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| Line 1,438: |
Line 1,438: |
| | <h6>9.28</h6> | | <h6>9.28</h6> |
| | <h6>Colony PCR</h6> | | <h6>Colony PCR</h6> |
| − | <h6>Successfully align the sequencing results with the objective sequences:</h6>
| + | <h6>Zhenyi Guo</h6> |
| − | <h6>Zhenyi Guo</h6>
| + | |
| − | <table border="1">
| + | |
| − | <tr> | + | |
| − | <td>Name
| + | |
| − | <td>Enzyme</td>
| + | |
| − | </tr>
| + | |
| − | <tr>
| + | |
| − | <td>prd29a
| + | |
| − | <td>XbaI and PstI</td>
| + | |
| − | </tr>
| + | |
| − | </table>
| + | |
| − | <img src="https://static.igem.org/mediawiki/2016/0/05/T--HUST-China--notebook-prd29a-fig02.png" alt="">
| + | |
| − | </p>
| + | |
| − | <p>
| + | |
| − | <h6>9.27</h6>
| + | |
| − | <h6>Restriction Enzyme Digestion</h6>
| + | |
| − | <h6>Zhenyi Guo</h6>
| + | |
| | <table border="1"> | | <table border="1"> |
| | <tr> | | <tr> |
Revision as of 13:48, 18 October 2016
Notebook
Notebook
see how these parts were constructed？
CII
7.24
PCR
Yufeng Mo
| Name
| Total |
| CII 1
| 50 μl |
| CII 2
| 50 μl |
| CII control group
| 30 μl |
7.25
Restriction Enzyme Digestion
Yufeng Mo
| Name
| Enzyme |
| CII
| XbaI and PstI |
7.26
DNA ligation&transformation
Yufeng Mo
| Name |
Backbone |
Strain |
Medium |
| CII |
psb1c3(Xba I and Pst I cut) |
DH5α |
LB(chl added) |
7.27
Colony PCR
Yufeng Mo
CIII
7.14
PCR
Yufeng Mo
7.15
Restriction Enzyme Digestion
Yufeng Mo
| Name
| Enzyme |
| plasmid psb1c3 1
| XbaI and SpeI |
| CIII
| XbaI and SpeI |
7.15
Dephosphorylation
Yufeng Mo
7.17
DNA ligation&transformation
Yufeng Mo
| Name
| Backbone
| Strain
| Medium |
| CIII
| psb1c3(XbaI and SpeI cut)
| DH5α
| LB(chl added) |
7.18
Colony PCR
Yufeng Mo
| Number
| Name |
| 1
| CIII 1 |
| 2
| CIII 2 |
| 3
| CIII 3 |
| 4
| CIII 4 |
| 5
| CIII 5 |
| 6
| CIII 6 |
RBS-CII
PCR
Yufeng Mo
| Name
| Total |
| RBS-CII 2
| 40 μl |
| RBS-CII 1
| 40 μl |
| RBS-CII 0
| 20 μl |
9.4
Restriction Enzyme Digestion
Yufeng Mo
| Name
| Enzyme |
| RBS-CII
| XbaI and PstI |
9.4
DNA ligation&transformation
Yufeng Mo
| Name
| Backbone
| Strain
| Medium |
| RBS-CII
| psb1c3(Xba I and Pst I cut)
| DH5α
| LB(chl added) |
9.5
Colony PCR
Yufeng Mo
| Number
| Name |
| 1
| RBS-CII 1 |
| 2
| RBS-CII 2 |
RBS-CIII
8.7
