 | 1.78 |
I732020(RBS-lacZα-DT) -(1) | 198 | 1.71 | 1.93 |
I13504(RBS-GFP-DT) -(1) | 248 | 1.69 | 2.03 |
I13504(RBS-GFP-DT) -(2) | 246 | 1.55 | 1.73 |
I0462(RBS-luxR-DT) -(1) | 326 | 1.65 | 1.74 |
Restriction Enzyme Digestion
8/11 J23100 -(2) | SpeI | PstI | Buffer | MilliQ | total | |
---|---|---|---|---|---|---|
2 cuts | 4.0µL | 1.0µL | 1.0µL | 3.0µL | 21µL | 30µL |
1 cut | 1.3µL | 0.2µL | -- | 1.0µL | 7.5µL | 10µL |
1 cut | 1.3µL | -- | 0.2µL | 1.0µL | 7.5µL | 10µL |
NC | 1.3µL | -- | -- | 1.0µL | 7.7µL | 10µL |
8/15 B0034(RBS) -(2) | SpeI | PstI | Buffer | MilliQ | total | |
---|---|---|---|---|---|---|
2 cuts | 6.0µL | 1.0µL | 1.0µL | 3.0µL | 19µL | 30µL |
1 cut | 2.0µL | 0.2µL | -- | 1.0µL | 6.8µL | 10µL |
1 cut | 2.0µL | -- | 0.2µL | 1.0µL | 6.8µL | 10µL |
NC | 2.0µL | -- | -- | 1.0µL | 7.0µL | 10µL |
8/17 K117000(lysis1) -(1) | XbaI | PstI | BSA | Buffer | MilliQ | total | |
---|---|---|---|---|---|---|---|
2 cuts | 2.0µL | 1.0µL | 1.0µL | 3.0µL | 3.0µL | 20µL | 30µL |
1 cut | 1.0µL | 0.2µL | -- | 1.0µL | 1.0µL | 6.8µL | 10µL |
1 cut | 1.0µL | -- | 0.2µL | 1.0µL | 1.0µL | 6.8µL | 10µL |
NC | 1.0µL | -- | -- | 1.0µL | 1.0µL | 7.0µL | 10µL |
8/17 K748002(lysis3) -(1) | XbaI | PstI | BSA | Buffer | MilliQ | total | |
---|---|---|---|---|---|---|---|
2 cuts | 2.0µL | 1.0µL | 1.0µL | 3.0µL | 3.0µL | 20µL | 30µL |
1 cut | 1.0µL | 0.2µL | -- | 1.0µL | 1.0µL | 6.8µL | 10µL |
1 cut | 1.0µL | -- | 0.2µL | 1.0µL | 1.0µL | 6.8µL | 10µL |
NC | 1.0µL | -- | -- | 1.0µL | 1.0µL | 7.0µL | 10µL |
8/17 I13504(RBS-GFP-DT) -(1) | XbaI | PstI | BSA | Buffer | MilliQ | total | |
---|---|---|---|---|---|---|---|
2 cuts | 2.0µL | 1.0µL | 1.0µL | 3.0µL | 3.0µL | 20µL | 30µL |
1 cut | 1.0µL | 0.2µL | -- | 1.0µL | 1.0µL | 6.8µL | 10µL |
1 cut | 1.0µL | -- | 0.2µL | 1.0µL | 1.0µL | 6.8µL | 10µL |
NC | 1.0µL | -- | -- | 1.0µL | 1.0µL | 7.0µL | 10µL |
8/17 I0462(RBS-luxR-DT) -(1) | XbaI | PstI | BSA | Buffer | MilliQ | total | |
---|---|---|---|---|---|---|---|
2 cuts | 2.0µL | 1.0µL | 1.0µL | 3.0µL | 3.0µL | 20µL | 30µL |
1 cut | 1.0µL | 0.2µL | -- | 1.0µL | 1.0µL | 6.8µL | 10µL |
1 cut | 1.0µL | -- | 0.2µL | 1.0µL | 1.0µL | 6.8µL | 10µL |
NC | 1.0µL | -- | -- | 1.0µL | 1.0µL | 7.0µL | 10µL |
