Supplemental Material:

Supplemental Materials and Methods
Immunofluorescence
Immunostaining of ovaries, S2 cells and OSCs was performed as previously described(Saito et al. 2006; Saito et al. 2009). The primary antibodies used were: culture
supernatants of anti-Armi hybridoma cells (1:1 dilution), culture supernatants ofanti-Piwi hybridoma cells (P4D2; 1:1 dilution) (Saito et al. 2006), a mouse monoclonal
antibody for the myc-tag (1:1,000 dilution; Sigma), a guinea pig antibody against TJ(1:2,000 dilution) (Li et al. 2003) and a rabbit polyclonal antibody against Yb (1:250
dilution) (Szakmary et al. 2009). The secondary antibodies used were: Alexa
488-conjugated anti-mouse IgG (Molecular Probes), Alexa 546-conjugated anti-mouse
IgG (Invitrogen), Alexa 488-conjugated anti-rabbit IgG (Invitrogen), Alexa
546-conjugated anti-guinea pig IgG (Invitrogen) and Alexa 546-conjugated anti-rabbit
IgG (Invitrogen). DNA was stained with 4′,6-diamidino-2-phenylindole (DAPI). Before
fixation, the cells were incubated with MitoTracker Orange CMXRos (Invitrogen)targeted at the active mitochondria in living cells. All images were collected using a
confocal microscope (Zeiss LSM5 EXCITER; Zeiss).

Immunoprecipitation
Immunoprecipitation from S2 lysates was performed essentially as previously described(Miyoshi et al. 2005). Immunoprecipitation from OSCs was also performed essentially1

as previously described (Saito et al. 2009). Briefly, 1 × 108 OSCs were homogenized inhomogenization buffer [20 mM HEPES-KOH (pH7.3), 2 mM MgCl2, 1 mM
dithiothreitol (DTT), 150 mM NaCl, 2 µg/ml pepstatin, 2 µg/ml leupeptin, 0.5%
aprotinin and 0.1% NP-40] to prepare OSC lysates. Anti-Piwi (P3G11) (Saito et al.
2006), anti-Armi and anti-Yb (Szakmary et al. 2009) antibodies were immobilized on
Dynabeads protein G (Invitrogen). Reaction mixtures were rocked at 4°C for 2 h andthe beads were washed four times with homogenization buffer. Total RNAs wereisolated from the immunoprecipitates with phenol-chloroform and were precipitatedwith ethanol. RNAs were dephosphorylated with calf intestinal phosphatase (TaKaRa)
and labeled with 32P-gATP using T4 PNK (TaKaRa) for visualization.

Analysis of piRNA-intermediate chemical structures
Total RNAs were isolated from the anti-Armi immunoprecipitates. Phosphorylation ofRNAs was performed in the presence of 1 mM ATP with T4 PNK (NEB). Alkaline
dephosphorylation was performed with calf intestinal phosphatase (NEB). Acidic
dephosphorylation was carried out with T4 PNK (Takara) in 50 mM Tris-HCl (pH 6.5),
10 mM MgCl2 and 5mM DTT. Enzymes were inactivated by phenol/chloroform
extraction, followed by RNA precipitation. Precipitates were dissolved in H2O.
Polyadenylation was performed with poly (A) polymerase (NEB) and 1 mM ATP in 1 ×poly (A) polymerase reaction buffer. RNAs were resolved on a 12% denaturing
polyacrylamide gel and blotted on Hybond N+ (GE Healthcare) or Hybond N (GEHealthcare) membrane. To cross-link RNAs, UV irradiation or EDC reagent was used.
2

Drosophila strains
Yellow white (y w) and Oregon R were employed as WT strains. The armi alleles usedwere armi72.1 and armi1 (Cook et al. 2004). All stocks were maintained at 25°C.

