(reporterkram rein) |
(reporter verbessert) |
||
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<td><p><h6>Reporter System</h6> | <td><p><h6>Reporter System</h6> | ||
<b>BBa_K1976002:</b><br> | <b>BBa_K1976002:</b><br> | ||
− | Clones were screened with colony PCR using VF2 and VR oligonucleotides. Plasmid DNA was isolated from positive clones. < | + | Clones were screened with colony PCR using VF2 and VR oligonucleotides. Plasmid DNA was isolated from positive clones. <br> |
− | |||
− | + | <h6>Metabolic burden</h6> | |
<b>BBa_K1976001:</b><br> | <b>BBa_K1976001:</b><br> | ||
The BioBrick BBa_I11020 was synthezised with a LVA‑tag and with B0034 as a ribosome binding site by IDT.</p> | The BioBrick BBa_I11020 was synthezised with a LVA‑tag and with B0034 as a ribosome binding site by IDT.</p> | ||
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<td><p><h6>Reporter System</h6> | <td><p><h6>Reporter System</h6> | ||
<b>BBa_K1976002:</b><br> | <b>BBa_K1976002:</b><br> | ||
− | Isolated Plasmid DNA Sequence was verified by Sanger Sequencing. < | + | Isolated Plasmid DNA Sequence was verified by Sanger Sequencing. <br> |
− | + | <h6>Metabolic burden</h6> | |
− | + | ||
<b>BBa_K1976001:</b><br> | <b>BBa_K1976001:</b><br> | ||
The LVA‑tag was deleted by PCR with the primer Del_LVA_fw and Del_LVA_rev.</p> | The LVA‑tag was deleted by PCR with the primer Del_LVA_fw and Del_LVA_rev.</p> | ||
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<td><p><h6>Reporter System</h6> | <td><p><h6>Reporter System</h6> | ||
<b>BBa_K1976003:</b><br> | <b>BBa_K1976003:</b><br> | ||
− | The mVenus-LVA-pSB1C3 plasmid was cut with <i>EcoRI</i> and <i>PstI</i> and ligated into a T7-RBS-pSB1A3 plasmid which was cut with <i>SpeI</i> and <i>PstI.</i> < | + | The mVenus-LVA-pSB1C3 plasmid was cut with <i>EcoRI</i> and <i>PstI</i> and ligated into a T7-RBS-pSB1A3 plasmid which was cut with <i>SpeI</i> and <i>PstI.</i> <br> |
− | + | ||
− | + | <h6>Metabolic burden</h6> | |
<b>BBa_K1976000:</b><br> | <b>BBa_K1976000:</b><br> | ||
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<td><p><h6>Reporter System</h6> | <td><p><h6>Reporter System</h6> | ||
<b>BBa_K1976003:</b><br> | <b>BBa_K1976003:</b><br> | ||
− | The ligation product was transformed into <i>E.coli</i> Top 10 by heat shock transformation and the cells were spread out on a AMP agar plate. < | + | The ligation product was transformed into <i>E.coli</i> Top 10 by heat shock transformation and the cells were spread out on a AMP agar plate. <br> |
− | + | ||
− | + | <h6>Metabolic burden</h6> | |
<b>BBa_K1976000:</b> | <b>BBa_K1976000:</b> | ||
Line 422: | Line 418: | ||
<td><p><h6>Reporter System</h6> | <td><p><h6>Reporter System</h6> | ||
<b>BBa_K1976003:</b><br> | <b>BBa_K1976003:</b><br> | ||
− | Clones were screened with colony PCR using VF2 and VR oligonucleotides. Plasmid DNA was isolated from positive clones . < | + | Clones were screened with colony PCR using VF2 and VR oligonucleotides. Plasmid DNA was isolated from positive clones . <br> |
− | + | ||
− | + | <h6>Metabolic burden</h6> | |
<b>BBa_K1976000:</b><br> | <b>BBa_K1976000:</b><br> | ||
Line 448: | Line 444: | ||
<td><p><h6>Reporter System</h6> | <td><p><h6>Reporter System</h6> | ||
<b>BBa_K1976003:</b><br> | <b>BBa_K1976003:</b><br> | ||
− | The isolated Plasmid DNA was verified by Sanger sequencing.< | + | The isolated Plasmid DNA was verified by Sanger sequencing.<br> |
− | + | <h6>Metabolic burden</h6> | |
− | + | ||
<b>BBa_K1976000:</b><br> | <b>BBa_K1976000:</b><br> | ||
Line 470: | Line 465: | ||
<td><p><h6>Reporter System</h6> | <td><p><h6>Reporter System</h6> | ||
<b>BBa_K1976003:</b><br> | <b>BBa_K1976003:</b><br> | ||
− | Subsequently the psB1A3 vector containing brick was cut with <i>EcoRI</i> and <i>PstI</i> and ligated into a pSB1C3 vector using T4‑Ligase.< | + | Subsequently the psB1A3 vector containing brick was cut with <i>EcoRI</i> and <i>PstI</i> and ligated into a pSB1C3 vector using T4‑Ligase.<br> |
− | + | <h6>Metabolic burden</h6> | |
− | + | ||
<b>BBa_K1976000:</b><br> | <b>BBa_K1976000:</b><br> | ||
Line 496: | Line 490: | ||
<div class="labWeek"><h5>Lab Week 27: <div class="week_date"> October 17 - October 23</div></h5></div></td> | <div class="labWeek"><h5>Lab Week 27: <div class="week_date"> October 17 - October 23</div></h5></div></td> | ||
<td><img class="labjournalIcon" src="https://static.igem.org/mediawiki/2016/b/b5/T--TU_Darmstadt--iconLab3.png"/></td> | <td><img class="labjournalIcon" src="https://static.igem.org/mediawiki/2016/b/b5/T--TU_Darmstadt--iconLab3.png"/></td> | ||
− | + | <p><h6>Metabolic burden</h6> | |
Plasmid curing could not be performed due to lack of time. | Plasmid curing could not be performed due to lack of time. | ||
</p> | </p> |
Revision as of 17:28, 19 October 2016
![iGEM TU Darmstadt 2016](https://static.igem.org/mediawiki/2016/8/83/T--TU_Darmstadt--titel.png)
NOTEBOOK
Labjournal 2016
For half a year we were working in the wet lab , developed a pipetting robot
and simulated the molecular dynamics of our proteins
.
As well we would like to highlight certain social events
which helped us to further improve our team spirit.
Lab Week 1: |
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Hier könnte Ihr labjournal stehen! |
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robotics | |
