Team:HokkaidoU Japan/Notebook/H Stem/Construction

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Notebook

Lab Documents

H-Stem Construction

Start
mini-Prep
- Sun Aug 31
1C3 Vector
PCR
- Sun Aug 31, Mon Sep 1, Wed Sep 3, Thu Sep 4
Plac (plasmid) with 100up_F & 200dn_R, Ptet-B0034-mRFP-dT (plasmid) with XhoI-RBS & NcoI-mRFP (anti-sense), double-Terminator (plasmid) with 100up & 200dn
Digestion
- Sun Aug 31, Mon Sep 1, Wed Sep 3, Thu Sep 4
Plac with SpeI, anti-sense with NcoI, anti-sense with XhoI, dT with XbaI
Primer Annealing
- Sun Aug 31, Thu Sep 4
stem_upstream_F & stem_upstream_R, stem_downstream_F & stem_downstream_R
Ligation
- Fri Sep 5
Plac & stem-up & as-up, as-dn & stem-dn & dT
PCR
- Fri Sep 5
Plac-stem-as with 100up_F & XhoI_RBS_R, as-stem-dT with NcoI_mRFP_F & 200dn_R
PCR
- Mon Sep 8, Tue Sep 9, Tue Sep 9
Plac-stem-as & as-stem-dT with 100up_F & 200dn_R
Digestion
- Wed Sep 10
Plac-stem-as-stem-dT with EcoRI & PstI, 1C3 with EcoRI & PstI
Ligation
- Wed Sep 10
Plac-stem-as-stem-dT & 1C3
Transformation
- Thu Sep 11
Plac-stem-as-stem-dT on 1C3
Colony PCR
- Fri Sep 12, Sat Sep 13, Sat Sep 13
Plac-stem-as-stem-dT on 1C3 with stem_up_F & stem_d_R / 100up_F & 200dn_R / EX_F & PS_R
Complete!