Difference between revisions of "20.109(F16): TA notes for M1"

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For Hsinhwa
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Module 1 Measuring Genomic Instability
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===M1D1===
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Number of groups: 9, so prepare 11 aliquots
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Per group, aliquot
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*50 mL 1x PBS
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*in TC room, 15 mL of “TK6 media”
  
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Week 1: September 12, 2016
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===M1D2===
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Day 1 (T/W)
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Per group, aliquot
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Per group:
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*in main lab
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• Aliquot 50 mL 1x PBS per group (label: PBS)
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**10 mL 1x PBS  
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• In TC room, aliquot 15 mL of “TK6 media” (label: TK6 media)
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**2 eppendorf tubes of 0.3% LMP agarose (in eppendorf water bath)
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*and separately, in the TC hood,  
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**10 mL 1x PBS
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**25 mL TK6 media
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**2 eppendorf tubes of 0.3% LMP agarose (in water bath '''at 43 &deg;C''')
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**2x 15 mL of TK6 cells at 500,000 cells/mL
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<br>
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Set up hoods in TC room with, per group:
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*4 binder clips
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*1 glass slide
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*1 bottomless 96-well plate
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*1 razor blade
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*Pasteur pipets
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*sharp jar
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*pipet aid
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*pen
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*team sticker to identify glass slides
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*printout of today's protocol
  
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Day 2 (R/F)
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===M1D3===
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Per group:
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Per group, aliquot:
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• Aliquot 10 mL 1x PBS (label: PBS)
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*1% LMP agarose
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• And separately, in the TC hood, aliquot 10 mL 1x PBS (label: PBS)
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*H2O2 and MMS
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Leslie & Maxine will prepare:
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*Alkyline lysis solution
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• Prepare 4 x 2 mL tubes of 0.3% LMP agarose, per group
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*Alkyline electrophoresis buffer
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*Neutralization buffer
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*SYBR Gold buffer
  
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Week 2: September 19, 2016
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===M1D4===
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Day 3 (T/W)
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Per group, aliquot:
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Leslie & Maxine will prepare:
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*15 mL 1x PBS
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• 1% LMP agarose
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*H2O2  
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• H2O2 and MMS
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*Alkyline lysis solution
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• Alkyline lysis solution
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*Warm M059J/K culture medium
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• Alkyline electrophoresis buffer
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• Neutralization buffer
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• SYBR Gold buffer
+
  
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Week 3: September 26, 2016
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===M1D5===
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Day 4 (T/W)
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Per group, aliquot in TC room:
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Per group:
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*5 mL of 0.1% gelatin
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• Aliquot 15 mL 1x PBS (label: PBS)
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*10 mL of PBS  
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Leslie & Maxine will prepare:
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*3 mL of 1X trypsin
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• H2O2
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*15 mL of M059J/K media
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• Alkyline lysis solution
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*M059J/K in T25 flasks
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• Warm culture medium
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===M1D6===
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Per group, aliquot in TC room:
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*10 mL of 1x TBS
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*500 uL of 10% BSA, frozen stock in -20C
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*Triton X-100 at front bench
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*Primary antibodies
  
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Day 5 (R/F)
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===M1D7===
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Per group:
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Per group, aliquot
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In TC room for all aliquots, leave at 4C:
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*5 mL of 1x TBS
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• Aliquot 5 mL of 0.1% gelatin (label: Gelatin)
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• Aliquot 10 mL of PBS (label: PBS)
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• Aliquot 3 mL of 1X trypsin (label: Trypsin)
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• Aliquot 15 mL of M059J/K media (label: Media)
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Leslie & Maxine will prepare:
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• M059J/K in T25 flasks
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−
 
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Week 4: October 3, 2016
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Day 6 (T/W)
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Per group:
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• Aliquot 10 mL of 1x TBS (label: TBS)
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• Aliquot 500 uL of 10% BSA, frozen stock in -20C (label: 10% BSA)
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Leslie & Maxine will prepare:
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• 10% BSA and freeze at -20C
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• Triton X-100 at front bench
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• Primary antibodies
+
−
 
+
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Day 7 (R/F)
+
−
Per group:
+
−
• Aliquot 5 mL of 1x TBS (label: TBS)
+

Revision as of 13:32, 16 September 2016

20.109(F16): Laboratory Fundamentals of Biological Engineering

Engelward PNAS 2006.png

Schedule Fall 2016        Announcements        Assignments        Homework        Communication
       1. Measuring Genomic Instability        2. Manipulating Metabolism        3. Engineering Biomaterials              

M1D1

Per group, aliquot

  • 50 mL 1x PBS
  • in TC room, 15 mL of “TK6 media”

M1D2

Per group, aliquot

  • in main lab
    • 10 mL 1x PBS
    • 2 eppendorf tubes of 0.3% LMP agarose (in eppendorf water bath)
  • and separately, in the TC hood,
    • 10 mL 1x PBS
    • 25 mL TK6 media
    • 2 eppendorf tubes of 0.3% LMP agarose (in water bath at 43 °C)
    • 2x 15 mL of TK6 cells at 500,000 cells/mL


Set up hoods in TC room with, per group:

  • 4 binder clips
  • 1 glass slide
  • 1 bottomless 96-well plate
  • 1 razor blade
  • Pasteur pipets
  • sharp jar
  • pipet aid
  • pen
  • team sticker to identify glass slides
  • printout of today's protocol

M1D3

Per group, aliquot:

  • 1% LMP agarose
  • H2O2 and MMS
  • Alkyline lysis solution
  • Alkyline electrophoresis buffer
  • Neutralization buffer
  • SYBR Gold buffer

M1D4

Per group, aliquot:

  • 15 mL 1x PBS
  • H2O2
  • Alkyline lysis solution
  • Warm M059J/K culture medium

M1D5

Per group, aliquot in TC room:

  • 5 mL of 0.1% gelatin
  • 10 mL of PBS
  • 3 mL of 1X trypsin
  • 15 mL of M059J/K media
  • M059J/K in T25 flasks

M1D6

Per group, aliquot in TC room:

  • 10 mL of 1x TBS
  • 500 uL of 10% BSA, frozen stock in -20C
  • Triton X-100 at front bench
  • Primary antibodies

M1D7

Per group, aliquot

  • 5 mL of 1x TBS
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