Team:Nevada/Notebook

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(→'Team Nevada Notebook')
(→Team Nevada Notebook)
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'''Week of May 2-8:'''
'''Week of May 2-8:'''
 +
 +
* ''Elaine''
 +
** Ran 0.8% agarose gel of EcoRI digest
 +
** Made LB/KAN plates
 +
** Spread for colonies
 +
** Did miniprep for pBIB liquid cultures
'''Week of May 9-15:'''
'''Week of May 9-15:'''
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** Ran 0.8% agarose gel of pBIB post-phenol:chloroform cleanup.
** Ran 0.8% agarose gel of pBIB post-phenol:chloroform cleanup.
** Did minipreps on additional pBIB liquid cultures.
** Did minipreps on additional pBIB liquid cultures.
 +
 +
* ''Elaine''
 +
** Did XbaI and EcoRI digest of pBIB
 +
** Ran 2 0.8% gels of each digest
 +
** Did a miniprep and a Phenol:chloroform clean-up
 +
** Ran 0.8% gel of the XbaI and EcoRI digest with the uncut pBIB
'''Week of May 16-22:'''
'''Week of May 16-22:'''
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* ''Randy Pares and Vidim Gladwell''
* ''Randy Pares and Vidim Gladwell''
** Designed primers pBIB-RB-F, pBIB-RB-R, NOS 3'-Foward Nos3' reverse for sequencing the pBIB plant plasmid.
** Designed primers pBIB-RB-F, pBIB-RB-R, NOS 3'-Foward Nos3' reverse for sequencing the pBIB plant plasmid.
 +
 +
* ''Elaine''
 +
** Made LB/KAN plates
 +
** Made 50mg/ml stock of KAN
 +
** Made 1X TAE buffer
'''Week of May 23-29:'''
'''Week of May 23-29:'''
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** EcoRI digest of uncut sample 3
** EcoRI digest of uncut sample 3
** Prepared 5 mM dNTP stock
** Prepared 5 mM dNTP stock
 +
 +
* ''Elaine''
 +
** Incubated 50 mL liquid culture of ''E. coli'' with pBIB (sample 4 & sample 5)
 +
** Qiaprep miniprep of sample 4 & sample 5 according to manufacturer's modified protocol for large/low-copy plasmids
 +
** Nanodrop of DNA recovery of miniprepped sample 4 & sample 5
'''Week of May 30-June 5:'''
'''Week of May 30-June 5:'''
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** Klenow reactions of pBIB-pool and pBIB-maxi
** Klenow reactions of pBIB-pool and pBIB-maxi
** Made glycerol stocks of pBIB samples 1-5
** Made glycerol stocks of pBIB samples 1-5
 +
 +
* ''Elaine''
 +
** EcoRI digest of uncut pBIB sample 4 and 5
 +
** HinD3 digest of uncut sample 4 & sample 5 as a positive control
 +
** Ran 0.8% gels of samples 1-5 to see if EcoRI digest was successful
 +
** Ethanol precipitation of the EcoRI digests of sample 4 & sample 5
 +
** Nanodrop resulted to a low DNA content
 +
** Worked with Bryson for the EcoRI digest of the 5µg of pBIB maxi-prep
 +
** Ran 0.8% agarose gel of the pBIB maxi-prep EcoRI digest
 +
** Modified all protocols of the Binary vector
'''Week of June 6-12:'''
'''Week of June 6-12:'''
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** Ran agarose gel of pBIB cut and uncut, pBIB# uncut, pBIB# cut with EcoRI and pBIB# cut with HinDIII to confirm the absence of EcoRI site in pBIB#
** Ran agarose gel of pBIB cut and uncut, pBIB# uncut, pBIB# cut with EcoRI and pBIB# cut with HinDIII to confirm the absence of EcoRI site in pBIB#
** Single-colony streaked pBIB# 2 on a fresh LB-Kan plate
** Single-colony streaked pBIB# 2 on a fresh LB-Kan plate
 +
 +
* ''Elaine''
 +
** Worked with Chris to incubate 30 liquid cell cultures
 +
** Ran 0.8% gels of all 30 samples
'''Week of June 20-26:'''
'''Week of June 20-26:'''

Revision as of 18:58, 22 June 2010

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Team Nevada Notebook

Week of April 11-17:

  • Bryson, Michael, Senny, Tyler
    • Made tobacco cell (NT-1) media in Dr. Shintani's lab

Week of April 18-24:

  • Bryson
    • EcoRI digest of pBIB
    • Made Na acetate buffer

Week of April 25-May 1:

  • Bryson
    • Ran agarose gel of EcoRI digest

Week of May 2-8:

