Team:Yale/MeetingNotes

From 2010.igem.org

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The phsABC plasmid worked!  We have metal precipitation!  Woooo!
The phsABC plasmid worked!  We have metal precipitation!  Woooo!
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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.
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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).

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

July 15