PCR
Ziyi Jiang
| Name
| Total |
| RBS-CIII
| 50 μl |
| RBS-CIII
| 50 μl |
8.7
Restriction Enzyme Digestion
Ziyi Jiang
| Name
| Enzyme |
| psb1c3
| XbaI and PstI |
| RBS-CIII
| XbaI and PstI |
8.10
DNA ligation&transformation
Ziyi Jiang
| Name
| Backbone
| Strain
| Medium |
| RBS-CIII
| psb1c3(XbaI and PstI cut)
| DH5α
| LB(chl added) |
8.11
Colony PCR
Ziyi Jiang
RBS-CIII-RBS-CIII
8.18
Restriction Enzyme Digestion
Ziyi Jiang
| Name
| Enzyme |
| RBS-CIII
| EcoRI and SpeI |
| RBS-CIII
| EcoRI and XbaI |
8.18
DNA ligation&Transformation
Ziyi Jiang
| Name
| Backbone
| Strain
| Medium |
| CIII-RBS
| CIII-RBS-psb1c3(EcoRI and XbaI cut)
| DH5α
| LB(chl added) |
8.19
Colony PCR
Ziyi Jiang
| Number
| Name |
| 13
| RBS-CIII-RBS-CIII-psb1c3 |
| 19
| RBS-CIII-RBS-CIII-psb1c3 |
RBS-CII-RBS-CII
9.12
PCR
Yufeng Mo
9.14
Restriction Enzyme Digestion
Yufeng Mo
| Name
| Enzyme |
| RBS-CII-psb1c3
| EcoRI and XbaI |
| RBS-CII-psb1c3
| EcoRI and SpeI |
9.14
DNA ligation&transformation
Yufeng Mo
| Name
| Backbone
| Strain
| Medium |
| RBS-CII
| RBS-CII-psb1c3(EcoRI and XbaI cut)
| DH5α
| LB(chl added) |
9.15
Colony PCR
Zhi Zeng
| Number
| Name |
| 1
| RBS-CII-RBS-CII-psb1c3 |
pRM-RBS-GFP-LAVtag
7.18
PCR
Xiaoyang Li
| Name
| Total |
| GFP-First
| 50 μl |
| Name
| Total |
| pRM-RBS-GFP-LAVtag
| 30 μl |
| pRM-RBS-GFP-LAVtag
| 30 μl |
| pRM-RBS-GFP-LAVtag
| 30 μl |
7.19
Restriction Enzyme Digestion
Xiaoyang Li
| Name
| Enzyme |
| GFP
| EcoRI and SpeI |
| psb1c3
| EcoRI and SpeI |
7.20
DNA ligation&transformation
Xiaoyang Li
| Name
| Backbone
| Strain
| Medium |
| pRM-RBS-GFP-LAVtag
| psb1c3(EcoRI and SpeI cut)
| DH5α
| LB(chl added) |
7.24
Colony PCR
Xiaoyang Li
| Number
| Name |
| 1
| pRM-RBS-GFP-LVAtag |
8.21
Restriction Enzyme Digestion
Yufeng Mo
| Name
| Enzyme |
| pRM-RBS-GFP-LVAtag
| EcoRI and PstI |
| pETDuet-1
| EcoRI and PstI |
8.21
DNA ligation&transformation
Yufeng Mo
| Name
| Backbone
| Strain
| Medium |
| pRM-RBS-GFP-LVAtag
| pETDuet-1(EcoRI and PstI cut)
| DH5α
| LB(Amp added) |
8.22
Colony PCR
Yufeng Mo
| Number
| Name |
| 1
| pRM-RBS-GFP-LVAtag-pETDuet 1 |
| 3
| pRM-RBS-GFP-LVAtag-pETDuet 3 |
| 4
| pRM-RBS-GFP-LVAtag-pETDuet 4 |
| 5
| pRM-RBS-GFP-LVAtag-pETDuet 5 |
| 6
| pRM-RBS-GFP-LVAtag-pETDuet 6 |
pRE-RBS-GFP-LVAtag
