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Team:Yale/MeetingNotes
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The phsABC plasmid worked! We have metal precipitation! Woooo! | The phsABC plasmid worked! We have metal precipitation! Woooo! | ||
| - | 1. Cloning issues | + | 1. Cloning issues |
| - | Did the double digestion of the terminator work? | + | *Did the double digestion of the terminator work? |
| - | * The EcoRI was old and might have failed | + | ** The EcoRI was old and might have failed, check each enzyme on a gel |
*SpeI not very efficient in EcoRI buffer, will try to incubate longer | *SpeI not very efficient in EcoRI buffer, will try to incubate longer | ||
* The ligation didn't work, but there are lots of ways to keep trying. This is a great way for people to come help during the week. | * The ligation didn't work, but there are lots of ways to keep trying. This is a great way for people to come help during the week. | ||
| - | 2.Copper precipitation assay | + | 2. Copper precipitation assay |
*Pick a concentration from the cell growth assay, like 1 mM Cu? | *Pick a concentration from the cell growth assay, like 1 mM Cu? | ||
*Grow shaking, at time points spin out the cells and measure the absorbance | *Grow shaking, at time points spin out the cells and measure the absorbance | ||
Latest revision as of 21:24, 8 July 2010
July 8
The phsABC plasmid worked! We have metal precipitation! Woooo!
1. Cloning issues
- Did the double digestion of the terminator work?
- The EcoRI was old and might have failed, check each enzyme on a gel
- SpeI not very efficient in EcoRI buffer, will try to incubate longer
- The ligation didn't work, but there are lots of ways to keep trying. This is a great way for people to come help during the week.
2. Copper precipitation assay
- Pick a concentration from the cell growth assay, like 1 mM Cu?
- Grow shaking, at time points spin out the cells and measure the absorbance
- this is assaying for loss of copper
3. Does phsABC confer copper resistance?
- 96-well plate assay
- same as before, but with the phsABC plasmid
- Since Leah has this assay dialed-in, this is a great job for a part-time helper to run and analyze!
4. Cu resistance
- will be shipping in a day or two
- pRJ1004 (Cu efflux protein) (Jon will email to follow up)
- massive plasmid, no antibiotic resistance
- Get together next week to request several plasmids (Tuesday morning at 10 am).
- pRJ1004 (Cu efflux protein) (Jon will email to follow up)
5. Email Kara about the mold (Jon)
6. Ways to get students involved (Dan email the group)
- Ask when they're available, delegate a task
- make sure someone is there to help them