  • Elaine
    • Ran 0.8% agarose gel of EcoRI digest
    • Made LB/KAN plates
    • Spread for colonies
    • Did miniprep for pBIB liquid cultures

Week of May 9-15:

  • Bryson
    • Ran 0.8% agarose gel of pBIB post-phenol:chloroform cleanup.
    • Did minipreps on additional pBIB liquid cultures.
  • Elaine
    • Did XbaI and EcoRI digest of pBIB
    • Ran 2 0.8% gels of each digest
    • Did a miniprep and a Phenol:chloroform clean-up
    • Ran 0.8% gel of the XbaI and EcoRI digest with the uncut pBIB

Week of May 16-22:

  • Bryson
    • Klenow reactions of EcoRI digests
    • Phenol:chloroform cleanup of pBIB prior to ligation
    • Blunt-end ligation of klenowed pBIB
  • Randy Pares and Vidim Gladwell
    • Designed primers pBIB-RB-F, pBIB-RB-R, NOS 3'-Foward Nos3' reverse for sequencing the pBIB plant plasmid.
  • Elaine
    • Made LB/KAN plates
    • Made 50mg/ml stock of KAN
    • Made 1X TAE buffer

Week of May 23-29:

  • Bryson
    • Incubated 50 mL liquid culture of E. coli with pBIB (samples 3).
    • Miniprepped sample 3, using modified protocol for large/low-copy plasmids.
    • EcoRI digest of uncut sample 3
    • Prepared 5 mM dNTP stock
  • Elaine
    • Incubated 50 mL liquid culture of E. coli with pBIB (sample 4 & sample 5)
    • Qiaprep miniprep of sample 4 & sample 5 according to manufacturer's modified protocol for large/low-copy plasmids
    • Nanodrop of DNA recovery of miniprepped sample 4 & sample 5

Week of May 30-June 5:

  • Bryson
    • HinD3 digest of uncut sample 3
    • Ran 08% gels of samples 1-5 to verify complete digestion by EcoRI
    • Pooled uncut samples 3, 4 and 5 (pBIB-pool)
    • Recieved 5 µg of pBIB from Randy and Vadim's maxi-prep (pBIB-maxi)
    • EcoRI digest of pBIB-pool and pBIB-maxi
    • Ran 0.8% agarose gel of EcoRI digests
    • Klenow reactions of pBIB-pool and pBIB-maxi
    • Made glycerol stocks of pBIB samples 1-5
  • Elaine
    • EcoRI digest of uncut pBIB sample 4 and 5
    • HinD3 digest of uncut sample 4 & sample 5 as a positive control
    • Ran 0.8% gels of samples 1-5 to see if EcoRI digest was successful
    • Ethanol precipitation of the EcoRI digests of sample 4 & sample 5
    • Nanodrop resulted to a low DNA content
    • Worked with Bryson for the EcoRI digest of the 5µg of pBIB maxi-prep
    • Ran 0.8% agarose gel of the pBIB maxi-prep EcoRI digest
    • Modified all protocols of the Binary vector

Week of June 6-12:

  • Bryson
    • Ligation reactions for pBIB-pool and pBIB-maxi
    • Digested pBIB-pool and pBIB-maxi again with EcoRI to linearize any unmodified pBIB
    • Transformed Top-10 Cells with modified pBIB (designated pBIB#)
      • Obtained two colonies after overnight incubation
    • Line-streaked the two colonies (pBIB# 1 and pBIB# 2) on an LB-Kan plate

Week of June 13-19:

  • Bryson
    • Prepared liquid cultures of pBIB# 1 and pBIB# 2
    • Miniprepped liquid cultures of pBIB#
    • EcoRI and HinDIII digests of pBIB#
    • Ran agarose gel of pBIB cut and uncut, pBIB# uncut, pBIB# cut with EcoRI and pBIB# cut with HinDIII to confirm the absence of EcoRI site in pBIB#
    • Single-colony streaked pBIB# 2 on a fresh LB-Kan plate
  • Elaine
    • Worked with Chris to incubate 30 liquid cell cultures
    • Ran 0.8% gels of all 30 samples

Week of June 20-26:

  • Bryson
    • Selected 3 colonies from pBIB# 2 (2-1, 2-2, 2-3) and streaked on a fresh LB-Kan plate
    • Started 20 mL liquid cultures of 2-1, 2-2, and 2-3
    • Miniprepped samples
    • EcoRI and XbaI digests of samples to confirm that the EcoRI was gone and 2-1, 2-2, 2-3 arose from the same colony
    • 0.8% agarose gel of digests

Week of June 27-July 3:

Week of July 4-10:

Week of July 11-17:

Week of July 18-24:

Week of July 25-31:

Week of August 1-7:

Week of August 8-14:

Week of August 15-21:

Week of August 22-28:

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