B DAPIDNAmerge S P A CENP-A HDAC1 Aurora B C Centrifugation on 15% sucrose 200 g, 10 min Centrifugation on 30% sucrose 2500 g, 20 min Centrifugation on.

B

DAPI DNA merge

S

P

A

CENP-A

HDAC1

Aurora B

C

Centrifugation on 15% sucrose 200 g, 10 min

Centrifugation on 30% sucrose 2500 g, 20 min

Centrifugation on 30% sucrose 27000 g, 30 min

Elimination of unbroken nuclei

Elimination of nucleoli

Supernatant (S) = low density fraction

Pellet (P) = chromocenters

Nuclei + 2 mM TEA, 0.2 mM MgCl2, 0.2% TritonSonication

Isolation of the chromocenters

200400

100

bp

S P

- + - +

minsat

E

Sir2

Survivin

S P

Ferri et al. Supplemental Figure 1

S P

minsat

200100

400600bp

15

40

15

60

120

kDa

D

minsat

rDNA

H3A

c

H3K

9Me3

CE

NP

-A

bea

ds

INP

UT

IP SUP

H3A

c

H3K

9Me3

CE

NP

-A

bea

ds

Tot

al D

NA

Mar

ker

100b

p

majsat

Ferri et al. Supplemental Figure 2

300500

100

300500

100

bp

300100

200300

100

400600

15002000

bp

- +- +AS NOC

min

sat

-actin

AS

0 400

0

100

200

200 600

G0/G1: 33%S: 57%

G2/M: 10%NOC

0 400

0

100

200

200 600

G0/G1: 2%S: 12%

G2/M: 86%

Ferri et al. Supplemental Figure 3

PI

coun

ts

PI

coun

ts 200

300400500

bp

1: RNA 2’OMe (1-27) 5’biot-GGAAACAUGAUAAAAACCACAGUGUAG-3’

2: DNA (1-27) 5’biot-GGAAACATGATAAAAACCACAGTGTAG-3’

3: DNA/lna (4-27) 5’-aACaTGaTAaAAaCCaCAgTGtAG-3’biot

4: DNA/lna (89-115) 5’-tGTgTAtATcAAtGAgTTaCAaTG-3’biot

5: DNA/lna (4-27) 5’biot-aACaTGaTAaAAaCCaCAgTGtAG-3’

murine minor satellites 120 nt repeat unit consensus sequence :

5’GGAAACATGATAAAAACCACAGTGTAGAACATATTAGATGAGTGAGTTACACTGAAA

AACACATTCGTTGGAAACGGGATTTGTAGAACAGTGTATATCAATGAGTTACAATGAG

AAACAT-3’

Ferri et al. Supplemental Figure 4

inpu

t

1 2 3 4 5

RNA minsat

MEL nuclear extractsStreptavidin

beads

Biotinylated oligonucleotide

In vitro transcribed RNA