Instruct: Identify the main category of Biorxiv papers based on the titles and abstracts
Query: Comparison of CRISPR/Cas9-mediated megabase-scale genome deletion methods in mouse embryonic stem cells The genome contains large functional units ranging in size from hundreds of kilobases to megabases, such as gene clusters, promoter-enhancer loops, and topologically associating domains. To analyze these large functional units, the technique of deleting the entire functional unit is effective. However, deletion of such large regions is less efficient than conventional genome editing, especially in cultured cells, and a method that can ensure success is anticipated. Here, we compared methods to delete the 2.5-Mb Kruppel-associated box zinc finger protein (KRAB-ZFP) gene cluster on chromosome 4 in mouse embryonic stem cells using CRISPR/Cas9. Three methods were used: first, deletion by non-homologous end joining (NHEJ); second, homology-directed repair (HDR) using a single-stranded oligodeoxynucleotide (ssODN) with 70-bp homology arms; and third, HDR employing targeting vectors with a selectable marker and 1-kb homology arms. NHEJ-mediated deletion was achieved in 9% of the transfected cells. The deletion frequency of NHEJ and HDR was found to be comparable when the ssODN was transfected. Deletion frequency was highest when targeting vectors were introduced, with deletions occurring in 31-63% of the drug-resistant clones. Biallelic deletion was observed when targeting vectors were used. This study will serve as a benchmark for the introduction of large deletions into the genome.
15
The categories of this paper are: developmental
2
The categories of this paper are: biology
24
The categories of this paper are: cancer
6
The categories of this paper are: plant
1
The categories of this paper are: bioinformatics
28
The categories of this paper are: communication
7
The categories of this paper are: ecology
21
The categories of this paper are: toxicology
11
The categories of this paper are: cognition
14
The categories of this paper are: biophysics
16
The categories of this paper are: microbiology
30
The categories of this paper are: scientific
13
The categories of this paper are: animal
12
The categories of this paper are: behavior
3
The categories of this paper are: evolutionary
17
The categories of this paper are: molecular