8.23
Restriction Enzyme Digestion
Yufeng Mo
| Name
| Enzyme |
| pRE-RBS-GFP-LVAtag
| XbaI and PstI |
8.23
DNA ligation&transformation
Yufeng Mo
| Name
| Backbone
| Strain
| Medium |
| pRE-RBS-GFP-LVAtag
| psb1c3(XbaI and PstI cut)
| DH5α
| LB(chl added) |
8.24
Colony PCR
Yufeng Mo
| Number
| Name |
| 1
| pRE-RBS-GFP-LVAtag-psb1c3 |
8.29
Restriction Enzyme Digestion
Yufeng Mo,Zhenyi Guo
| Name
| Enzyme |
| pRE-RBS-GFP-LVAtag-psb1c3
| EcoRI and PstI |
8.29
DNA ligation&transformation
Xiao Deng,Zhenyi Guo
| Name
| Backbone
| Strain
| Medium |
| pRE-RBS-GFP-LVAtag
| petDuet-1(EcoRI and PstI cut)
| DH5α
| LB(Amp added) |
8.30
Colony PCR
Yufeng Mo
| Number
| Name |
| 1
| pRE-RBS-GFP-LVAtag-petDuet-1 1 |
| 3
| pRE-RBS-GFP-LVAtag-petDuet-1 3 |
| 5
| pRE-RBS-GFP-LVAtag-petDuet-1 5 |
| 8
| pRE-RBS-GFP-LVAtag-petDuet-1 8 |
pR-RBS-GFP-LAVtag
8.2
PCR
Zhenyi Guo
| Name
| Total |
| pR-RBS-GFP-tag
| 50 μl |
| pR-RBS-GFP-tag
| 50 μl |
| pR-RBS-GFP-tag
| 50 μl |
8.2
Restriction Enzyme Digestion
Zhenyi Guo
| Name
| Enzyme |
| psb1c3
| EcoRI and PstI |
| pR-RBS-GFP-tag
| EcoRI and PstI |
8.2
DNA ligation&Transformation
Zhenyi Guo
| Name
| Backbone
| Strain
| Medium |
| pR-RBS-GFP-tag
| psb1c3(EcoRI and PstI cut)
| DH5α
| LB(chl added) |
8.4
Colony PCR
Zhenyi Guo
| Number
| Name |
| 2
| pR-RBS-GFP-tag |
8.24
Restriction Enzyme Digestion
Zhenyi Guo
| Name
| Enzyme |
| pR-RBS-GFP-tag
| EcoRI and PstI |
8.24
DNA ligation&transformation
Yufeng Mo
| Name
| Backbone
| Strain
| Medium |
| pR-RBS-GFP-tag
| pETDuet-1(EcoRI and PstI cut)
| DH5α
| LB(Amp added) |
8.26
Colony PCR
Yufeng Mo
| Number
| Name |
| 4
| pR-RBS-GFP-tag-pETDuet-1 4 |
| 6
| pR-RBS-GFP-tag-pETDuet-1 6 |
| 7
| pR-RBS-GFP-tag-pETDuet-1 7 |
SnRK2.2
9.10
PCR
Yufeng Mo
| Name
| Total |
| SnRK2.2 1
| 40 μl |
| SnRK2.2 2
| 40 μl |
| SnRK2.2 0
| 20 μl |
9.10
Restriction Enzyme Digestion
Yufeng Mo
| Name
| Enzyme |
| SnRK2.2
| XbaI and PstI |
9.10
DNA ligation&transformation
Yufeng Mo
| Name
| Backbone
| Strain
| Medium |
| SnRK2.2
| psb1c3(XbaI and PstI cut)
| DH5α
| LB(chl added) |
9.12
Colony PCR
Yufeng Mo
| Number
| Name |
| 1
| SnRK2.2-psb1c3 |
ABF2
9.10
PCR
Yufeng Mo
| Name
| Total |
| ABF2 1
| 40 μl |
| ABF2 2
| 40 μl |
| ABF2 0
| 20 μl |
9.10
Restriction Enzyme Digestion