8/17 I732020(RBS-lacZα-DT) -(1) | XbaI | PstI | BSA | Buffer | MilliQ | total | |
---|---|---|---|---|---|---|---|
2 cuts | 2.0µL | 1.0µL | 1.0µL | 3.0µL | 3.0µL | 20µL | 30µL |
1 cut | 1.0µL | 0.2µL | -- | 1.0µL | 1.0µL | 6.8µL | 10µL |
1 cut | 1.0µL | -- | 0.2µL | 1.0µL | 1.0µL | 6.8µL | 10µL |
NC | 1.0µL | -- | -- | 1.0µL | 1.0µL | 7.0µL | 10µL |
8/11 R6061 -(1) | SpeI | PstI | Buffer | MilliQ | total | |
---|---|---|---|---|---|---|
2 cuts | 8.0µL | 1.0µL | 1.0µL | 3.0µL | 17µL | 30µL |
1 cut | 3.0µL | 0.2µL | -- | 1.0µL | 5.8µL | 10µL |
1 cut | 3.0µL | -- | 0.2µL | 1.0µL | 5.8µL | 10µL |
NC | 3.0µL | -- | -- | 1.0µL | 6.0µL | 10µL |
- incubate at 37 °C, for 1 h
Electrophoresis
Lane | Sample | Enzyme1 | Enzyme2 |
---|---|---|---|
1 | J23100 -② | SpeI | PstI |
2 | J23100 -② | SpeI | -- |
3 | J23100 -② | -- | PstI |
4 | J23100 -② | -- | -- |
5 | B0034(RBS) -(2) | SpeI | PstI |
6 | B0034(RBS) -(2) | SpeI | -- |
7 | B0034(RBS) -(2) | -- | PstI |
8 | B0034(RBS) -(2) | -- | -- |
9 | 1kb ladder | -- | -- |
10 | K11700(lysis1) -(1) | XbaI | PstI |
11 | K11700(lysis1) -(1) | XbaI | -- |
12 | K11700(lysis1) -(1) | -- | PstI |
13 | K11700(lysis1) -(1) | -- | -- |
14 | K748002(lysis3) -(1) | XbaI | PstI |
15 | K748002(lysis3) -(1) | XbaI | -- |
16 | K748002(lysis3) -(1) | -- | PstI |
17 | K748002(lysis3) -(1) | -- | -- |
Lane | Sample | Enzyme1 | Enzyme2 |
---|---|---|---|
18 | I13504(RBS-GFP-DT) -(1) | XbaI | PstI |
19 | I13504(RBS-GFP-DT) -(1) | XbaI | -- |
20 | I13504(RBS-GFP-DT) -(1) | -- | PstI |
21 | I13504(RBS-GFP-DT) -(1) | -- | -- |
22 | I0462(RBS-luxR-DT) -(1) | XbaI | PstI |
23 | I0462(RBS-luxR-DT) -(1) | XbaI | -- |
24 | I0462(RBS-luxR-DT) -(1) | -- | PstI |
25 | I0462(RBS-luxR-DT) -(1) | -- | -- |
26 | 1kb ladder | -- | -- |
27 | I732020(RBS-lacZα-DT) -(1) | XbaI | PstI |
28 | I732020(RBS-lacZα-DT) -(1) | XbaI | -- |
29 | I732020(RBS-lacZα-DT) -(1) | -- | PstI |
30 | I732020(RBS-lacZα-DT) -(1) | -- | -- |
31 | R0061(Plux) -(1) | SpeI | PstI |
32 | R0061(Plux) -(1) | SpeI | -- |
33 | R0061(Plux) -(1) | -- | PstI |
34 | R0061(Plux) -(1) | -- | -- |
Miniprep
DNA | concentration[µg/mL] | 260/280 | 260/230 |
---|---|---|---|
8/17 PBR322(TaKaRa) -(1) | 47 | 1.68 | 1.55 |
8/17 K112806(lysis2) -(1) | 64 | 1.67 | 1.70 |