Heat shock transformation
[DOC File]BIOTECH Laboratory Activity:
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Heat shock: Brief exposure to 42oC “thermal imbalance” ... so a transformation method with a better efficiency of transformation is required. To prepare cells which are more competent (i.e., have a higher efficiency of transformation) must start with bacteria actively dividing in mid-log phase broth culture.
Heat shock transformation History, Methods, Applications ...
Heat shock 42oC for 45 sec. Incubate on ice for 2 min . Add in LB medium 500 ul to each tube . Shaker in 37oC for 20 min . 50 ul plate on agar plates . Detailed procedure (Later afternoon, duration ~ 2 hours) Get the ice bucket and keep DH5a (-80C) chemical competent cells in cie for 5-10 min for them to thaw.
[DOC File]Session #4 Labs 2 & 5: Rapid colony transformation
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Step 1: Transformation (Two days before maxi-prep. Starting at late afternoon. Duration: ~2 hours) 1. Get the ice bucket and keep DH5a chemical competent cells (from the -80癈e freezer) in ice for 5~10 minutes for them to thaw. 2. Turn on the heat shock machine to low setting at 42°C.
[DOC File]Transformation of plasmid into Top 10 E
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Heat shock the transformation mix (competent cells) at 42 ˚C for 45 seconds then put on ice for a couple of min. Add 500 µl of LB and incubate in warm room (37 ˚C) with shaking for 1 hour. Spin down cells and resuspend in 100 µl LB. Plate everything on an LB-Amp plate and incubate plate overnight in warm room. Pick single colonies and screen.
[DOC File]Quick ligations using Roche Rapid DNA Ligation Kit
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TRANSFORMATION LAB. Discussion. Please read carefully: Start by recalling your experiment and the purpose of the various steps (heat shock, recovery in shaking incubator) and reagents (CaCl2 , LB, ice, plasmids). (If necessary, do some research on the details of the protocol we used---“Google it”.
[DOC File]TRANSFORMATION LAB
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Bacterial Transformation. I. Objectives. After completing this activity, you should be able to: Describe the process of transformation and outline the steps involved in this technique. Explain the purpose of the use of calcium chloride and the heat shock step. Explain why antibiotic-resistance genes are …
[DOC File]Maxi-prep Overall Procedure (by Steven Chang)
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Yeast Transformation * inoculate 5-10 ml culture with single colony and grow ON * measure OD600nm and dilute to OD600nm = 0.3 in 50 ml * grow for 2-4 hrs ... * heat shock at 42 °C for 7 min * spin, resuspend cells in 1 ml 1 x TE * spin, resuspend cells in 0.5 ml 1 x TE * plate 100 - 200 µl / plate and incubate at 30 °C.
[DOC File]Transformation Procedure - University of California, San Diego
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Leave the tube on ice for 30 minutes. Meanwhile, thaw the SOC medium (-20˚C). Fill the heat shock wells with ¾ water. Heat shock the tube @ 42˚C for exactly 40 seconds. Leave the tubes on ice for 2 minutes. Add 200ul of RT SOC medium to the tube. Shake and incubate the tube at 37˚C for 1 …
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