"
Team:CAU China/J Sep
From 2013.igem.org
 |
 |
 |
 |
 |
|
 |
 |
|
 |
 |
|
SeptemberSeptember 1stLab meeting Discussion for experiment and wiki September 2nd Wet lab PCR for ta0841 and Nadh1 amplification Gel electrophoresis of PCR product Gel purification of PCR product Nanoview: the concentration is OK. \(^o^)/YEH~ Digestion of pET-28a(+) by BamH1-HF and Sal1-HF for ta0841; Not1-HF and BamH1-HF for Nadh1 Nanoview: the concentration is not high (⊙_⊙?) Digestion of ta0841 with BamH1-HF and Sal1-HF Digestion of Nadh1 with Not1-HF and BamH1-HF Column-purification of digested gene Nanoview: the concentration is OK O(∩_∩)O~ Ligation of ta0841 with pET-28a(+) Ligation of Nadh1 with pET-28a(+) Dry lab Building enzyme kinetic equations to describe the enzyme catalysis reactions September 3rdWet lab Transformation of ligated pET-28a(+)-ta0841 (3rd) and pET-28a(+)-Nadh1 (7st) Colony PCR for transformation product identification Gel check for colony PCR Result:positive results! ( ⊙o⊙ ) Small inoculation of pET-28a(+)- (3rd) and pET-28a(+)-Nadh1 (7st) for restriction anlysis Dry lab Building enzyme kinetic equations to describe the enzyme catalysis reactions September 4stWet lab Plasmid extraction of pET-28a(+)-ta0841 (3rd) and pET-28a(+)-Nadh1 (7st) Restriction analysis for identification Result:positive results!!! ~\(≧▽≦)/~ Bravo~ 
September 5thWet lab DNA sequencing for pET-28a(+)-ta0841 (3rd) and pET-28a(+)-Nadh1 (7st) Result:positive! positive! positive! ~\(≧▽≦)/~ Dry lab September 6thWet lab Digestion PSB1C3 backbone by PstI+EcoRI and gel check Gel purification of digested PSB1C3 Dry lab Design primers for biobricks September 8thLab meeting Discussion for experiment and T-shirt September 9thWet lab Primers for biobricks arrive Digestion of Sadh2with EcoR1 and Pst1 Digestion of Nadh1 with EcoR1 and Pst1 Column-purification of digested gene Nanoview: the concentration is OK O(∩_∩)O~ Ligation of Sadh2 with pET-28a(+) Ligation of Nadh1 with pET-28a(+) Dry lab September 10thWet lab Transformation of ligated PSB1C3- Sadh2 (1st) andPSB1C3-Nadh1 (1st) Result: no colony! ( ⊙o⊙ ) Dry lab September 11thTransformation of ligated PSB1C3- Sadh2 (2st) andPSB1C3-Nadh1 (2st) Result: no colony! o(︶︿︶)o Analysis for the bad result Dry lab September 12thWet lab Digestion of Sadh2 with EcoR1 and Pst1 Digestion of Nadh1 with EcoR1 and Pst1 Column-purification of digested gene Nanoview: the concentration is high O(∩_∩)O~ Ligation of Sadh2 with pET-28a(+) Ligation of Nadh1 with pET-28a(+) Dry lab September 13thWet lab Transformation of ligated PSB1C3- Sadh2 (3rd) and PSB1C3-Nadh1 (3rd) Result: look! The plate of Sadh2 has colony! Yeh~ Small inoculation of PSB1C3-Sadh2 (3rd) Dry lab September 14thWet lab Plasmid extraction of PSB1C3-Sadh2 (3rd) for identification Result:positive results!!! ~\(≧▽≦)/~ Bravo~ Dry lab September 15thLab meeting Presentation rehearsing September 16thWet lab Error-pone PCR of Sadh2, Nadh1 and ta0841 for random mutagenesis Mailing our biobrick Plasmid maximum extraction(alkaline lysis method) Gel electrophoresis of Error-pone PCR product Gel purification of Error-pone PCR product Transformation of pET-28a(+)-Sadh2 into BL21(Rosetta) Dry lab September 17thWet lab Inoculation for pET-28a(+)-Sadh2 Protein induction with 0.6mM IPTG,16℃,over night Dry lab September 18thCell lysis Affinity chromatography with a Nickel column Purification by gel-filtration chromatography using the Superdex 200 High Performance column SDS-PAGE for protein induced assay Result:our protein has been induced! \(^o^)/
  
Dry lab September 19thWet lab Digestion of Error-Pone Sadh2and ta0841with Bam1-HF and Sal1-HF Digestion of Error-Pone Nadh1 with BamH1-HF and Not1-HF Column-purification of digested gene Nanoview: the concentration is high O(∩_∩)O~ Ligation of purification products with pET-28a(+) Transformation of pET-28a(+)-Nadh1 and pET-28a(+)-ta0841 into BL21(Rosetta) Dry lab Mathematical modeling Information about the Judging Forms can be found in our Model September 20thWet lab Transformation of conjunction into BL21 competent cells Inoculation for pET-28a(+)-ta0841 and pET-28a(+)-Nadh1 Protein induction with 0.6mM IPTG,16℃,over night Dry lab September 21stCell lysis Affinity chromatography with a Nickel column Purification by gel-filtration chromatography using the Superdex 200 High Performance column SDS-PAGE for protein induced assay Result: ta0841 has been induced,but Nadh1 isn’t.
    |
