Using a schematic diagram, summarize the following steps in preparing competent cells for transformation:Inoculate a single colony of E. coli into 5 ml LB broth and incubate overnight at 37°C with moderate shaking (250 rpm).Add 200 μl of the culture into 50 ml LB broth and incubate overnight at 37°C with moderate shaking (250 rpm) to an OD600 = 1.3 to 1.5.Aliquot culture into five 15-ml pre-chilled, conical tubes. Leave tube on ice 5 to 10 min.Centrifuge cells 7 min at 1,600 × g (3,000 rpm), 4°C.Pour off supernatant and resuspend each pellet in 10 ml ice-cold CaCl2 solution (50 mM CaCl2), perform resuspension very gently, and keep on ice.Centrifuge cells 5 min at 1,100 × g (2,500 rpm), 4°C. Discard supernatant and resuspend each pellet in 10 ml ice-cold CaCl2 solution. Keep resuspended on ice for 30 min.Centrifuge cells 5 min at 1,100 × g, 4°C. Discard supernatant and resuspend each pellet in 10 ml ice-cold CaCl2 solution.Dispense cells (250 μl) into pre-chilled, sterile conical tubes and freeze immediately at −70°C.

prepare competent cells for transformation ;- Add 20 µL of each ligation reaction or 1 ng of uncut…

Answered: Using a schematic diagram, summarize…

Human Anatomy & Physiology (11th Edition)

11th Edition

ISBN:9780134580999

Author:Elaine N. Marieb, Katja N. Hoehn

Publisher:Elaine N. Marieb, Katja N. Hoehn

Chapter1: The Human Body: An Orientation

Section: Chapter Questions

Problem 1RQ: The correct sequence of levels forming the structural hierarchy is A. (a) organ, organ system,...

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Using a schematic diagram, summarize the following steps in preparing competent cells for transformation:

Inoculate a single colony of E. coli into 5 ml LB broth and incubate overnight at 37°C with moderate shaking (250 rpm).
Add 200 μl of the culture into 50 ml LB broth and incubate overnight at 37°C with moderate shaking (250 rpm) to an OD600 = 1.3 to 1.5.
Aliquot culture into five 15-ml pre-chilled, conical tubes. Leave tube on ice 5 to 10 min.
Centrifuge cells 7 min at 1,600 × g (3,000 rpm), 4°C.
Pour off supernatant and resuspend each pellet in 10 ml ice-cold CaCl2 solution (50 mM CaCl2), perform resuspension very gently, and keep on ice.
Centrifuge cells 5 min at 1,100 × g (2,500 rpm), 4°C. Discard supernatant and resuspend each pellet in 10 ml ice-cold CaCl2 solution. Keep resuspended on ice for 30 min.
Centrifuge cells 5 min at 1,100 × g, 4°C. Discard supernatant and resuspend each pellet in 10 ml ice-cold CaCl2 solution.
Dispense cells (250 μl) into pre-chilled, sterile conical tubes and freeze immediately at −70°C.

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