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From 2008.igem.org

Contents 1 Test Competent E. coli DH5a 1.1 Methods 1.1.1 Materials 1.1.2 Procedure 1.2 Results 1.3 Discussion

Test Competent E. coli DH5a

• Test our competent cells against Callahan competent cells

Methods

Materials

• Transported 50uL of competent cells from Callahan Lab on ice
• 100uL batch 1 competent cells made at OD₆₀₀ ~0.6 (split into two 50 uL aliquots 4-18a,4-18b)
• 100uL batch 2 competent cells made at OD₆₀₀ ~0.3 (split into two 50 uL aliquots 2-17a,2-17b)

Procedure

1. Thaw plasmid on ice
2. Add 1uL plasmid pBluescript
3. Incubate plasmid+competent cells on ice for 30 minutes
4. Heat shock 60 sec at 42C
5. Add 250 μl SOC
6. Incubate at 37 C for 1 hour in 2 ml centrifuge tubes rotated using 2ml centrifuge tubes for transformation and regrowth works well because the small volumes flow well when rotated, increasing aeration.
7. Plate 20 uL dilutions 1/1 1/10, 1/100 to test competency
   1. For plasmids (pSB1AC3, pSB1AT3) which are chloramphenicol and tetracycline resistant, growing for 2 hours yields many more colonies
   2. Ampicillin and kanamycin resistant transformants appear to do fine with 1 hour growth

Results

• 
  

Discussion

• What was learned and how to do future experiments differently.

Insanity is doing the same thing over and over again and expecting different results. - Albert Einstein

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