JoVE Logo
Faculty Resource Center

Sign In

Large-Scale Transformation of UAS-cDNA/ORF Plasmids from CRISPR-Based Modular Assembly into E. coli

18 Views

01:46 min

May 17th, 2024

DOI :

10.3791/201692-v

May 17th, 2024

18 Views

1Clinical Research Institute, The Affiliated Nanhua Hospital, Hengyang Medical School, University of South China, 2Department of Neurology, The Affiliated Nanhua Hospital, Hengyang Medical School, University of South China, 3Department of Clinical Laboratory, The Affiliated Nanhua Hospital, Hengyang Medical School, University of South China, 4Shanghai Diabetes Institute, Shanghai Key Laboratory of Diabetes Mellitus, Shanghai Clinical Center for Diabetes, Shanghai Sixth People's Hospital Affiliated to Shanghai Jiao Tong University School of Medicine
* These authors contributed equally

Explore More Videos

Transformation Of UAS cDNA ORF Plasmids From CRISPR Based Modular Assembly Into E Coli 1 Pre chill 10 L Tips And 1 5 ML Tubes In An Aluminum Cooling Block On Ice 2 Thaw Competent E Coli Cells On Ice For 5 Minutes 3 Add 10 L Of Competent Cells To Each Pre chilled 1 5 ML Tube 4 Add 1 L Of The UAS CDNA ORF Plasmid Products To Each Tube 5 Gently Swirl The Tubes To Mix And Incubate On Ice For 30 Minutes 6 Heat Shock The Cells At 42 C For 1 Minute 7 Immediately Chill The Tubes On Ice For 2 Minutes 8 Add 100 L Of Pre warmed 37 C SOC Medium To Each Tube 9 Incubate The Tubes In A Shaking Incubator At 250 RPM And 37 C For 1 Hour 10 Warm The LB Plates Containing The Appropriate Antibiotic To 37 C 11 Spread The Cells Onto The Warm Plates And Incubate At 37 C Overnight

From the series

A High-Throughput UAS-cDNA/ORF Plasmid Library Construction Using CRISPRmass
JoVE Logo

Privacy

Terms of Use

Policies

Research

Education

ABOUT JoVE

Copyright © 2024 MyJoVE Corporation. All rights reserved