myc-Piwi expression and RNAi in OSCs
To yield RNAi-resistant piwi cDNA (Piwi-r), 5′ and 3′ fragments of piwi cDNA wereindividually amplified by KOD plus DNA polymerase (Toyobo) using the following
primers: 5′ fragment, Piwi-F and Piwi-mt-R; 3′ fragment, Piwi-mt-F and Piwi-R(primers are listed in Supplemental Table S2). To yield Piwi PAZ mutant cDNA
(myc-Piwi-PAZmt), 5′ and 3′ fragments of piwi cDNA were individually amplified by
KOD plus DNA polymerase (Toyobo) using the following primers: 5′ fragment, Piwi-F
and Piwi-PAZmt-R; 3′ fragment, Piwi-PAZmt-F and Piwi-R (primers are shown in
Table S2). The resulting DNA fragments were gel-purified, mixed and used as
templates for PCR amplification. DNA fragments were amplified by PCR with Piwi-F
and Piwi-R. The resulting PCR products were cloned into the pAcM vector (Saito et al.
2009) and the sequence was validated by DNA sequence analysis. A short DNA
fragment encoding a myc-tag was inserted between the Xho I and Bam HI sites ofpAcM to produce pAcM-Myc. Full-length zuc cDNA was inserted between the Nhe I
and Xho I sites of pAcM-Myc to yield pAc-Zuc-myc. For RNAi and expressing
myc-tagged proteins in OSCs, trypsinized OSCs (3 × 106 cells) were suspended in 100µl of Solution V of the Cell Line Nucleofector Kit V (Amaxa Biosystems) together with3

either 200 pmol of siRNA duplex or 5 µg of plasmid vector. Transfection was
conducted in electroporation cuvettes using a Nucleofector device (Amaxa Biosystems).
The transfected cells were transferred to fresh OSC medium and incubated at 26°C for 2
d for further experiments. The siRNAs utilized for RNAi are summarized in Table S2.
The control siRNA was enhanced green fluorescent protein (EGFP) siRNA (Saito et al.
2009). Quantitative RT-PCR was performed as previously described (Kawamura et al.
2008). Total RNA (0.5 µg) was used to reverse transcribe target sequences using
oligo(dT) primer. The resulting cDNA was analyzed by quantitative RT-PCR using a
LightCycler real-time PCR system (Roche Diagnostics) using SYBR Premix Ex Taq(Takara). Relative steady-state mRNA levels were determined from the threshold cycle
for amplification. Table S2 lists the PCR primer sequences. Ribosomal protein 49(Rp49) was used as an internal control.

Cloning of small RNAs and processing sequence tags
Armi

was

immunopurified

from

OSCs

using

a

specific

antibody.

Afterimmunoprecipitation, total RNAs were isolated from the anti-Armi immunoprecipitates.
Acidic dephosphorylation was carried out with T4 PNK (Takara) in 50 mM Tris-HCl(pH 6.5), 10 mM MgCl2 and 5mM DTT. Enzymes were inactivated byphenol/chloroform extraction, followed by RNA precipitation. Precipitates weredissolved in H2O. RNAs were resolved on a 12% denaturing polyacrylamide gel with
5′-labelled Decade Marker (Ambion) as a size marker. RNAs of 50-75 nt were excisedfrom the gel. Cloning of RNAs was carried out as described previously (Saito et al.
4

2009). The sequencing was performed on a GS FLX system (Roche).

Genome mapping, annotation and frequency mapping
We followed a previously described procedure to map and annotate sequences (Saito etal. 2009). We mapped 116,680 piR-IL sequences to the Drosophila melanogaster (dm3)genome. The annotation results are shown as a pie-chart in Supplemental Fig. S4A.
Maps for piL-ILs were generated (Supplemental Fig. S5 and S6A), representing thenumber of piR-ILs that are 100% identical to the genomic sequence. RNA sequences ofpiR-ILs have been deposited at the GEO database under accession number GSE23931.

Northern blotting
Northern blotting was performed as previously described (Saito et al. 2006). DNA
oligonucleotides used are summarized in Table S2.

5

Supplemental Figures

120 k-

n.i.

anti-Armi
anti-Armi

220 k160 k-+- + S2 lysate

B
S2

ovary

A

Figure S1A-D
Saito et al.

- Armi
220 k-

100 k-

160 k-

80 k-- Armi120 k-

70 k-

100 k-

60 k80 k-

50 k-

70 k-

40 k(Da)

Armi

60 k-

- Tubulin

50 k-

1188

40 k-

Helicase domain
(Da)

*
D
OSC

C

Armi

Armi/DAPI

Armi/DAPI

Armi

Armi/DAPI

germariumArmi

egg chamberArmicontrolArmi / DAPI

Armi
RNAi
Armi

Armi / DAPI

Figure S1E-G
Saito et al.