Yufeng Mo
| Name
| Enzyme |
| ABF2
| XbaI and PstI |
9.10
DNA ligation&transformation
Yufeng Mo
| Name
| Backbone
| Strain
| Medium |
| ABF2
| psb1c3(XbaI and PstI cut)
| DH5α
| LB(chl added) |
9.11
Colony PCR
Yufeng Mo
| Number
| Name |
| 2
| ABF2 2 |
| 3
| ABF2 3 |
| 4
| ABF2 4 |
PP2CA
7.25
PCR
Yufeng Mo
| Name
| Total |
| PP2CA 1
| 25 μl |
| PP2CA 2
| 25 μl |
| PP2CA control group
| 25 μl |
7.26
Restriction enzyme digestion
Yufeng Mo
| Name
| Enzyme |
| PP2CA
| XbaI and PstI |
7.26
DNA ligation&transformation
Yufeng Mo
| Name
| Backbone
| Strain
| Medium |
| PP2CA
| psb1c3(Xba I and Pst I cut)
| E.coli DH5α
| LB(chl added) |
7.27
Colony PCR
Yufeng Mo
prd29A
9.27
PCR
Zhi Zeng
| Name
| Total |
| prd29a 1
| 50 μl |
| prd29a 2
| 50 μl |
| prd29a 3
| 50 μl |
| prd29a 4
| 50 μl |
9.27
Restriction Enzyme Digestion
Zhenyi Guo
| Name
| Enzyme |
| prd29a
| XbaI and PstI |
9.28
Colony PCR
Zhenyi Guo
| Number
| Name |
| 1
| prd29a-psb1c3 |
| 2
| prd29a-psb1c3 |
CI-TT-pR-RBS-GFP-LVAtag&CII-TT-pRE-RBS-GFP-LVAtag
8.17
Restriction Enzyme Digestion
Lin Di
| Name
| Enzyme |
| Duet-pet
| EcoRI and PstI |
| CI-TT-pR-RBS-GFP-LVAtag
| EcoRI and PstI |
| CII-TT-pRE-RBS-GFP-LVAtag
| EcoRI and PstI |
8.17
DNA ligation&Transformation
Lin Di
| Name
| Backbone
| Strain
| Medium |
| CI-TTADH1-pR-RBS-GFP-LVAtag
| Duet-pet(T7)
| DH5α
| LB(Amp added) |
| CII-TTADH1-pRE-RBS-GFP-LVAtag
| Duet-pet(T7)
| DH5α
| LB(Amp added) |
8.18
Colony PCR
Successfully align the sequencing results with the objective sequences:
Lin Di
| Number
| Name |
| 6
| CI-TTADH1-pR-RBS-GFP-LVAtag-petDuet-1 6 |
| 5
| CII-TTADH1-pRE-RBS-GFP-LVAtag-petDuet-1 5 |
| 11
| CII-TTADH1-pRE-RBS-GFP-LVAtag -petDuet-1 11 |
Cro-TT-pRM-RBS-GFP-LVAtag-PSB1C3
8.14
PCR
Zhi Zeng
8.15
Restriction Enzyme Digestion
Zhi Zeng
| Name
| Enzyme |
| pRM-RBS-GFP-LVAtag-psb1c3
| EcoRI and XbaI |
| psb1c3
| XbaI and PstI |
8.16
In-Fusion
Zhi Zeng
| Insert
| Vector
| Total |
| Cro&TT
| pRM-RBS-GFP-LVAtag-PSB1C3(Cut by EcoRⅠ&XbaⅠ)
| 5uL |
8.16
Transformation
Zhi Zeng
| Name
| Strain
| Medium |
| Cro-TT-pRM-RBS-GFP-LVAtag-PSB1C3 (ligate by Infusion)
| DH5α
| LB(Chl added) |
8.17
Colony PCR
Zhi Zeng
| Name
| Total |
| Cro-TT-pRM-RBS-GFP-LVAtag-psb1c3
| 20uL |
8.18
Restriction Enzyme Digestion
Zhi Zeng
