Team:Columbia-Cooper/GenspaceNotebook
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+ | <h2>July 5</h2> | ||
+ | <p>- Grew out BBa_k325219 and BBa_k325909 in media C.</p> | ||
+ | <p>- Restriction digest of BBa_k231000 and BBa_k231001 with EcoRI and PstI</p> | ||
+ | <h2>July 15</h2> | ||
+ | <p>- CDS7 isolation and ligation</p> | ||
+ | |||
+ | <h2>July 19</h2> | ||
+ | <p>- NcoI and BamHI digest of pET Vector</p> | ||
+ | |||
+ | <h2>July 20</h2> | ||
+ | <p>- Another attempt at NcoI and BamHI digest of pET Vector</p> | ||
+ | |||
+ | <h2>July 21</h2> | ||
+ | <p>- Gel Purified pET Vector Digest </p> | ||
+ | <p>- Grew out more pET vector</p> | ||
+ | |||
+ | <h2>July 24</h2> | ||
+ | <p>- Received new NcoI and BamHI enzymes </p> | ||
+ | <p>- pET Vector digest with new NcoI and new BamHI</p> | ||
+ | <p>- pET Vector digest with old NcoI and old BamHI</p> | ||
+ | <p>- pET Vector digest with new NcoI and another with old NcoI</p> | ||
+ | <p>- pET Vector digest with BamHI and anther with old BamHI</p> | ||
+ | |||
+ | <h2>July 25</h2> | ||
+ | <p>- Ran a gel with all pET Vector digests from July 24th</p> | ||
+ | <p>- Fermentas transformation on pET Vector</p> | ||
+ | |||
+ | <h2>July 28</h2> | ||
+ | <p>- CDS7 digest with NcoI and BamHI</p> | ||
+ | <p>- Prepared plates of kanamycin and pET Vector</p> | ||
+ | <p>- Oligo annealed with Silver Lab protocol</p> | ||
+ | <p>- Maxiprep using ~50mL CDS7 growth</p> | ||
+ | <p>- pET Vector inoculation</p> | ||
+ | |||
+ | <h2>July 29</h2> | ||
+ | <p>- pET Vector digest with BamHI and NcoI</p> | ||
+ | <p>- Gel purification of pET Vector digest</p> | ||
+ | <p>- Inoculated more pET Vector</p> | ||
+ | <p>- Inoculated C-media for transformations</p> | ||
+ | <p>- Miniprep of pET Vector </p> | ||
+ | <p>- Ligated 4 oligos</p> | ||
+ | <p>- Prepare transformation ready cells using C-media culture prepared the previous day</p> | ||
+ | |||
+ | |||
+ | <h2>July 30</h2> | ||
+ | <p>- Miniprep of pET Vector</p> | ||
+ | <p>- Digestion of pET Vector with NcoI and BamHI</p> | ||
+ | <p>- Oligo PCR</p> | ||
+ | <p>-Digest pUC19 with EcoRI and PstI</p> | ||
+ | |||
+ | <h2>August 1</h2> | ||
+ | <p>- Ran gels to verify pET and PUC double digests</p> | ||
+ | <p>- Restriction digest of annealed oligo sequences with EcoRI and PstI</p> | ||
+ | <p>- Oligo PCR verification</p> | ||
+ | <p>- Digested pSB1C3 with EcoRI and PstI</p> | ||
+ | |||
+ | <h2>August 2</h2> | ||
+ | <p>- pUC19 and pET Vector were isolated and gel purified </p> | ||
+ | <p>- Miniprep of pUC19, pET Vector, and pSB1C3</p> | ||
+ | |||
+ | <h2>August 3</h2> | ||
+ | <p>- Ligated pUC19 and all the oligos</p> | ||
+ | |||
+ | <h2>August 5</h2> | ||
+ | <p>- PCR with new CDS7 was successful. Sung has “never seen a brighter 70 bp lane in [his] life”</p> | ||
+ | <p>- pUC19 – quantum dot synthesis oligos worked</p> | ||
+ | <p>- Miniprep, digest with EcoRI and PstI, and gel run of CDS7</p> | ||
+ | |||
+ | <h2>August 6</h2> | ||
+ | <p>- Gel extraction of pUC19 and pET Vector compatible CDS7 amplicon</p> | ||
+ | |||
+ | <h2>August 7</h2> | ||
+ | <p>- Inoculated oligo ligation cells in ampicillin media</p> | ||
+ | <p>- Digested and ran more of oligo ligations</p> | ||
+ | <p>- Inoculated more of the ligations </p> | ||
+ | <p>- Inoculated 6 CDS7 colonies</p> | ||
+ | |||
+ | <h2>August 8</h2> | ||
+ | <p>- Optimized PCR</p> | ||
+ | |||
+ | <h2>August 22</h2> | ||
