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Team:Hong Kong HKUST/notebook/mod2
From 2013.igem.org
(Difference between revisions)
| Line 301: | Line 301: | ||
<a href='#' class='head'>Week 4</a> | <a href='#' class='head'>Week 4</a> | ||
<div class='content' align="left"> | <div class='content' align="left"> | ||
| - | June 24 <br> | + | <br><b>June 24</b><br> |
Wet lab <br> | Wet lab <br> | ||
· Inoculation of pBlueScript KS(+) for training <br> | · Inoculation of pBlueScript KS(+) for training <br> | ||
| Line 309: | Line 309: | ||
Dry lab <br> | Dry lab <br> | ||
Protocols review <br> | Protocols review <br> | ||
| - | <br> | + | |
| - | June 25 <br> | + | <br><b> |
| + | June 25 </b><br> | ||
Wet lab <br> | Wet lab <br> | ||
· Plasmid extraction of pBlueScript KS(+) <br> | · Plasmid extraction of pBlueScript KS(+) <br> | ||
| Line 317: | Line 318: | ||
· Primers design for FABP1, PPARa, GRP78 <br> | · Primers design for FABP1, PPARa, GRP78 <br> | ||
· Informal meeting <br> | · Informal meeting <br> | ||
| - | <br> | + | |
| - | June 26 <br> | + | <br><b> |
| + | June 26 </b><br> | ||
Wet lab <br> | Wet lab <br> | ||
· Extraction of gDNA from HepG2 Cells for FABP1 and PPARa <br> | · Extraction of gDNA from HepG2 Cells for FABP1 and PPARa <br> | ||
· Transformation of pEGFP-N1 and BBa_K817002 (pFadBA) <br> | · Transformation of pEGFP-N1 and BBa_K817002 (pFadBA) <br> | ||
| - | <br> | + | |
| - | June 27 <br> | + | <br><b> |
| + | June 27 </b><br> | ||
Wet lab <br> | Wet lab <br> | ||
· Inoculation of pEGFP-N1 and BBa_K817002 (pFadBA) for plasmid extraction <br> | · Inoculation of pEGFP-N1 and BBa_K817002 (pFadBA) for plasmid extraction <br> | ||
| - | <br> | + | |
| - | June 28 <br> | + | <br><b> |
| + | June 28</b> <br> | ||
Wet lab <br> | Wet lab <br> | ||
· Extraction of pEGFP-N1 and BBa_K817002 (pFadBA) plasmids by miniprep <br> | · Extraction of pEGFP-N1 and BBa_K817002 (pFadBA) plasmids by miniprep <br> | ||
| Line 333: | Line 337: | ||
· Restriction of c(pFadBA) by EcoR1 and Pst1 <br> | · Restriction of c(pFadBA) by EcoR1 and Pst1 <br> | ||
· Ran 0.8% gel for pEGFP-N1 and 2% gel for BBa_K817002 (pFadBA) <br> | · Ran 0.8% gel for pEGFP-N1 and 2% gel for BBa_K817002 (pFadBA) <br> | ||
| - | <br> | + | |
| - | June 30 <br> | + | <br><b> |
| + | June 30 </b><br> | ||
Dry lab <br> | Dry lab <br> | ||
Experiment planning and protocols revision for next week work <br><br> | Experiment planning and protocols revision for next week work <br><br> | ||
| + | |||
</div> | </div> | ||
</div> | </div> | ||
Revision as of 14:58, 19 September 2013
June 2013
Week 4
June 24
Wet lab
· Inoculation of pBlueScript KS(+) for training
· Autoclave basic materials
· Preparing LB
· LB-Ampicillin plates poured Dry lab
Protocols review
June 25
Wet lab
· Plasmid extraction of pBlueScript KS(+)
· LB-Chloramphenicol plates poured
Dry lab
· Primers design for FABP1, PPARa, GRP78
· Informal meeting
June 26
Wet lab
· Extraction of gDNA from HepG2 Cells for FABP1 and PPARa
· Transformation of pEGFP-N1 and BBa_K817002 (pFadBA)
June 27
Wet lab
· Inoculation of pEGFP-N1 and BBa_K817002 (pFadBA) for plasmid extraction
June 28
Wet lab
· Extraction of pEGFP-N1 and BBa_K817002 (pFadBA) plasmids by miniprep
· Restriction of pEGFP-N1 Restriction digestion by Ase1 and bamH1 for PPARa; Ase1 and Xho1 for GRP78, Xba1 and BamH1 for FABP1, Pst1 and Not1; followed by gel check, extraction and purification
· Restriction of c(pFadBA) by EcoR1 and Pst1
· Ran 0.8% gel for pEGFP-N1 and 2% gel for BBa_K817002 (pFadBA)
June 30
Dry lab
Experiment planning and protocols revision for next week work
June 24
Wet lab
· Inoculation of pBlueScript KS(+) for training
