"
August/10 August 2009
From 2009.igem.org
1.Min prep
Result of Nanodrop concentration check
(sample No.) (concentration) [http://partsregistry.org/wiki/index.php?title=Part:BBa_R0062 1-6O] 35.2 ng/uL [http://partsregistry.org/wiki/index.php?title=Part:BBa_I14017 1-18L] 41.8 ng/uL [http://partsregistry.org/wiki/index.php?title=Part:BBa_R0077 1-10K] 39.1 ng/uL
2.make competent cells(with CaCl2)
3.digestion and ligation
digestion with restriction enzyme
| Vector1 | Insert1 | ||
|---|---|---|---|
| [http://partsregistry.org/wiki/index.php?title=Part:BBa_B0015 1-23L] | 12 | [http://partsregistry.org/wiki/index.php?title=Part:BBa_K118003 2-16H] | 6 |
| EcoRI | 1 | EcoRI | 1 |
| SpeI | 1 | XbaI | 1 |
| No.2 Buffer | 2 | No.2 | 2 |
| dH2O | 4 | dH2O | 10 |
| total | 20uL | total | 20uL |
| Vector2 | Insert2 | ||
|---|---|---|---|
| [http://partsregistry.org/wiki/index.php?title=Part:BBa_B0015 1-23L] | 12 | 3-18O | 5 |
| EcoRI | 1 | EcoRI | 1 |
| SpeI | 1 | XbaI | 1 |
| No.2 Buffer | 2 | No.2 | 2 |
| dH2O | 4 | dH2O | 11 |
| total | 20uL | total | 20uL |
↓
vector1 insert1 vector2 insert2 ladder
failure
digestion with restriction enzyme insert : E+S vector : E+x
4.Transformation
to get new plasmid
[http://partsregistry.org/wiki/index.php?title=Part:BBa_B0015 1-23L] A [http://partsregistry.org/wiki/index.php?title=Part:BBa_J23100 1-18C] A [http://partsregistry.org/wiki/index.php?title=Part:BBa_I15009 1-20F] K [http://partsregistry.org/wiki/index.php?title=Part:BBa_C0070 1-12H] K [http://partsregistry.org/wiki/index.php?title=Part:BBa_R0040 1-6I] A [http://partsregistry.org/wiki/index.php?title=Part:BBa_K091101 2-6P] A [http://partsregistry.org/wiki/index.php?title=Part:BBa_I719005 1-15N] A used '8/10 competent cell