+ | <p>- Oligo annealed </p> | ||
+ | <p>- Ethanol precipitation of DNA</p> | ||
+ | |||
+ | <h2>August 23</h2> | ||
+ | <p>- Precipitation of CDS7 and restriction digest with EcoRI and PstI</p> | ||
+ | <p>- Oligo annealed</p> | ||
+ | |||
+ | |||
+ | <h2>August 24</h2> | ||
+ | <p>- Dephosphorylated backbone and created ligation mixtures</p> | ||
+ | <p>- Annealed biobrick part</p> | ||
+ | <p>- Made master mix for biobrick backbone</p> | ||
+ | <p>- Oligo annealed</p> | ||
+ | <p>- Digested pUC19 with EcoR1 and PstI</p> | ||
+ | |||
+ | <h2>August 30</h2> | ||
+ | <p>- Attempted ligation of biobrick</p> | ||
+ | |||
+ | <h2>August 31</h2> | ||
+ | <p>- Tested previous day’s ligation in a gel and it failed</p> | ||
+ | |||
+ | <h2>September 1</h2> | ||
+ | <p>- pUC19 digest with EcoRI and PstI </p> | ||
+ | <p>- Gel purification of pUC19 digest</p> | ||
+ | <p>- Alcohol precipitated CDS7</p> | ||
+ | |||
+ | <h2>September 2</h2> | ||
+ | <p>- Received new primers</p> | ||
+ | <p>- Ran three PCR reactions, CDS7 forward with RBS, reverse, and control</p> | ||
+ | <p>- Ran 1/100 dilution of gel</p> | ||
+ | |||
+ | <h2>September 7</h2> | ||
+ | <p>- Another CDS7 PCR</p> | ||
+ | |||
+ | <h2>September 8</h2> | ||
+ | <p>- Restriction digest of J140 with EcoRI and PstI</p> | ||
+ | |||
+ | <h2>September 9</h2> | ||
+ | <p>- Ran gel of pET Vector, j140, pSB1C3, CDS7, and CDS7S</p> | ||
+ | |||
+ | <h2>September 13</h2> | ||
+ | <p>- Restriction digest of master mix with EcoR1 and Pst1</p> | ||
+ | <p>- Made a ligation mix</p> | ||
+ | <p>- Transformed with ligation mix</p> | ||
+ | |||
+ | <h2>September 14</h2> | ||
+ | <p>- pET Vector digest with Nco1 and BamH1</p> | ||
+ | <p>- CDS7 PCR</p> | ||
+ | <p>- CDS7 digest with EcoRI and PstI</p> | ||
+ | <p>- CDS7 digest with XbaI and PstI</p> | ||
+ | <p>- Gel verification and purification of pET Vector and CDS7</p> | ||
+ | <p>- Ligation of CDS7 EcoRI and PstI cut with pSB1C3</p> | ||
+ | |||
+ | |||
+ | <h2>September 20</h2> | ||
+ | <p>- VF and VR PCR verification</p> | ||
+ | <p>- Restriction digest on pSB1C3 with Pst1 and EcoR1</p> | ||
+ | <p>- Gel verification of digest</p> | ||
+ | |||
+ | <h2>September 21</h2> | ||
+ | <p>- Verified base pair numbers of primers and verified they work</p> | ||
+ | <p>- PCR of CDS7</p> | ||
+ | <p>- Gel purification of CDS7</p> | ||
+ | <p>- Digest CDS7 with EcoR1 and Pst1</p> | ||
+ | <p>- Phosphotased pSB1C3</p> | ||
+ | <p>- Made ligation mix with all the parts</p> | ||
+ | <p>- Transformed with the ligation mix</p> | ||
+ | |||
+ | <h2>September 23</h2> | ||
+ | <p>- Ligation mix from previous day failed</p> | ||
+ | <p>- Re-did everything from September 21st except this time made a new master mix with different PCR settings</p> | ||
+ | <p>- Ligated and transformed with the new master mix</p> | ||
+ | |||
+ | <h2>September 24</h2> | ||
+ | <p>- Grew colonies overnight from previous days ligation and transformation</p> | ||
+ | <p>- Grew out colonies with LB chloramphenicol </p> | ||
+ | <p>- Made minipreps of CDS7</p> | ||
+ | |||
+ | <h2>September 26</h2> | ||
+ | <p>- PCR a sequencing segment from plasmid with VF2 and VR</p> | ||
+ | <p>- Sent segment off for sequencing</p> | ||
+ | <p>- Reviewed other transformations</p> | ||
</div> | </div> | ||
</body> | </body> | ||
</html> | </html> |
Revision as of 00:34, 29 September 2011
Genspace Notebook
July 5
- Grew out BBa_k325219 and BBa_k325909 in media C.