GArmi / DAPI

armi / armi

/ armi1

1

6.6 5.0 -] long form
] short form

(kb)

1

1

armi / +yw

72.1

3.6 2.6 -

1

F

armi

armi / armi

yw

1

armi / +1

E- Armi- Tubulin

Supplemental Figure S1. (A) Western blot analyses on ovary and S2 cell lysates using anti-Armi
antibodies. Anti-tubulin was utilized as an internal control. The antigen (the N-terminal end of
Armi; aa 1-200; indicated as *) used for immunizing mice is shown at the bottom. (B)Immunoprecipitation was performed from S2 cells using anti-Armi antibodies. Mass spectrometricanalysis determined that the ~150 kDa protein is Armi. (C) Immunofluorescence of Armi in OSCsand ovaries. DAPI staining (blue) shows the locations of the nuclei. (D) Immunostaining of OSCs
before and after the treatment with RNAi for Armi indicates that the dotted signals in the cytoplasm reflect the subcellular localization of Armi in the cells. (E) Immunostaining pattern of Armiin the armi transheterozygous mutant ovary. (F) Western blot analysis shows that Armi isexpressed even in the armi homozygous mutant ovaries. (G) Northern blotting shows that the armitranscripts (short form) are expressed in the armi homozygous mutant ovaries.

Saito et al.
Figure S2

A
DCP1

DCP1/Yb

Yb

Yb / DAPI

DCP1/Yb/DAPI

B

Yb

controlArmi

Armi / DAPI
control

Yb / DAPI
Armi RNAi

Armi

Armi / DAPIYb RNAi

Supplemental Figure S2.
(A) Coimmunostaining of OSCs with anti-Yb and anti-Dcp1
antibodies shows that Yb bodies do not colocalize with P bodies. (B) Depletion of Armi
did not affect the cellular localization of Yb whereas depletion of Yb caused Armi to belocalized to the cytoplasm.

Saito et al.
Figure S3A-D

C
Yb

Armi

Piwi

control

RNAi in OSC

Yb

Armi

RNAi
Piwi

controlA- Piwi] Armi- Yb

50 nt 40 nt 30 nt - tj piRNA- Tubulin
20 nt -- miR-310

BPiwi- sl-1 esiRNA

controlArmi RNAi
Piwi

D

DAPI

TJ

myc-Piwi-PAZmt

myc-Piwi-PAZmt/DAPI

myc-Piwi

myc-Piwi/DAPI

DAPI

TJ

Saito et al.
Figure S3E-F

E
anti-Yb

anti-Piwi

kDa

n.i.

input

IP from OSC

175 -- AGO2
AGO1
80 -

F
IP
n.i.
1

anti-Piwi
1

1/20 1/40 1/60

antiArmi
1

dilution- Piwi

Supplemental Figure S3.
(A) Western blotting was performed using anti-Piwi, anti-Armi and antiYb antibodies on OSC lysates in which Piwi, Armi and Yb were depleted by RNAi. Anti-Tubulin
was used as an internal control. siRNA for EGFP was employed as a negative control (control). (B)Depletion of Armi caused Piwi (green) to be mislocalized to the cytoplasm. In contrast, the nuclear
localization of TJ protein (red) was unaffected by Armi depletion. (C) Yb depletion causes a severe
reduction of the piRNA level in OSCs (~25% of the control signal). (D) The Piwi PAZ mutant(green) (myc-Piwi-PAZmt; containing two mutations, Y327A and Y328A) accumulates in the cytoplasm while myc-Piwi wt (green) localizes to the nucleus, supporting the idea that piRNA loading isrequired for the nuclear localization of Piwi. DAPI staining (blue) shows the locations of nuclei in thecells. (E) The complexes immunoprecipitated with anti-Yb and anti-Piwi antibodies contain neither
AGO1 nor AGO2. (F) Western blot analysis shows that the amounts of Piwi are approximately equalin the complexes immunoisolated with anti-Piwi and anti-Armi antibodies only when the anti-Piwi
complex was diluted 1:40 before loading.

