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利用 CRISPR/Cas9 基因编辑研究细胞因子依赖性造血细胞中突变钙的致癌活性
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JoVE Journal
Cancer Research
Using CRISPR/Cas9 Gene Editing to Investigate the Oncogenic Activity of Mutant Calreticulin in Cytokine Dependent Hematopoietic Cells
Please note that all translations are automatically generated.
Click here for the English version.
利用 CRISPR/Cas9 基因编辑研究细胞因子依赖性造血细胞中突变钙的致癌活性
DOI:
10.3791/56726-v
•
10:21 min
•
January 05, 2018
•
Nouran S. Abdelfattah
,
Ann Mullally
2,3
1
Division of Hematology, Department of Medicine
,
Brigham and Women’s Hospital, Harvard Medical School
,
2
Broad Institute
,
3
Dana-Farber Cancer Institute
,
Harvard Medical School
Chapters
00:05
Title
00:44
Cloning sgRNA Oligos
01:08
Digestion of LentiGuide-puroVector
02:04
Ligation of Annealed Oligos into Digested Backbone
03:02
Generation of Cell Lines Stably Expressing spCas9
05:40
Reporter Assay for Cas9 Activity
06:10
Spinfection of sgRNAs into Cas9 Expressing Cells
06:31
Ba/F3 Cellular Transformation and Positive Selection using m-IL3 Withdrawal
07:23
Results: Use of CRISPR/Cas9 Tool and Ba/F3 Cellular Transformation to Study Oncogenic Potential of Mutated Calreticulin
09:02
Conclusion
Summary
Automatic Translation
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Automatic Translation
有针对性的基因编辑使用 CRISPR/Cas9 极大地促进了对基因的生物功能的理解。在这里, 我们利用 CRISPR/Cas9 的方法, 模型钙突变的细胞因子依赖性造血细胞, 以研究其致癌活动。
Tags
CRISPR/Cas9
Gene Editing
Calreticulin
Oncogenic Activity
Cytokine Dependent Hematopoietic Cells
Hematological Malignancies
LentiGuide-Puro Vector
BsmB1 Restriction Enzyme
Phosphatase
Ligation
STBL3 Cells
HEK293T Cells
Transfection
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