- Restriction digest of BBa_k231000 and BBa_k231001 with EcoRI and PstI
July 15
- CDS7 isolation and ligation
July 19
- NcoI and BamHI digest of pET Vector
July 20
- Another attempt at NcoI and BamHI digest of pET Vector
July 21
- Gel Purified pET Vector Digest
- Grew out more pET vector
July 24
- Received new NcoI and BamHI enzymes
- pET Vector digest with new NcoI and new BamHI
- pET Vector digest with old NcoI and old BamHI
- pET Vector digest with new NcoI and another with old NcoI
- pET Vector digest with BamHI and anther with old BamHI
July 25
- Ran a gel with all pET Vector digests from July 24th
- Fermentas transformation on pET Vector
July 28
- CDS7 digest with NcoI and BamHI
- Prepared plates of kanamycin and pET Vector
- Oligo annealed with Silver Lab protocol
- Maxiprep using ~50mL CDS7 growth
- pET Vector inoculation
July 29
- pET Vector digest with BamHI and NcoI
- Gel purification of pET Vector digest
- Inoculated more pET Vector
- Inoculated C-media for transformations
- Miniprep of pET Vector
- Ligated 4 oligos
- Prepare transformation ready cells using C-media culture prepared the previous day
July 30
- Miniprep of pET Vector
- Digestion of pET Vector with NcoI and BamHI
- Oligo PCR
-Digest pUC19 with EcoRI and PstI
August 1
- Ran gels to verify pET and PUC double digests
- Restriction digest of annealed oligo sequences with EcoRI and PstI
- Oligo PCR verification
- Digested pSB1C3 with EcoRI and PstI
August 2
- pUC19 and pET Vector were isolated and gel purified
- Miniprep of pUC19, pET Vector, and pSB1C3
August 3
- Ligated pUC19 and all the oligos
August 5
- PCR with new CDS7 was successful. Sung has “never seen a brighter 70 bp lane in [his] life”
- pUC19 – quantum dot synthesis oligos worked
- Miniprep, digest with EcoRI and PstI, and gel run of CDS7
August 6
- Gel extraction of pUC19 and pET Vector compatible CDS7 amplicon
August 7
- Inoculated oligo ligation cells in ampicillin media
- Digested and ran more of oligo ligations
- Inoculated more of the ligations
- Inoculated 6 CDS7 colonies
August 8
- Optimized PCR
August 22
- Oligo annealed
- Ethanol precipitation of DNA
August 23
- Precipitation of CDS7 and restriction digest with EcoRI and PstI
- Oligo annealed
August 24
- Dephosphorylated backbone and created ligation mixtures
- Annealed biobrick part
- Made master mix for biobrick backbone
- Oligo annealed
- Digested pUC19 with EcoR1 and PstI
August 30
- Attempted ligation of biobrick
August 31
- Tested previous day’s ligation in a gel and it failed
September 1
- pUC19 digest with EcoRI and PstI
- Gel purification of pUC19 digest
- Alcohol precipitated CDS7
September 2
- Received new primers
- Ran three PCR reactions, CDS7 forward with RBS, reverse, and control
- Ran 1/100 dilution of gel
September 7
- Another CDS7 PCR
September 8
- Restriction digest of J140 with EcoRI and PstI
September 9
- Ran gel of pET Vector, j140, pSB1C3, CDS7, and CDS7S
September 13
- Restriction digest of master mix with EcoR1 and Pst1
- Made a ligation mix
- Transformed with ligation mix
September 14
- pET Vector digest with Nco1 and BamH1
- CDS7 PCR
- CDS7 digest with EcoRI and PstI
- CDS7 digest with XbaI and PstI
- Gel verification and purification of pET Vector and CDS7
- Ligation of CDS7 EcoRI and PstI cut with pSB1C3
September 20
- VF and VR PCR verification
- Restriction digest on pSB1C3 with Pst1 and EcoR1
- Gel verification of digest
September 21
- Verified base pair numbers of primers and verified they work
- PCR of CDS7
- Gel purification of CDS7
- Digest CDS7 with EcoR1 and Pst1
- Phosphotased pSB1C3
- Made ligation mix with all the parts
- Transformed with the ligation mix
September 23
- Ligation mix from previous day failed
- Re-did everything from September 21st except this time made a new master mix with different PCR settings
- Ligated and transformed with the new master mix
September 24
- Grew colonies overnight from previous days ligation and transformation
- Grew out colonies with LB chloramphenicol
- Made minipreps of CDS7
September 26
- PCR a sequencing segment from plasmid with VF2 and VR
- Sent segment off for sequencing
- Reviewed other transformations