Saito et al.
Figure S4A-B

A
no annotation
6.2%
transposons
16.6%
protein-coding genes
protein-coding gene
35.9%

snoRNA
1.4%
snRNA
3.5%
rRNA
35.7%

rRNA

ncRNA

tRNA

repeatsnRNAtransposons

snoRNAno annotation

actin 5c ORF
probe 1

actin 5c ORF
probe 2

tj ORFprobe 1

control oligo
antisense oligoIPtotal RNAn.i.
anti-Armi

control oligo
antisense oligoIPtotal RNAn.i.
anti-Armi

control oligo
antisense oligoIPtotal RNAn.i.
anti-Armi

total RNAn.i.
anti-Armi

IP

control oligo
antisense oligoB

tj ORFprobe 2

Supplemental Figure S4.
piR-IL analyses. (A) The small RNA content (50-75 nt) in the ArmiPiwi-Yb complex perfectly matches the Drosophila genome sequence. (B) Northern blotting
analyses. RNAs isolated from OSCs (total RNAs) and the anti-Armi immunoprecipitates (antiArmi IP) were probed with DNA oligos corresponding to actin 5c and tj ORF. n.i. was used as anegative control for immunoprecipitation. The sequences of the probes used are;5'-GCCTCGGTCAGCAGCACGGGGTGCT (actin 5c-probe1)5'-CGCAGGATGGCATGGGGAAGGGCAT (actin 5c-probe2)5'-TCAACGTGGTCAGCATGTCGTCATT (tj-probe1)5'-ACCCGAAGCACTACCGTACGAGTTC (tj-probe2)
RNAs isolated from the Armi-IP complex appeared negative although control oligos were- positive, suggesting that clones corresponding to actin 5c and tj ORF are background.

A

Saito et al.
Figure S5A
21,550,000

21,600,000

21,650,000

piRNAs (black shows piRNAs overlapping with piR-ILs)
Unique mappers

OSC Piwi-associated piRNAs

Chr. X
21,500,000

OSC piR-ILs

Unique mappers

piR-ILs (black shows piR-ILs overlapping with piRNAs)

FlyBase Protein-Coding Genes

DIP1

Repeating Elements by RepeatMasker

SINE
LINE
LTR
DNA
SimpleLow Complexity
SatelliteRNA
Other
UnknownpiRNAs21,504,000

21,506,000

piRNAs (black shows piRNAs overlapping with piR-ILs)

piR-ILs (black shows piR-ILs overlapping with piRNAs)

Unique mappers

piR-ILs

Unique mappers

21,502,000

DIP1
SINE
LINE
LTR
DNA
SimpleLow Complexity
SatelliteRNA
Other
Unknown

FlyBase Protein-Coding Genes
Repeating Elements by RepeatMasker

21,508,000

21,510,000

Saito et al.
Figure S5B

B
chrX:21,508,080-21,508,550

OSC Piwi-associated piRNAs

Unique mappers (plus)

OSC piR-ILs

Unique mappers (plus)

21,508,100

21,508,200

21,508,300

21,508,400

21,508,500

piRNAs (black shows piRNAs overlapping with piR-ILs)

piR-ILs (black shows piR-ILs overlapping with piRNAs)

Repeating Elements by RepeatMasker
SINE
LINE
LTR
DNA
SimpleLow Complexity
SatelliteRNA
Other
Unknown

Supplemental Figure S5. (A) Upper: The distribution patterns of piRNAs (Saito et al. 2009)and piR-ILs on the piRNA locus flam sequence. Only uniquely mapped ones are shown.
Black bars show piRNAs/piR-ILs overlapping with piR-ILs/piRNAs. Green bars show
piRNAs/piR-ILs non-overlapping with piR-ILs/piRNAs. Lower: The magnification of theregion specified in (A). (B) The distribution of uniquely mapped piR-ILs and mature piRNAs
on the chromosome X, position 21,508,080-21,508,550. Black bars show piRNAs/piR-ILs
overlapping with piR-ILs/piRNAs. Green bars show piRNAs/piR-ILs non-overlapping with
piR-ILs/piRNAs.

Saito et al.
Figure S6A
chr2L

piRNAs

19,465,000

19,466,000

19,467,000

19,468,000

piRNAs (black shows piRNAs overlapping with piR-ILs)

piR-ILs

piR-ILs (black shows piR-ILs overlapping with piRNAs)

5'UTR
traffic jam

3'UTR
ORF

B 100%
C
80%

G

60%

40%

U

20%

A
0%

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20

Supplemental Figure S6.
(A) The distribution patterns of piRNAs and piR-ILs on the tj genesequence. Some piR-ILs were mapped to the tj ORF. However, the signals for the clones were notdetected in the Armi-Piwi-Yb complex (Supplemental Fig. S4B) by Northern blotting, suggestingthat they may be considered as 'background' of the small RNA library. piR-ILs were detected in
the complex by Northern blotting (Fig. 3C and 3D). (B) The 5' end of piR-ILs mapped totransposons was not U-rich as oppose to that of mature transposon-piRNAs. Only a small numberof transposn-piR-ILs (~4%) has G at their 5' end, although the reason for this appearance remains
unknown.

