Berkeley LBL/Methods
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===Constructs=== | ===Constructs=== | ||
| - | The T7 expression vector pET3a was used for the subcloning of the genes for magnesium-chelatase. The following constructs were built by inserting | + | The T7 expression vector pET3a was used for the subcloning of the genes for magnesium-chelatase. The following constructs were built by inserting various fragments genes from the three organisms into pET3a. (Note: The pET3a vector contains a ribosome binding site downstream of the T7 promoter region; this rbs would not be indicated below) |
| - | Rhodobacter sphaeroides | + | '''Magnesium-Chelatase'''<br> |
| - | Synechocystis sp. | + | Rhodobacter sphaeroides<br> |
| - | Heliobacillus mobilis | + | pET3a-bchH<br> |
| + | pET3a-bchI<br> | ||
| + | pET3a-bchD<br> | ||
| + | pET3a-bchHID<br> | ||
| + | Synechocystis sp.<br> | ||
| + | pET3a-chlH<br> | ||
| + | pET3a-chlI<br> | ||
| + | pET3a-chlD<br> | ||
| + | pET3a-chlHID | ||
| + | |||
| + | Heliobacillus mobilis<br> | ||
==Protocols== | ==Protocols== | ||
Revision as of 21:27, 26 October 2007
| Home | Project Description | Methods | Notebook | Results and Discussion | Resources |
Experimental
Constructs
The T7 expression vector pET3a was used for the subcloning of the genes for magnesium-chelatase. The following constructs were built by inserting various fragments genes from the three organisms into pET3a. (Note: The pET3a vector contains a ribosome binding site downstream of the T7 promoter region; this rbs would not be indicated below)
Magnesium-Chelatase
Rhodobacter sphaeroides
pET3a-bchH
pET3a-bchI
pET3a-bchD
pET3a-bchHID
Synechocystis sp.
pET3a-chlH
pET3a-chlI
pET3a-chlD
pET3a-chlHID
Heliobacillus mobilis
Protocols
PCR (Using Phusion Polymerase)
PCR (Using TaKaRa Ex Taq Polymerase)
Digestion for PCR Product or Miniprepped DNA
KCM Competent Cell Transformation
Electroporation Transformation
