SBB09Ntbk-Dirkvs: Difference between revisions
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==[[User:Dirk Van Swaay|Dirk]] 13:21, 18 February 2009 (EST)== | ==[[User:Dirk Van Swaay|Dirk]] 13:21, 18 February 2009 (EST)== | ||
Today, I set up the PCR reactions for the three parts. | Today, I set up the PCR reactions for the three parts. | ||
# Dilute oligonucleotides for construct 17 | |||
## Add to the tube a volume of ddH20 10x ul the molar amount of DNA | |||
## Mix and spin | |||
## Add 1 ul of oligo dilution to 9 ul of ddH20 | |||
## Mix and spin | |||
# Prepare the PCR reaction for construct 17 A (Restriction site removal) | |||
** 24 uL water | |||
** 3.3 uL Expand 10x Buffer 2 | |||
** 3.3 uL 10x dNTPs (2 mM in each; 0.2 mM final conc) | |||
** 1 uL Oligo Odvs007F (10uM) | |||
** 1 uL Oligo Odvs008rR (10uM) | |||
** 0.55 uL Expand Polymerase 1 | |||
## Run this PCR on program 55 | |||
# Prepare the PCR reaction for construct 17 A (Restriction site removal) | |||
** 24 uL water | |||
** 3.3 uL Expand 10x Buffer 2 | |||
** 3.3 uL 10x dNTPs (2 mM in each; 0.2 mM final conc) | |||
** 1 uL Oligo Odvs007R (10uM) | |||
** 1 uL Oligo Odvs008rF (10uM) | |||
** 0.55 uL Expand Polymerase 1 | |||
## Run this PCR on program 55 | |||
** 29 uL water | |||
** 5 uL Expand 10x Buffer 2 | |||
** 5 uL 10x dNTPs (2 mM in each; 0.2 mM final conc) | |||
** 5 uL Oligo 1 (100uM) | |||
** 5 uL Oligo 2 (100uM) | |||
** 0.75 uL Expand Polymerase 1 | |||
=== === | === === | ||
Revision as of 13:55, 18 February 2009
Dirk 13:21, 18 February 2009 (EST)
Today, I set up the PCR reactions for the three parts.
- Dilute oligonucleotides for construct 17
- Add to the tube a volume of ddH20 10x ul the molar amount of DNA
- Mix and spin
- Add 1 ul of oligo dilution to 9 ul of ddH20
- Mix and spin
- Prepare the PCR reaction for construct 17 A (Restriction site removal)
- 24 uL water
- 3.3 uL Expand 10x Buffer 2
- 3.3 uL 10x dNTPs (2 mM in each; 0.2 mM final conc)
- 1 uL Oligo Odvs007F (10uM)
- 1 uL Oligo Odvs008rR (10uM)
- 0.55 uL Expand Polymerase 1
- Run this PCR on program 55
- Prepare the PCR reaction for construct 17 A (Restriction site removal)
- 24 uL water
- 3.3 uL Expand 10x Buffer 2
- 3.3 uL 10x dNTPs (2 mM in each; 0.2 mM final conc)
- 1 uL Oligo Odvs007R (10uM)
- 1 uL Oligo Odvs008rF (10uM)
- 0.55 uL Expand Polymerase 1
- Run this PCR on program 55
- 29 uL water
- 5 uL Expand 10x Buffer 2
- 5 uL 10x dNTPs (2 mM in each; 0.2 mM final conc)
- 5 uL Oligo 1 (100uM)
- 5 uL Oligo 2 (100uM)
- 0.75 uL Expand Polymerase 1
to do
Dirk 00:39, 10 February 2009 (EST)
Carried out the oligos and construction file for the Cellulose Binding Knottin CBK (Bdvs003, or M10046).
- Construction file to be found here (M10046)
- Oligonucleotide sequences to be found here (M10046)
Dirk 00:17, 10 February 2009 (EST)
Carried out the oligos and construction file for the Silver-binding peptide AG4 (Bdvs002, or M10023).
- Construction file to be found here (M10023)
- Oligonucleotide sequences to be found here (M10023)
* Design Oligos and make construction file for Cellulse Binding Knottin (M10046)
Dirk 15:01, 9 February 2009 (EST)
Got the project ball on Wednesday last week. Assigned project 15954 - Beta Roll and Silver Peptide. Also assigned Cellulose Binding Knottin, from 37738. The project page can be found here (SBB09_15954).
Spent the lab time designing the oligos and writing the construction file for the Beta Roll (Bdvs001, or M10022).
- Construction file to be found here (M10022)
- Oligonucleotide sequences to be found here (M10022)
* Design oligos and make construction file for Silver Peptide (M10023) * Design Oligos and make construction file for Cellulse Binding Knottin (M10046)