Saito et al.
Figure S7A

Bcontrol

Zuc RNAi

Yb / Armi / DAPIYb / Armi / DAPI

1.0
0.8
0.6
0.4
0.2

D
control

0

Zuc
RNAi

n.i.

anti-myc

1/8 1/8

1

1

n.i.

controlanti-myc

C

Mito

Zuc

Mito / Zuc

Mito / Zuc / DAPIZucRNAi

relative Zuc mRNA

1.2

E
dilutionMito

Yb

40 nt -

30 nt -

Yb / Zuc / DAPI

Mito / Yb Mito / Yb / DAPI

20 nt -

< 0.01< 0.011.0
0.18

Supplemental Figure S7.
(A) The efficiency of Zuc depletion in OSC was determined by
qRT-PCR. (B) Cellular localization of Armi in OSC was not affected by Zuc depletion. (C)32 P end-labeling showed that when Zuc was depleted, myc-Piwi was loaded with far fewer
piRNAs compared with loading under normal conditions. (D) Zuc (green) localizes to mitochondria (red) when expressed in OSCs. (E) Left panel; Zuc signals (green) can be detected
in close proximity to Yb bodies (red). Right four panels; Yb signals (green) are detected inclose proximity to mitochondria (red).

Saito et al.
Figure S8

Banti-PiwiA

anti-Armi-- + - - + T4 PNK

anti-Armi
anti-AGO1-- + - - + T4 PNK
CIP

CIP

100 nt -

+
-

+ PAP
+ T4 PNK (Acidic)

100 nt 75 nt -50 nt 40 nt 30 nt -EDC

30 nt -

EDC
50 nt -

20 nt -

20 nt -

40 nt miR-310

30 nt 20 nt -100 nt 75 nt 50 nt 40 nt 30 nt -UV

tj-piRNA

20 nt tj-piRNA

Supplemental Figure S8.
(A) piR-ILs are undetectable by northern blotting in which the RNAs
were cross-linked by EDC. They become detectable after phosphorylation, suggesting that piR-ILs
are free from phosphate groups at both ends. It should be noted that mature tj-piRNA and miR-310
are detectable by the EDC method even without phosphorylation. (B) piRNA-ILs are barely
polyadenylated unless they were pre-treated with PNK under acidic conditions, suggesting thatthey contain a cyclic phosphate at their 3' end. From this result, together with those in (A), it issuggested that piR-ILs are free from phosphate groups at 5' end but contain a cyclic phosphate attheir 3' end.

A

Saito et al.
Figure S9

C

160
200

140

250

180
160

40

200

140
120

20

150

100

6

80

100

60

4

40
20

2

1

1
0

siEGFP

B

70

50

10

0

siPiwi

D

E

80

300
60

70

50

60

40

250
200

50
40

150

30
30

20

100
20

10

10

1

0

0

50
0

control Piwi RNAi

control

Piwi RNAi

Supplemental Figure S9. (A) Piwi depletion in OSCs causes derepression of various transposons.
(B) Piwi and Armi depletion by RNAi in OSCs caused derepression of mdg1 transposon. RNAi was
performed for 48 hours. (C) Yb and Zuc depletion by RNAi in OSC caused derepression of mdg1.
RNAi was performed for 96 hours. (D) Quantitative RT-PCR analysis for the mdg1 transposon. In
OSCs where endogenous Piwi was depleted, myc-Piwi-r (RNAi-insensitive Piwi), but not myc-Piwi
or myc-Piwi-ΔN (Piwi lacking its nuclear localization signal), was able to rescue transposon derepression. (E) RNAi-insensitive Piwi-r and Piwi-DDAA-r rescued transposon silencing in Piwidepleted OSC.

