Nucleic Acids Research | Oxford Academic - Oxford Journals

Keka, et al., Supplemental Figure 1 B SmaI

ild -ty pe SM AR CA L 1 +/ SM AR CA L 1 +/ SM AR CA L 1 -/-

A XhoI

19 kb

W

Probe 19 kb

1

7.7 kb

C W

ild

-ty

pe

XhoI

D

SM AR CA

7.7 kb SmaI

L 1 -/-

Southern blotting

Marker

400bp

SM AR CA SM L1 AR C AL SM 1 AR CA L1

'3 0/-

E

'30 construct

Marker

+/-

SMARCAL1

RT-PCR

+/-

+/+

RPA-binding motif

400bp Marker

310bp RT-PCR

Popout marker

SMARCAL1 310bp

'30/-

TAGCCAAAGGAGGCCGCC- - - - - -ACATGGGCCTGGGGAAGA 15 base

kb

Wi ldtyp

TALEN induced DSB

#1

SM AR CA

B

SMARCAL1

e

A

L 1 -/ -

Keka, et al., Supplemental Figure 2

#2

#3

6 ATG

8th 3.2 TALEN nuclease

Marker

Southern blotting

LoxP

Smarcal1

*

6 kb

Actin

Exon 8th

Western blotting Probe

Marker 3.2 kb

XbaI XbaI

XbaI

C ICRF193

100

J-rays

100

Wild-type 10 10

1 1 0 0.1

0

100

200 nM

300

1 Gy

400

Wild-type LIG4 -/-/-

-/SMARCAL1 clone #1 -/SMARCAL1 clone #2 -/SMARCAL1 clone #3

2

Keka, et al., Supplemental Figure 3 B

LIGASE4 (LIG4)

Wi

ldtyp e

guide RNA AGATTCATCACCGCTTTG ATG G

LI caG4 -/nd /iat es

A

#1 #2 #3 #4 #5 #6

LIG4

* Western Blotting

Marker

D

NEO primer

PURO primer

e

typ

15 base TTGGCTCCCAGCCAGCT- - - - - - - -TGAAGGCAGGTCCTGTGA

2nd

#1 #2

Wi

Wi

TALEN-induced DSB

ld-

ld-

typ

RAD54

e

C

LoxP

#1 #2

2.3 kb Genomic PCR

R TALEN nuclease

Marker

LoxP

F

DN

DN SM A-P AR Kcs -/ CA L1 -/-

A-P

pe d-ty Wil

guide RNA GCAAATGCACCGTTGTGGTCCGG

Marker

F

DNA-PKcs

#1 #2 #3 #1 #2

DNA-PKcs GAPDH

32nd

Kc s /A-P DN

pe

LoxP

d-ty

Marker

DN SM A-P AR Kcs -/ CA L1 -/ -

RT-PCR

R

F

Wil

E

Kc s /-

2.3 kb

#1 #2 #3 #1 #2

DNA-PKcs ACTIN Western blotting

Keka, et al., Supplemental Figure 4

A J-rays

10 Wild-type DNA-PKcs-/- clone#1 DNA-PKcs -/- clone#2 DNA-PKcs -/- clone#3 1 0

J-rays

100

Cell survival (%)

Cell survival (%)

100

1 Gy

2

Wild-type 10 -/-/SMARCAL1 / DNA-PKcs clone #1 -/-/SMARCAL1 / DNA-PKcs clone #2 1

0

1 Gy

B ICRF193

Cell survival (%)

100

Wild-type SMARCAL1 -/-/DNA-PKcs -/-/SMARCAL1 / DNA-PKcs

10

1

0

100

200 nM

300

400

C J-rays

Cell survival (%)

100

Wild-type 10

SMARCAL1

SMARCAL1

1

0

1 Gy

2

-/-

'30

2

4 -/ -/ LIG

Wild-type

SM AR CA

L 1 -/ -

A

SM AR /LI CA G 4 - L 1 -/ / -/ -

Keka, et al., Supplemental Figure 5

Asynchronous

G1 -synchronized

DNA content (PI)

B Merged JH2AX

Merged

JH2AX

53BP1

Merged

53BP1

Wild-type

Wild-type SMARCAL1

Merged

-/-

SMARCAL1 4h 0h G1 -synchronized, 1 Gy

LIG4

-/-

-/-/-

4h 0h G1 -synchronized, 1 Gy

Keka., et al. Supplemental Figure 6

A

B

Thymidine kinase (TK) gene alleles

Wild-type

I-Sce1

TK

100 bases Exon4

+/-

3 kb 2 kb 1.6 kb 1 kb

5 6 7

SMARCAL1 Wild-type and SMARCAL1 Fa

-/Ra

TK Exon 4

5

-/-

3 kb 1.6 kb2 kb 1 kb

6 7 LIG4

1.9 kb LIG4

-/-

-/-/-

6 kb

-/-/-

Fa

Rb

3 kb 2 kb

TK Exon 4

-/-

5 6 7 5.8 kb

C

D

~3 kb 12%

E ~3 kb 9%

~4 kb 7%