· Autoclave basic materials
· Preparing LB
· LB-Ampicillin plates poured Dry lab
Protocols review
June 25
Wet lab
· Plasmid extraction of pBlueScript KS(+)
· LB-Chloramphenicol plates poured
Dry lab
· Primers design for FABP1, PPARa, GRP78
· Informal meeting
June 26
Wet lab
· Extraction of gDNA from HepG2 Cells for FABP1 and PPARa
· Transformation of pEGFP-N1 and BBa_K817002 (pFadBA)
June 27
Wet lab
· Inoculation of pEGFP-N1 and BBa_K817002 (pFadBA) for plasmid extraction
June 28
Wet lab
· Extraction of pEGFP-N1 and BBa_K817002 (pFadBA) plasmids by miniprep
· Restriction of pEGFP-N1 Restriction digestion by Ase1 and bamH1 for PPARa; Ase1 and Xho1 for GRP78, Xba1 and BamH1 for FABP1, Pst1 and Not1; followed by gel check, extraction and purification
· Restriction of c(pFadBA) by EcoR1 and Pst1
· Ran 0.8% gel for pEGFP-N1 and 2% gel for BBa_K817002 (pFadBA)
June 30
Dry lab
Experiment planning and protocols revision for next week work
July 2013
Week 1
July 2
Wet lab
· Digestion of pEGFP-N1 by Ase1 and bamH1 for PPARa; , Xba1 and BamH1 for FABP1
· LB-Chloramphenicol plates (the previous ones were found contaminated)
· Inoculation of pEGFP-N1 for plasmid extraction
· Transformation of pFadBA due chloramphenicol plates contamination
July 3
· Extraction of pEGFP-N1 plasmids by miniprep
· PCR for PPARa promoter amplification
· Digestion of pEGFP-N1 by Ase1 and bamH1 for PPARa; , Xba1 and BamH1 for FABP1
July 4
Wet lab
· Inoculation of BBa_K817002 pFadBA
· PCR for PPARa promoter cloning from gDNA
· PCR for FABP1 promoter cloning from gDNA
· Ran gel for digestion of pEGFP-N1 by Ase1 and bamH1 for PPARa; Xba1 and BamH1 for FABP1; Ase1 and Xhol1 for GRP78
July 5
Wet lab
· Extraction of BBa_K817002 (pFadBA)
· Inoculation of pEGFP-N1
· PCR for PPARa promoter cloning from gDNA
· PCR for FABP1 promoter cloning from gDNA
· Gel check, 0.8% gel for previously gDNA extraction
· Ran gel for PCR check of PPARa
· Ran gel for PCR chack of FABP1
· Poured new LB-Kanamycin plates
· Transformation of BBa_J176171
Dry lab
· Discussion about re-design constructions, possible change for BBa_J176171 as mammalian expression vector
· Primer redesign for PPARa and FABP1
July 2
Wet lab
· Digestion of pEGFP-N1 by Ase1 and bamH1 for PPARa; , Xba1 and BamH1 for FABP1
· LB-Chloramphenicol plates (the previous ones were found contaminated)
· Inoculation of pEGFP-N1 for plasmid extraction
· Transformation of pFadBA due chloramphenicol plates contamination
July 3
· Extraction of pEGFP-N1 plasmids by miniprep
· PCR for PPARa promoter amplification
· Digestion of pEGFP-N1 by Ase1 and bamH1 for PPARa; , Xba1 and BamH1 for FABP1
July 4
Wet lab
· Inoculation of BBa_K817002 pFadBA
· PCR for PPARa promoter cloning from gDNA
· PCR for FABP1 promoter cloning from gDNA
· Ran gel for digestion of pEGFP-N1 by Ase1 and bamH1 for PPARa; Xba1 and BamH1 for FABP1; Ase1 and Xhol1 for GRP78
July 5
Wet lab
· Extraction of BBa_K817002 (pFadBA)
· Inoculation of pEGFP-N1
· PCR for PPARa promoter cloning from gDNA
· PCR for FABP1 promoter cloning from gDNA
· Gel check, 0.8% gel for previously gDNA extraction
· Ran gel for PCR check of PPARa
· Ran gel for PCR chack of FABP1
· Poured new LB-Kanamycin plates
· Transformation of BBa_J176171
Dry lab
· Discussion about re-design constructions, possible change for BBa_J176171 as mammalian expression vector
· Primer redesign for PPARa and FABP1
Week 2
July 8
Wet lab
· gDNA extraction from HepG2 cells
· restriction of BBa_K817002 pFAdBA
· Gel check for gDNA extraction
· Inoculation of BBa_J176171
· Inoculation of BBa_K817002 pFadBA Inoculation of pEGFP-N1 for plasmid extraction
· New primers for PPARa and FABP1 arrival