| Name
| Enzyme |
| Duet-pet
| EcoRI and PstI |
| Cro-TT-pRM-RBS-GFP-LVAtag
| EcoRI and PstI |
8.18
DNA ligation&Transformation
Zhi Zeng
| Name
| Backbone
| Strain
| Medium |
| Cro-TT-pRM-RBS-GFP-LVAtag
| Duet-pet(T7)
| DH5α
| LB(Amp added) |
PP2CA-TTADH1
8.14
Restriction Enzyme Digestion
Ziyi Jiang
| Name
| Enzyme |
| TTADH1-psb1c3
| EcoRI and XbaI |
| PP2CA-psb1c3
| EcoRI and SpeI |
8.16
Colony PCR
Successfully align the sequencing results with the objective sequences:
Ziyi Jiang
| Number
| Name |
| 5
| PP2CA-TTADH1-psb1c3 5 |
| 7
| PP2CA-TTADH1-psb1c3 7 |
| 12
| PP2CA-TTADH1-psb1c3 12 |
PP2CA-TTADH1-pCyc
8.22
Overlap PCR
Zhi Zeng
| Name
| Total |
| PP2CA-TTADH1-Pcyc
| 80uL |
8.22
Restriction Enzyme Digestion
Zhi Zeng
| Backbone
| Enzyme |
| psb1c3
| PstI and XbaI |
8.23
In-Fusion
Zhi Zeng
| Insert
| Vector
| Total |
| PP2CA-TTADH1-pCyc
| psb1c3(Cut by PstⅠ&XbaⅠ)
| 5uL |
8.23
Transformation
Zhi Zeng
| Name
| Strain
| Medium |
| PP2CA-TTADH1-pCyc-psb1c3 (ligate by In-Fusion)
| DH5α
| LB(Chl added) |
8.24
Colony PCR
Zhi Zeng
| Number
| Name |
| 3
| PP2CA-TTADH1-pCyc-psb1c3 |
| 4
| PP2CA-TTADH1-pCyc-psb1c3 |
| 7
| PP2CA-TTADH1-pCyc-psb1c3 |
| 13
| PP2CA-TTADH1-pCyc-psb1c3 |
| 14
| PP2CA-TTADH1-pCyc-psb1c3 |
Kozak-SnRK2.2-TTADH1-pCyc
8.30
Overlap PCR
| Name
| Total |
| Kozak-Snrk2.2-TTADH1-pCyc
| 80uL |
9.5
In-Fusion
Zhi Zeng
| Insert
| Vector
| Total |
| Kozak-Snrk2.2-TTADH1-pCyc
| psb1c3(Cut by PstⅠ&XbaⅠ)
| 5uL |
9.5
Transformation
Zhi Zeng
| Name
| Strain
| Medium |
| Kozak-Snrk2.2-TTADH1-pCyc-psb1c3(ligate by In-Fusion)
| DH5α
| LB(Chl added) |
9.7
Colony PCR
Zhi Zeng
| Name
| Total |
| Kozak-Snrk2.2-TTADH1-pCyc-psb1c3
| 20uL |
placm-pRE-RBS-Cro-RBS-CII-TT
9.6
Overlap PCR
Jiyue Qin
| Name
| Total |
| Placm-pRE-RBS-Cro
| 50uL |
9.7
In-Fusion
Jiyue Qin
| Insert
| Vector
| Total |
| Placm-pRE-RBS-Cro&RBS-cII-TT
| psb1c3(Cut by PstⅠ&XbaⅠ)
| 10uL |
9.6
Transformation
Jiyue Qin
| Name
| Strain
| Medium |
| Placm-pRE-RBS-Cro-RBS-CII-TT-psb1c3(ligate by In-Fusion)
| DH5α
| LB(Chl added) |
9.8
Colony PCR
Jiyue Qin
| Number
| Name |
| 1
| Placm-pRE-RBS-Cro-RBS-CII-TT-psb1c3 |
patp2-RBS-CI-TTADH1-pR-RBS-CIII
9.2
Overlap PCR
Jiyue Qin
| Name
| Total |
| Patp2-RBS-CI-TT
| 50uL |
9.3
In-Fusion
Jiyue Qin
| Insert
| Vector
