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Sensor: Week 8 July 5-8
From 2011.igem.org
Contents |
Tuesday, July 5
cI Repressor Test Assembly, Day 27
Sequencing Results: The following constructs were sent to sequencing to verify their DNA sequences:
| Construct | Sequencing Result |
|---|---|
| J23113 + B0034 + C0051 | Good |
| KOR + K648000 | Good |
Clone OR and CRO
| Protocol | Part Involved with Protocol | |
|---|---|---|
| Restriction Digest | Inserts using XbaI and PstI: | OR CRO |
| Vector using XbaI and PstI: | K3 | |
| Ligation | K3 + OR K3 + CRO | |
| Transformation/Plating | The above ligations were transformed into Escherichia coli cells and plated onto kanamycin resistant plates. | |
Wednesday, July 6
cI Repressor Test Assembly, Day 28
The sensor group attempted to place two ribosome binding sites behind the cI repressor.
| Protocol | Part Involved with Protocol | |
|---|---|---|
| Insert PCR | B0033 B0032 | |
| Restriction Digest | Inserts using EcoRI and SpeI: | B0032 B0033 |
| Vector using EcoRI and XbaI: | C0051 | |
| Ligation | C0051 + B0032 C0051 + B0033 | |
| Transformation/Plating | The above ligations were transformed into Escherichia coli cells and plated onto ampicillin resistant plates. | |
Clone OR and CRO, Take 2
No colonies grew on the kanamycin resistant plates from 7/5, so the cloning was attempted again.
| Protocol | Part Involved with Protocol | |
|---|---|---|
| Restriction Digest | Inserts using XbaI and PstI: | OR CRO |
| Vector using XbaI and PstI: | K3 | |
| Ligation | K3 + OR K3 + CRO | |
| Transformation/Plating | The above ligations were transformed into Escherichia coli cells and plated onto kanamycin resistant plates. | |
Thursday, July 7
cI Repressor Test Assembly, Day 29
The sensor group verified their assemblies from 7/6 through agarose gel electrophoresis. The C0051+B0033 looked good, but the C0051+B0032 did not.
Friday, July 8
The C0051+B0033 and K3+CRO constructs were sent to the sequencing center. The C0051+B0033 DNA sequence came back positive, but the K3+CRO did not, so this cloning attempt must be done again.
Since the sequencing results came back positive, freezer stocks were made containing J23113+B0034+C0051 and KOR+K648000.
Colonies of the following constructs were grown up:
KCRO
KOR
B0032+C0051
B0033+C0051
cI Repressor Test Assembly, Day 30
| Protocol | Part Involved with Protocol | |
|---|---|---|
| Insert PCR | J23113 + B0031 + C0051 J23113 + B0034 + C0051 J2313 + B0030 + C0051 | |
| Restriction Digest | Inserts using XbaI and PstI: | J23113 + B0031 + C0051 J23113 + B0034 + C0051 J2313 + B0030 + C0051 |
| Vector using SpeI and PstI: | OR | |
| Ligation | OR + K648000 + J23113 + B0031 + C0051 OR + K648000 + J23113 + B0034 + C0051 OR + K648000 + J23113 + B0030 + C0051 | |
| Transformation/Plating | The above ligations were transformed into Escherichia coli cells and plated onto kanamycin resistant plates. | |