Table S1.
Saito et al.
Supplemental Table 1. Genes depleted in OSCs.
Gene

Function

Referencearmi

piRNA pathway(Vagin et al., 2006; Malone et al., 2009)

mael

piRNA pathway(Lim and Kai, 2007)

spn-E

piRNA pathway(Vagin et al., 2006; Malone et al., 2009)

zuc

piRNA pathway(Malone et al., 2009; Pane et al., 2007;
Saito et al., 2009)

dicer1

miRNA pathway(Vagin et al., 2006)

dicer2siRNA pathway(Vagin et al., 2006)

Table S2.
Saito et al.
Supplemental Table 2
Experiment

Primer Name

Primer sequence

Piwi-F
Piwi-R
Piwi-mt-F

ATAAGCTTGCTGATGATCAGGGACGTGATAGATCTTTATAGATAATAAAACTTCTTTTCGAGCGAGGAGGCACCGCGCAGGGAGGGAGGCCCGCCAGAGCGAAAGCCG
CGGGCCTCCCTCCCTGCGCGGTGCCTCCTCGAGAGCTCCTCTCTCTCGATATCAGTTTCGTGGAAGCCGCTCTCACTAAATATAATATACGCATTCGCG
GCGTATATTATATTTAGTGAGAGCGGCTTCCACGAAACTGATATCTCT
ACC
TCGAGGGAGAGCAGAAACTGATCTCTGAAGAAGACCTGAACTAAGGATCCTTAGTTCAGGTCTTCTTCAGAGATCAGTTTCTGCTCTCCC
TTGCTAGCCACCATGTTGATTACCCAAATAATTATGAAACAA
ATCTCGAGCTTGAGCTGGATTTGGCTCC

Piwi-mt-R

Vectorconstruction

Piwi-PAZmt-F
Piwi-PAZmt-R
Myc-tag-5
Myc-tag-3
Zuc-F
Zuc-R

Experiment

qRT-PCR

Gene

Primer sequence (Forward, Reverse)

RP49
297
1731
I- element
HeT-A
Mdg1
Roo
ZAM
Zucchini

CCGCTTCAAGGGACAGTATCTG, ATCTCGCCGCAGTAAACGC
CTGGCAAAGGGATTTCATCA, TGCATTCCTAAGGCCAAATG
TATGGGCTGAGGCGATAAAC, CAAGTGGCTCACTGCTGGTAGACCAAATAAAAATAATACGACTTC, AACTAATTGCTGGCTTGTTATGCGCGCGGAACCCATCTTCAGA, CGCCGCAGTCGTTTGGTGAGTAACAGAAACGCCAGCAACAGC, CGTTCCCATGTCCGTTGTGATCGTCTGCAATGTACTGGCTCT, CGGCACTCCACTAACTTCTCC
ACTTGACCTGGATACACTCACAAC, GAGTATTACGGCGACTAGGGATACTGATTTGGAAGCTGGTGCAG, GCGACACTTGTGGTTTCTGG

Experiment

Gene

Probe sequence

Northern Blot

U6 snRNA
miR-310sl-1 esiRNATj-piRNA
Idefix-piRNA

GGGCCATGCTAATCTTCTCTGTA
AAAGGCCGGGAAGTGTGCAATAGGAGCGAACTTGTTGGAGTCAAGGTAATGGGAATGCACTTCTCTTGAAAAACTACTGGCAATCGTTTGGGAA

Experiment

RNAi

siRNA namesiRNA sense

siRNA antisense

EGFP-siPiwi-si
Zucchini –si
Armitage-si
Yb-si
Maelstrom-si
Spindle-E-si
Dicer1-si
Dicer2-si

GGCAAGCUGACCCUGAAGUTTGCUCCCAGGCGUGAAGGUGTTGCAUUGCCGUCAGCACUGUTTCAGCUUAAGGCGAAUCCAATT
GGGGUACAUCAACCAAGCUTTCGCCAAGAUGUCCCAUGAUTTCUCGCGGCAUGUAAAGAUUTTGAUUGUCAUGACAGCGCCGTT
CAGGGAGUGGAUGACUGGATTACUUCAGGGUCAGCUUGCCTT
CACCUUCACGCCUGGGAGCTTACAGUGCUGACGGCAAUGCTTUUGGAUUCGCCUUAAGCUGTTAGCUUGGUUGAUGUACCCCTT
AUCAUGGGACAUCUUGGCGTTAAUCUUUACAUGCCGCGAGTTCGGCGCUGUCAUGACAAUCTTUCCAGUCAUCCACUCCCUGTT