· Ran gel for BBa_K817002 pFadBA restriction check, gel extraction and purification
July 9
Wet lab
· Extraction of BBa_J176171 by miniprep
· Extraction of BBa_K817002 pFadBA by miniprep
· Extraction of pEGFP-N1 by miniprep
· Restriction of pEGFP-N1 Restriction digestion by Ase1 and bamH1 for PPARa; Ase1 and Xho1 for GRP78, Xba1 and BamH1 for FABP1, Pst1 and Not1; followed by gel check, extraction and purification
· Restriction of BBa_J176171 for pFadBA Ase1 and BamH1; FABP1 by Xba1 and Not1
· Ran gel for pEGFP-N1 restriction products
· Ran gel for BBa_J176171 restriction products
· PCR PPARa and FABP1 promoters cloning from gDNA
· BBa_J52034 (FADR) transformation
July 10
Wet lab
· Ran gel for PCR check of PPARa and FAPB1
· PCR for FABP1, PCR replication
· BBa_J176171 vector dephosphorylation by antartic phosphatase
· Ligation of BBa_K817002 and pFadBA and BBa_J176171
· BBa_J52034 (FADR) inoculation
· Digestion for BBa_K817002 pFadBA extraction
Gel check for BBa_K817002 pFadBA extraction
July 11
Wet lab
· Gel purification for BBa_K817002 pFadBA extraction
· Gel check for FABP1 PCR product
· PCR clean-up
· BBa_J52034 (FADR) miniprep
· BBa_J52034 restriction by Pst1 HF and Not1 HF
· PCR for PPARa
Dry lab
Re-design constructions, now using BBa_J176171 as mammalian expression vector for all related constructions
July 12
Wet lab
· Digestion of FABP1 PCR product
· Ran gel for PCR check of PPARa
· Digest pEGFP-n1 for BBa_J52034 FADR
· Ran gel for pEGFP-n1 for BBa_J52034 FADR followed by gel purification
· BBa_J176171 vector dephosphorylation by Antarctic phosphatase for FABP1 and FADR
Dry Lab
Primers design for pCMV cloning for FADR expression
July 8
Wet lab
· gDNA extraction from HepG2 cells
· restriction of BBa_K817002 pFAdBA
· Gel check for gDNA extraction
· Inoculation of BBa_J176171
· Inoculation of BBa_K817002 pFadBA Inoculation of pEGFP-N1 for plasmid extraction
· New primers for PPARa and FABP1 arrival
· Ran gel for BBa_K817002 pFadBA restriction check, gel extraction and purification
July 9
Wet lab
· Extraction of BBa_J176171 by miniprep
· Extraction of BBa_K817002 pFadBA by miniprep
· Extraction of pEGFP-N1 by miniprep
· Restriction of pEGFP-N1 Restriction digestion by Ase1 and bamH1 for PPARa; Ase1 and Xho1 for GRP78, Xba1 and BamH1 for FABP1, Pst1 and Not1; followed by gel check, extraction and purification
· Restriction of BBa_J176171 for pFadBA Ase1 and BamH1; FABP1 by Xba1 and Not1
· Ran gel for pEGFP-N1 restriction products
· Ran gel for BBa_J176171 restriction products
· PCR PPARa and FABP1 promoters cloning from gDNA
· BBa_J52034 (FADR) transformation
July 10
Wet lab
· Ran gel for PCR check of PPARa and FAPB1
· PCR for FABP1, PCR replication
· BBa_J176171 vector dephosphorylation by antartic phosphatase
· Ligation of BBa_K817002 and pFadBA and BBa_J176171
· BBa_J52034 (FADR) inoculation
· Digestion for BBa_K817002 pFadBA extraction
Gel check for BBa_K817002 pFadBA extraction
July 11
Wet lab
· Gel purification for BBa_K817002 pFadBA extraction
· Gel check for FABP1 PCR product
· PCR clean-up
· BBa_J52034 (FADR) miniprep
· BBa_J52034 restriction by Pst1 HF and Not1 HF
· PCR for PPARa
Dry lab
Re-design constructions, now using BBa_J176171 as mammalian expression vector for all related constructions
July 12
Wet lab
· Digestion of FABP1 PCR product
· Ran gel for PCR check of PPARa
· Digest pEGFP-n1 for BBa_J52034 FADR
· Ran gel for pEGFP-n1 for BBa_J52034 FADR followed by gel purification
· BBa_J176171 vector dephosphorylation by Antarctic phosphatase for FABP1 and FADR
Dry Lab
Primers design for pCMV cloning for FADR expression
Week 3
Content 1
Week 4
Content 1
Week 5
Content 1
