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Team:Bielefeld-CeBiTec/Notebook/Journal/C02-fixation/Jul
From 2014.igem.org
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<li><b>pHnCBcsoS1D</b></li> | <li><b>pHnCBcsoS1D</b></li> | ||
<ul> | <ul> | ||
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</ul> | </ul> | ||
</ul> | </ul> | ||
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| + | <br> | ||
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<li><b>csoS1D of the carboxysome</b></li> | <li><b>csoS1D of the carboxysome</b></li> | ||
<ul> | <ul> | ||
| - | <li>This week we tried to amplify | + | <li>This week we tried to amplify the backbone pSB1C3 for csoS1D and csoS1D itself.</li> |
<ul> | <ul> | ||
| - | <li><a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Protocols#PCR" target="_blank">PCR amplification</a> (<a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Primer# | + | <li><a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Protocols#PCR" target="_blank">PCR amplification</a> of csoS1D (<a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Primer#csoS1D_fwd" target="_blank">csoS1D_fwd</a>, <a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Primer#csoS1D_rev" target="_blank">csoS1D_rev</a>)</li> |
<ul> | <ul> | ||
| - | <li>Annealing temperature: | + | <li>Annealing temperature: 55 °C</li> |
<li>Bands as expected (~703 bp)</li> | <li>Bands as expected (~703 bp)</li> | ||
</ul> | </ul> | ||
| + | <li><a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Protocols#PCR" target="_blank">PCR amplification</a> of pSB1C3 (<a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Primer#pSB1C3_suf_csoS1D" target="_blank">pSB1C3_suf_csoS1D</a>, <a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Primer#pSB1C3_pre_csoS1D" target="_blank">pSB1C3_pre_csoS1D</a>)</li> | ||
| + | <ul> | ||
| + | <li>Annealing temperature: 55 °C</li> | ||
| + | <li>Bands as expected (~2070 bp)</li> | ||
| + | </ul> | ||
| + | |||
</ul> | </ul> | ||
</ul> | </ul> | ||
| + | |||
| + | <br> | ||
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<li><b>Carbonic anhydrase (can) of the carboxysome</b></li> | <li><b>Carbonic anhydrase (can) of the carboxysome</b></li> | ||
<ul> | <ul> | ||
| - | <li>This week we tried to amplify | + | <li>This week we tried to amplify the backbone pSB1C3 for can and can itself.</li> |
<ul> | <ul> | ||
| - | <li><a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Protocols#PCR" target="_blank">PCR amplification</a> (<a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Primer# | + | <li><a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Protocols#PCR" target="_blank">PCR amplification</a> of can (<a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Primer#csoS3_can_fwd" target="_blank">csoS3_can_fwd</a>, <a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Primer#csoS3_can_rev" target="_blank">csoS3_can_rev</a>)</li> |
<ul> | <ul> | ||
| - | <li>Annealing temperature: | + | <li>Annealing temperature: 55 °C</li> |
| - | <li>Bands | + | <li>Bands as expected (~1605 bp)</li> |
</ul> | </ul> | ||
| - | + | <li><a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Protocols#PCR" target="_blank">PCR amplification</a> of pSB1C3 (<a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Primer#pSB1C3_pre_can" target="_blank">pSB1C3_pre_can</a>, <a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Primer#pSB1C3_suf_can" target="_blank">pSB1C3_suf_can</a>)</li> | |
| + | <ul> | ||
| + | <li>Annealing temperature: 55 °C</li> | ||
| + | <li>Bands as expected (~2070 bp)</li> | ||
| + | </ul> | ||
| + | </ul> | ||
</ul> | </ul> | ||
| - | <li><b>Shell associated protein of the carboxysome (first protein)</b></li> | + | |
| + | <br> | ||
| + | |||
| + | <li><b>Shell associated protein of the carboxysome (first part of the protein)</b></li> | ||
<ul> | <ul> | ||
| - | <li>This week we tried to amplify | + | <li>This week we tried to amplify the backbone pSB1C3 for sap and the first part of sap: sap_1.</li> |
<ul> | <ul> | ||
| - | <li><a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Protocols#PCR" target="_blank">PCR amplification</a> (<a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Primer# | + | <li><a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Protocols#PCR" target="_blank">PCR amplification</a> of sap_1 (<a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Primer#csoS2_sap_1_fwd" target="_blank">csoS2_sap_1_fwd</a>, <a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Primer#csoS2_sap_1_rev" target="_blank">csoS2_sap_1_rev</a>)</li> |
<ul> | <ul> | ||
| - | <li>Annealing temperature: | + | <li>Annealing temperature: 55 °C</li> |
| - | <li>Bands as expected ( | + | <li>Bands as expected (~1235 bp)</li> |
</ul> | </ul> | ||
| - | + | <li><a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Protocols#PCR" target="_blank">PCR amplification</a> of pSB1C3 (<a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Primer#pSB1C3_pre_sap_1" target="_blank">pSB1C3_pre_sap_1</a>, <a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Primer#pSB1C3_suf_sap2" target="_blank">pSB1C3_suf_sap2</a>)</li> | |
| + | <ul> | ||
| + | <li>Annealing temperature: 55 °C</li> | ||
| + | <li>Bands as expected (~2070 bp)</li> | ||
| + | </ul> | ||
| + | </ul> | ||
</ul> | </ul> | ||
</ul> | </ul> | ||
Revision as of 15:56, 6 September 2014
 |
July |
 |
- pHnCBcsoS1D
- This week we aimed to isolate the plasmid of the strain ordered from addgene
- Plasmid isolation of pHnCBcsoS1D
- csoS1D of the carboxysome
- This week we tried to amplify the backbone pSB1C3 for csoS1D and csoS1D itself.
