19 Life Technologies
™
| Genome editing
Figure 4
A. 293FT cells, HPRT locus
Cleavage
efficiency
(%)
60
50
40
30
20
10
0
1 2 3 4
500 bp
400 bp
300 bp
200 bp
Cleavage efficiency: 32% 36% 51% 0%
11% 25% 51% 0% 10% 9% 32% 0% 4% 17% 49% 0%
CRISPR-Cas9 format Lane
All-in-one plasmid 1
Cas9 mRNA + Strings™ DNA
2
Cas9 mRNA + IVT gRNA 3
Control (Cas9 mRNA only) 4
C. U2OS cells, RELA locus
Cleavage
efficiency
(%)
60
50
40
30
20
10
0
1 2 3 4
300 bp
200 bp
CRISPR-Cas9 format Lane
All-in-one plasmid 1
Cas9 mRNA + Strings™ DNA
2
Cas9 mRNA + IVT gRNA 3
Control (Cas9 mRNA only) 4
Cleavage
efficiency
(%)
500 bp
400 bp
300 bp
200 bp
CRISPR-Cas9 format Lane
HPRT RELA
All-in-one plasmid 1 5
Cas9 mRNA + Strings™ DNA
2 6
Cas9 mRNA + IVT gRNA 3 7
Control (Cas9 mRNA only) 4 8
B. HCT116 cells, HPRT and RELA loci
60
HPRT RELA
50
40
30
20
10
0
1 2 3 4 5 6 7 8
Assay for:
System
Ordering information
Product Quantity Cat. No.
GeneArt
®
CRISPR Nuclease mRNA 15 μg A25640
GeneArt
®
Strings
™
U6 DNA >200 ng
Contact geneartsupport
@
lifetech.com
GeneArt
®
Strings
™
T7 DNA >200 ng
Contact geneartsupport
@
lifetech.com
Custom in vitro transcribed gRNA 250 nmol
Contact geneartsupport
@
lifetech.com
Figure 5. The GeneArt
®
CRISPR Nuclease mRNA System demonstrates efficient genome editing in a broad
range of cell types. (A) 293FT, (B) HCT116, and (C) U2OS human cell lines were transfected in 24-well plates
using the indicated CRISPR-Cas9 format, targeting the HPRT or RELA locus. The CRISPR-Cas9 formats and
corresponding sample lane numbers are listed in the tables. The GeneArt
®
CRISPR Nuclease Vector with
OFP reporter was transfected using Lipofectamine
®
3000 Transfection Reagent, the Strings
™
DNA format was
transfected using Lipofectamine
®
2000 Transfection Reagent, and the IVT gRNA format was in vitro transcribed
using delivered MEGAshortscript
™
T7 Transcription Kit (Cat. No. AM1354) and MEGAclear
™
Transcription Clean-
Up Kit (Cat. No. AM1908), and then transfected using Lipofectamine
®
MessengerMAX
™
Reagent. At 72 hours
post-transfection, cells were harvested and genome editing efficiency was quantified using the GeneArt
®
Genomic Cleavage Detection Kit (Cat. No. A24372). The cleavage products are indicated with arrowheads.
