Team:Vilnius-Lithuania/Labjournal

Parts construction

Written by: IR
Performed by: ŠT, IR, MB, IS.

About

pLac+cI.
P10 and P11 plasmids are used. pLac (P11) and repressor cI (P10) ligation.

Date	Materials	Experiment
06 30	P10 P11	P10 and P11 transformation into DH5α.
07 01		Bacteria inoculation (4 minipreps).
07 09	P10 P11	Plasmids’ restriction (3 ug each). P10 with EcoRI and XBal (+FastAp). P11 with EcoRI and SpeI. Extraction from gel L10 and L11 are produced. Conc.: L11 – 2,4 ng/uL.

Conclusions and Results:

07 14 – all of the colonies are successful, except third colony.

Parts construction

Part: P15.1 (BBa_)

Written by: IR
Performed by: ŠT, IR, MB, IS.

About

Cas3 with mutated restriction sites
L7+L15. Concentration - 261 ng/μL

Date	Materials	Experiment
06 30	P13 P7	P13 and P7 transformation into DH5α.
07 01		Transformants are inoculated (4 minipreps each).
07 02		Plasmid extraction. Concentrations: P7 – 174,1 ng/μL and 166 ng/μL. P13 - 216 ng/μL and 131, 5 ng/μL.
07 13	P13	Cas3 mutagenesis (with ST1/2, ST3/4, ST5/6 primers).
07 16	Mutated P13	Cas3 plasmids extraction.
07 21-23	Mutated P13	Restriction with EcoRI, XbaI and PstI.
07 27	Mutated P13	Cas3 mutagenesis.
07 29	P7	Restriction with EcoRI and SpeI. L7 is produced.
07 31	P13
08 02	Mutated P13	Mutated Cas3 restriction with EcoRI, XbaI and PstI.
08 05	Mutated P13	Cas3 mutagenesis.
08 07	Mutated P13	Mutated Cas3 restriction with PstI. Mutagenesis was successful. L15 is produced.
08 11	L7 L15	L7 and L15 ligation with pSB1C3 (EcoRI+XBal+FastAp). P15.1 is produced.
	P15.1	Sequencing.

Conclusions and Results:

07 21 – Mutagenesis was unsuccessful.
08 02 – successfully mutagenized EcoRI and XbaI sites.
08 07 – successful mutagenesis confirmed (XbaI and EcoRI sites are mutated).
08 07 – Successful mutagenesis (PstI, XbaI and EcoRI sites are mutated).

Parts construction

Part: P14.2 (BBa_K1773006)

Written by: IR
Performed by: ŠT, IR, MB, IS.

About

pLac+cI.
P10 and P11 plasmids are used. pLac (P11) and repressor cI (P10) ligation.

Date	Materials	Experiment
06 30	P10 P11	P10 and P11 transformation into DH5α.
07 01		Bacteria inoculation (4 minipreps).
07 09	P10 P11	Plasmids’ restriction (3 ug each). P10 with EcoRI and XBal (+FastAp). P11 with EcoRI and SpeI. Extraction from gel L10 and L11 are produced. Conc.: L11 – 2,4 ng/uL.

Conclusions and Results:

07 14 – all of the colonies are successful, except third colony.

Parts construction

Part: P14.2 (BBa_K1773006)

Written by: IR
Performed by: ŠT, IR, MB, IS.

About

pLac+cI.
P10 and P11 plasmids are used. pLac (P11) and repressor cI (P10) ligation.

Date	Materials	Experiment
06 30	P10 P11	P10 and P11 transformation into DH5α.
07 01		Bacteria inoculation (4 minipreps).
07 09	P10 P11	Plasmids’ restriction (3 ug each). P10 with EcoRI and XBal (+FastAp). P11 with EcoRI and SpeI. Extraction from gel L10 and L11 are produced. Conc.: L11 – 2,4 ng/uL.

Conclusions and Results:

07 14 – all of the colonies are successful, except third colony.

Parts construction

Part: P14.2 (BBa_K1773006)

Written by: IR
Performed by: ŠT, IR, MB, IS.

About

pLac+cI.
P10 and P11 plasmids are used. pLac (P11) and repressor cI (P10) ligation.

Date	Materials	Experiment
06 30	P10 P11	P10 and P11 transformation into DH5α.
07 01		Bacteria inoculation (4 minipreps).
07 09	P10 P11	Plasmids’ restriction (3 ug each). P10 with EcoRI and XBal (+FastAp). P11 with EcoRI and SpeI. Extraction from gel L10 and L11 are produced. Conc.: L11 – 2,4 ng/uL.

Conclusions and Results:

07 14 – all of the colonies are successful, except third colony.