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modeling | |
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social |
Lab Week 2: |
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Hier könnte Ihr labjournal stehen! |
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robotics | |
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modeling | |
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social |
Lab Week 3: |
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Hier könnte Ihr labjournal stehen! |
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robotics | |
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modeling | |
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social |
Lab Week 4: |
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Hier könnte Ihr labjournal stehen! |
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robotics | |
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modeling | |
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social |
Lab Week 5: |
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Hier könnte Ihr labjournal stehen! |
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robotics | |
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modeling | |
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social |
Lab Week 6: |
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Hier könnte Ihr labjournal stehen! |
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robotics | |
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social |
Lab Week 7: |
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Hier könnte Ihr labjournal stehen! |
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robotics | |
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modeling | |
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social |
Lab Week 8: |
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Hier könnte Ihr labjournal stehen! |
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robotics | |
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modeling | |
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social |
Lab Week 9: |
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Hier könnte Ihr labjournal stehen! |
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robotics | |
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modeling | |
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social |
Lab Week 10: |
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Hier könnte Ihr labjournal stehen! |
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robotics | |
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modeling | |
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social |
Lab Week 11: |
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Hier könnte Ihr labjournal stehen! |
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robotics | |
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modeling | |
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social |
Lab Week 12: |
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Hier könnte Ihr labjournal stehen! |
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robotics | |
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modeling | |
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social |
Lab Week 13: |
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Hier könnte Ihr labjournal stehen! |
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robotics | |
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modeling | |
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social |
Lab Week 14: |
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Hier könnte Ihr labjournal stehen! |
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robotics | |
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modeling | |
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social |
Lab Week 15: |
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Hier könnte Ihr labjournal stehen! |
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robotics | |
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modeling | |
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social |
Lab Week 16: |
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Hier könnte Ihr labjournal stehen! |
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robotics | |
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modeling | |
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social |
Lab Week 17: |
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Reporter SystemBBa_K1976002:The mVenus-LVA-gene was synthesized by idt and amplified through PCR using the SP-amplify-fw-Rep1 (fw) and SP-amplify-rev-Rep2 (REV) oligonucleotides. |
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robotics | |
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modeling | |
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social |
Lab Week 18: |
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Reporter SystemBBa_K1976002:The PCR product was verified by agarose gel electrophoresis and subsequently purified. The mVenus amplicon was cut with EcoRI and PstI and ligated into a pSB1C3 vector using T4-ligase. |
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robotics | |
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modeling | |
