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Journal
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Génétique
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A High-Throughput UAS-cDNA/ORF Plasmid Library Construction Using CRISPRmass
/
Large-Scale Transformation of UAS-cDNA/ORF Plasmids from CRISPR-Based Modular Assembly into
E. coli
Journal JoVE
Génétique
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Journal JoVE
Génétique
Large-Scale Transformation of UAS-cDNA/ORF Plasmids from CRISPR-Based Modular Assembly into
E. coli
Large-Scale Transformation of UAS-cDNA/ORF Plasmids from CRISPR-Based Modular Assembly into
E. coli
A High-Throughput UAS-cDNA/ORF Plasmid Library Construction Using CRISPRmass
DOI:
10.3791/201692-v
•
01:46 min
•
May 17, 2024
•
Si Xu*
1,2
,
Mengyu Chen*
1,2
,
Guang Chen
3
,
Si Luo
,
Wen Xue
2
,
Xuelian Liu
2
,
Jiwu Wang
2,3
,
Ping Wei
1
Clinical Research Institute, The Affiliated Nanhua Hospital, Hengyang Medical School
,
University of South China
,
2
Department of Neurology, The Affiliated Nanhua Hospital, Hengyang Medical School
,
University of South China
,
3
Department of Clinical Laboratory, The Affiliated Nanhua Hospital, Hengyang Medical School
,
University of South China
,
4
Shanghai 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
Tags
Transformation Of UAS-cDNA/ORF Plasmids From CRISPR-Based Modular Assembly Into E. Coli:1. Pre-chill 10 μL 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 μL Of Competent Cells To Each Pre-chilled 1.5 ML Tube.4. Add 1 μL 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 μL 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
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Author Spotlight: Simplifying Genome-Wide Plasmid Library Construction Using CRISPRmass
Large-Scale Construction of UAS-cDNA/ORF Plasmids by CRISPR-Based Modular Assembly (CRISPRmass)
Large-Scale Transformation of UAS-cDNA/ORF Plasmids from CRISPR-Based Modular Assembly into
E. coli
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