- PCR amplification of csoS1D (csoS1D_fwd, csoS1D_rev)
- Annealing temperature: 55 °C
- Bands as expected (~703 bp)
- PCR amplification of pSB1C3 (pSB1C3_suf_csoS1D, pSB1C3_pre_csoS1D)
- Annealing temperature: 55 °C
- Bands as expected (~2070 bp)
- Carbonic anhydrase (can) of the carboxysome
- This week we tried to amplify the backbone pSB1C3 for can and can itself.
- PCR amplification of can (csoS3_can_fwd, csoS3_can_rev)
- Annealing temperature: 55 °C
- Bands as expected (~1605 bp)
- PCR amplification of pSB1C3 (pSB1C3_pre_can, pSB1C3_suf_can)
- Annealing temperature: 55 °C
- Bands as expected (~2070 bp)
- Shell associated protein of the carboxysome (first part of the protein)
- This week we tried to amplify the backbone pSB1C3 for sap and the first part of sap: sap_1.
- PCR amplification of sap_1 (csoS2_sap_1_fwd, csoS2_sap_1_rev)
- Annealing temperature: 55 °C
- Bands as expected (~1235 bp)
- PCR amplification of pSB1C3 (pSB1C3_pre_sap_1, pSB1C3_suf_sap2)
- Annealing temperature: 55 °C
- Bands as expected (~2070 bp)
- csoS1D
- This week we tried to transform the construct.
- Gibson Assembly with csoS1D on pSB1C3
- Transformation with electrocompotetent cells
- Colony PCR (Primer1, Primer2)
- Annealing temperature: ...
- Bands not as expected (... bp). Size looked like template insert CFP.
- Restriction digestion with DpnI
- Bands (not) as expected (... bp)
- Carbonic anhydrase (can)
- This week we tried to transform the construct.
- Gibson Assembly with can and pSB1C3
- Transformation with electrocompotetent cells
- Transformation was not succesful.
- pHnCBcsoS1D backbone
- This week we tried to amplify the backbone of the plasmid.
- PCR amplification (Primer1, Primer2)
- Annealing temperature: ...
- Bands as expected (~13,200 bp)
- BioBricks (BBa_k731500 and BBa_Q01400)
- This week we tried to use promotors of two BioBricks of the parts distribution.
- Plasmid isolation of pSB1C3_pTac and pSB1C3_p_TetR
- csoS4AB
- This week we tried amplifiy shell proteins of the carboxysome.
- PCR amplification (Primer1, Primer2)
- Annealing temperature: ...
- Bands as expected (... bp)
- csoS1CAB
- This week we tried amplifiy shell proteins of the carboxysome.
- PCR amplification (Primer1, Primer2)
- Annealing temperature: ...
- Bands as expected (... bp)
- Carbonic anhydrase
- This week we tried to purify the construct from the gel.
- Purification of can with gel extraction
- Bands not as expected (... bp)
- Shell associated protein
- This week we tried to purify the construct from the gel.
- Purification of can with gel extraction
- Bands as expected (~1235 bp)
- Pore protein (csoS1D) of the carboxysom
- This week we tried to purify the construct from the gel.
- Purification of can with gel extraction
- Bands not as expected (~703 bp)
- SBPase (glpX)
- This week we tried to amplify both parts of glpX.
- PCR amplification (Primer1, Primer2 and Primer3, Primer4)
- Annealing temperature: ...
- Bands (not) as expected (... bp)
- Carbnonic anhydrase (can)
- This week we tried to transform the construct.
- Gibson Assembly with csoS1D and pSB1C3
- Restriction digestion with DpnI
- Bands (not) as expected (... bp)
- Colony PCR (Primer1, Primer2)
- Annealing temperature: ...
- Bands as expected (1900 bp)
- Plasmid isolation of can
- csoS1D
- This week we tried to transform the construct.
- Gibson Assembly with csoS1D and pSB1C3
- Restriction digestion with DpnI
- Bands (not) as expected (... bp)
- Colony PCR (Primer1, Primer2)
- Annealing temperature: ...
- Bands as expected (1000 bp)
- Restriction digestion with EcoRI and XbaI
- Bands (not) as expected (... bp)
- Plasmid isolation of csoS1D
- Phosphoribulokinase (prkA)
- This week we tried to transform the fragments of the gene synthesis
- Transformation with electrocompotetent cells
- Colony PCR (prkA_pHn_fwd, prkA_pHn_rev)
- Annealing temperature: ...
- Bands as expected (~1060 bp)
- Plasmid isolation of prkA
- sRNA:pfkA
- This week we tried to transform the fragments of the gene synthesis
- Transformation with electrocompotetent cells
- Plasmid isolation of sRNA:pfkA
- RuBisCO of H. neapolitanus
- This week we tried to transform the fragments of the gene synthesis
- Transformation with electrocompotetent cells
- Colony PCR (Primer1, Primer2)
- Annealing temperature: ...
- Bands as expected (~2100 bp)
- We plated sample number 1 for a plasmid isolation.
- Plasmid isolation of Hneap
- RuBisCO of S. elongatus
- This week we tried to transform the fragments of the gene synthesis
- Transformation with electrocompotetent cells
- BioBricks (BBa_I719005)
- This week we tried to use the promotor of the BioBrick of the parts distribution.
- Plasmid isolation of pSB1C3_T7 BBa_I719005