| Total |
| Patp2-RBS-CI-TT&pR-RBS-CIII
| psb1c3(Cut by PstⅠ&XbaⅠ)
| 10uL |
9.3
Transformation
Jiyue Qin
| Name
| Strain
| Medium |
| Patp2-RBS-CI-TT-pR-RBS-CIII-psb1c3
| DH5α
| LB(Chl added) |
9.4
Colony PCR
Jiyue Qin
| Number
| Name |
| 1
| Patp2-RBS-CI-TT-pR-RBS-CIII-psb1c3 |
RBS-iLDH-TT-pRM-RBS-beta-galactosidase
9.1
Restriction Enzyme Digestion
Jiyue Qin
| Name
| Enzyme |
| GFP-LVAtag-psb1c3
| XbaI and PstI |
9.1
In-Fusion
Jiyue Qin
| Insert
| Vector
| Total |
| RBS-iLDH-TT&pRM-RBS-beta-galactosidase
| psb1c3(Cut by PstⅠ&XbaⅠ)
| 10uL |
9.1
Transformation
Jiyue Qin
| Name
| Strain
| Medium |
| RBS-iLDH-TT-pRM-RBS-beta-galactosidase-psb1c3
| DH5α
| LB(Chl added) |
9.2
Colony PCR
Jiyue Qin
| Number
| Name |
| 1
| RBS-iLDH-TT-pRM-RBS-beta-galactosidase-psb1c3 |
plac-pRE-RBS-Cro-RBS-CII-TT-patp2-RBS-CI-TT-pR-RBS-CIII-RBS-iLDH-TT-pRM-RBS-beta-galactosidase
9.7
In-Fusion
Zhenyi Guo
| Insert
| Vector
| Total |
| plac-pRE-RBS-Cro-RBS-CII-TT&patp2-RBS-CI-TT-pR-RBS-CIII-RBS-iLDH-TT-pRM-RBS-beta-galactosidase
| psb1c3(Cut by PstⅠ&XbaⅠ)
| 10uL |
9.7
Transformation
Zhenyi Guo
| Name
| Strain
| Medium |
| plac-pRE-RBS-Cro-RBS-CII-TT-patp2-RBS-CI-TT-pR-RBS-CIII-RBS-iLDH-TT-pRM-RBS-beta-galactosidase
| DH5α
| LB(Chl added) |
9.9
Colony PCR
Zhenyi Guo
| Number
| Name |
| 1
| plac-pRE-RBS-Cro-RBS-CII-TT-patp2-RBS-CI-TT-pR-RBS-CIII-RBS-iLDH-TT-pRM-RBS-beta-galactosidase |
9.11
Restriction Enzyme Digestion
Zhenyi Guo
| Name
| Enzyme |
| Duet-pet
| EcoRI and PstI |
| plac-pRE-RBS-Cro-RBS-CII-TT-patp2-RBS-CI-TT-pR-RBS-CIII-RBS-iLDH-TT-pRM-RBS-beta-galactosidase -psb1c3
| EcoRI and PstI |
9.7
Transformation
Zhenyi Guo
| Name
| Strain
| Medium |
| plac-pRE-RBS-Cro-RBS-CII-TT-patp2-RBS-CI-TT-pR-RBS-CIII-RBS-iLDH-TT-pRM-RBS-beta-galactosidase
| DH5α
| LB(Chl added) |
9.17
DNA ligation&Transformation
Zhenyi Guo
| Name
| Backbone
| Strain
| Medium |
| plac-pRE-RBS-Cro-RBS-CII-TT-patp2-RBS-CI-TT-pR-RBS-CIII-RBS-iLDH-TT-pRM-RBS-beta-galactosidase
| Duet-pet(T7)(cut by EcoRI and PstI)
| DH5α
| LB(Amp added) |
9.18
Colony PCR
Successfully align the sequencing results with the objective sequences:
Zhenyi Guo
| Number
| Name |
| 1
| plac-pRE-RBS-Cro-RBS-CII-TT-patp2-RBS-CI-TT-pR-RBS-CIII-RBS-iLDH-TT-pRM-RBS-beta-galactosidase-Duet-pet |