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social |
Lab Week 19: |
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Reporter SystemBBa_K1976002:The ligation product was transformed into E.coli Top 10 by heat shock transformation and the cells were spread out on a CMP agar plate. |
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robotics | |
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modeling | |
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social |
Lab Week 20: |
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Reporter SystemBBa_K1976002:Clones were screened with colony PCR using VF2 and VR oligonucleotides. Plasmid DNA was isolated from positive clones. Metabolic burdenBBa_K1976001:The BioBrick BBa_I11020 was synthezised with a LVA‑tag and with B0034 as a ribosome binding site by IDT. |
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robotics | |
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modeling | |
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social |
Lab Week 21: |
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Reporter SystemBBa_K1976002:Isolated Plasmid DNA Sequence was verified by Sanger Sequencing. Metabolic burdenBBa_K1976001:The LVA‑tag was deleted by PCR with the primer Del_LVA_fw and Del_LVA_rev. |
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robotics | |
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modeling | |
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social |
Lab Week 22: |
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Reporter SystemBBa_K1976003:The mVenus-LVA-pSB1C3 plasmid was cut with EcoRI and PstI and ligated into a T7-RBS-pSB1A3 plasmid which was cut with SpeI and PstI. Metabolic burdenBBa_K1976000:The BioBrick BBa_E0240 was cut with the restriction enzymes XbaI and PstI. BBa_J61002 was cut with SpeI and PstI. The two restriction products were ligated by standard T4‑Ligation protocol. BBa_K1976001: B0034‑BBa_I11020 was cut with XbaI and PstI. BBa_J23101 was cut with SpeI and PstI. Both fragments were ligated using the T4‑Ligase by standard protocol. |
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robotics | |
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modeling | |
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social |
Lab Week 23: |
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Reporter SystemBBa_K1976003:The ligation product was transformed into E.coli Top 10 by heat shock transformation and the cells were spread out on a AMP agar plate. Metabolic burdenBBa_K1976000:After plasmid preparation, the plasmid was cut with XbaI and PstI. We had the attp‑site BBa_11023 flanked by the two additional bidirectional transcription terminators BBa_1001 synthesized by IDT. The attp‑site of BBa_11023 was not the original λ‑attp‑site. Instead the sequence corresponded to the attP2‑site used in the Gateway ® cloning system. The construct was cut with EcoRI and SpeI. The pSB1C3 backbone was cut with EcoRI and PstI. Together with the previously cut BBa_J61002‑BBa_E0240 construct the synthesized BBa_1001‑BBa_11023‑BBa_1001 construct was ligated in the cut pSB1C3. BBa_K1976001: The ligation product was transformed into E. coli Top 10 via heatshock. |
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robotics | |
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modeling | |
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social |
Lab Week 24: |
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Reporter SystemBBa_K1976003:Clones were screened with colony PCR using VF2 and VR oligonucleotides. Plasmid DNA was isolated from positive clones . Metabolic burdenBBa_K1976000:The attP2‑site was mutated to λ‑attp via Quick‑Change‑PCR, using the following primers: QC_attp_fw and QC_attp_rev (for more details see our primerlist). BBa_K1976001: To check whether BBa_I11020 was expressed properly a SDS‑Page was made. |
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robotics | |
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modeling | |
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social |
Lab Week 25: |
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Reporter SystemBBa_K1976003:The isolated Plasmid DNA was verified by Sanger sequencing. Metabolic burdenBBa_K1976000:The LVA‑tag was added by PCR with the overhang primer LVA_GFP_fw and LVA_GFP_rev. |
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Lab Week 26: |
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Reporter SystemBBa_K1976003:Subsequently the psB1A3 vector containing brick was cut with EcoRI and PstI and ligated into a pSB1C3 vector using T4‑Ligase. Metabolic burdenBBa_K1976000:For plasmid curing the final plasmid construct and the low copy backbone pSB4A5 were cut with EcoRI and PstI and then ligated. BBa_K1976001: For plasmid curing the final plasmid construct and the low copy vector pSB4K5 were cut with EcoRI and PstI and then ligated. BBa_K1976000 and BBa_K1976001 were cotransformed into E. coli Top 10. To check whether the genomic integration was successful a cPCR with attb_fw and VR primers was made. |
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modeling | |
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social |
Lab Week 27: |
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