plasmids - static-content.springer.com10.1038...pcag znf91 [1-11]. pcag znf91 was digested with xmai...

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Plasmids ZNF Army pCAG EV. pCAG EN (CEPKO Lab) was ordered (Addgene plasmid 11160) that contained multiple cloning sites for subcloning. pCAG GFP. pCAG GFP (Cepko Lab) was ordered (Addgene plasmid 11150) and used as a transfection efficiency control as well as a co-transfection plasmid fort FACS applications in the Tc11 Rescue experiments. pCAG ZNF91. ZNF91 cDNA sequence was synthesized as a codon-optimized pUC57 insert flanked by EcoRV sites from Genscript. This insert was EcoRV digested and inserted into the pCAG EN vector at EcoRV. pCAG ZNF91 HA. pCAG ZNF91 was used as a starting source to add a C-terminal HA tag via PCR for Co-IP and western blot analysis. pCAG ZNF33a. IMAGE clone 8991835 was ordered from ATCC and PCR amplified and ligated into pENTR/D/TOPO and moved to pCAG-DEST via LR clonase reaction. pCAG ZNF90. IMAGE clone 9021208 was ordered from ATCC and PCR amplified with a C-terminal HA tag into pENTR/D/TOPO and moved to pCAG-DEST via LR clonase reaction. pCAG ZNF93. cDNA sequence was synthesized as a codon-optimized pUC57 insert flanked by EcoRV sites from Genscript. This insert was EcoRV digested and inserted into the pCAG EN vector at EcoRV.

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Page 1: Plasmids - static-content.springer.com10.1038...pCAG ZNF91 [1-11]. pCAG ZNF91 was digested with XmaI and NotI and religated resulting in the removal of the C-terminal 12-36 domains

Plasmids

ZNF Army

pCAG EV. pCAG EN (CEPKO Lab) was ordered (Addgene plasmid 11160) that contained

multiple cloning sites for subcloning.

pCAG GFP. pCAG GFP (Cepko Lab) was ordered (Addgene plasmid 11150) and used as a

transfection efficiency control as well as a co-transfection plasmid fort FACS applications in the

Tc11 Rescue experiments.

pCAG ZNF91. ZNF91 cDNA sequence was synthesized as a codon-optimized pUC57 insert

flanked by EcoRV sites from Genscript. This insert was EcoRV digested and inserted into the

pCAG EN vector at EcoRV.

pCAG ZNF91 HA. pCAG ZNF91 was used as a starting source to add a C-terminal HA tag via

PCR for Co-IP and western blot analysis.

pCAG ZNF33a. IMAGE clone 8991835 was ordered from ATCC and PCR amplified

and ligated into pENTR/D/TOPO and moved to pCAG-DEST via LR clonase

reaction.

pCAG ZNF90. IMAGE clone 9021208 was ordered from ATCC and PCR amplified

with a C-terminal HA tag into pENTR/D/TOPO and moved to pCAG-DEST via LR

clonase reaction.

pCAG ZNF93. cDNA sequence was synthesized as a codon-optimized pUC57 insert flanked by

EcoRV sites from Genscript. This insert was EcoRV digested and inserted into the pCAG EN

vector at EcoRV.

Page 2: Plasmids - static-content.springer.com10.1038...pCAG ZNF91 [1-11]. pCAG ZNF91 was digested with XmaI and NotI and religated resulting in the removal of the C-terminal 12-36 domains

pCAG ZNF93serF HA. ZNF93 Human with the 4 contact residues substituted to Ser at fingers 8-

13 as described(Moore, Choo, & Klug, 2001) thanks to personal communication with Prof. Dave

Segal (UC Davis Genome Center). cDNA sequence was synthesized as a codon-optimized

pUC57 insert flanked by EcoRI/NotI sites from Genscript. This insert was EcoRI/NotI digested

and inserted into the pCAG EN vector at EcoRI/NotI.

pCAG ZNF254. IMAGE clone 5296940 was ordered from ATCC and PCR amplified.

This was ligated into pENTR/D/TOPO and moved to pCAG-DEST with LR reaction.

znf254_ENTR_DG_F-CAC CTG TTA CCA GCA GGT ATT GGA GAT CC

znf254_ENTR_DG_R-GCG CCT CTC TCA GGT TTG TAG TTT C

pCAG ZNF443. IMAGE 5583971  was   ordered   from  ATCC   and   using   restriction   digests  was  moved  form  pSPORT6  to  pCAG.    

pCAG ZNF460. hESC cDNA was used to PCR amplify ZNF460 and this was ligated to

pENTR/D/TOPO. This was moved to pCAG-DEST with LR reaction.

pCAG ZNF486. IMAGE clone 40125819 was ordered from ATCC and PCR amplified

with HA C-terminal primer from pCR4TOPO. This was ligated into pENTR/D/TOPO

and moved to pCAG-DEST with LR reaction.

pCAG ZNF519. IMAGE clone 4806717 was ordered form ATCC and moved from

pDNR-LIB using restriction enzymes to pCAG.

Page 3: Plasmids - static-content.springer.com10.1038...pCAG ZNF91 [1-11]. pCAG ZNF91 was digested with XmaI and NotI and religated resulting in the removal of the C-terminal 12-36 domains

pCAG ZNF544. IMAGE clone 6199648 in pSPORT6 was ordered from ATCC and using

restriction enzymes was moved to pCAG.

pCAG ZNF587. IMAGE clone 8991953   was   ordered   through   Thermo.   This was PCR

cloned from the IMAGE plasmid. The amplicon was ligated to pENTR/D/TOPO

and subsequently moved to pCAG-DEST with LR clonase.

F-  caccCCGTGACGGCGACCACTG

R-  GCTACTCAGGAGGCTGAGGTAGGAG

pCAG ZNF589. IMAGE clone 100066351   was   ordered   through   Thermo.   This was PCR

cloned from the IMAGE plasmid. The amplicon was ligated to pENTR/D/TOPO

and subsequently moved to pCAG-DEST with LR clonase.

pCAG ZNF714. IMAGE clone 4797729 was ordered through ATCC. This was PCR

cloned from pBluescript clone using primers below. This amplicon was ligated to

pENTR/D/TOPO and subsequently moved to pCAG-DEST with LR clonase. This

clone represents unitprot isoform 3 (Q96N38-­‐3).

ZNF714-F caccCTAGAAATGGGCGACCTGAG,    

ZNF714-­‐R  AGACACCACACCTGGCCTAC

pCAG ZNF721. cDNA sequence was synthesized as a codon-optimized pUC57 insert flanked

from Genscript. This insert was moved to the pCAG EN vector at EcoRV.

Domain deletion analysis of ZNF91

Page 4: Plasmids - static-content.springer.com10.1038...pCAG ZNF91 [1-11]. pCAG ZNF91 was digested with XmaI and NotI and religated resulting in the removal of the C-terminal 12-36 domains

pCAG ZNF91 [1-11]. pCAG ZNF91 was digested with XmaI and NotI and religated resulting in

the removal of the C-terminal 12-36 domains.

pCAG ZNF91 [1-11] HA. pCAG ZNF91 [1-11] was used as a starting source to add a C-terminal

HA tag via PCR and a N-terminal CACC site for ligation into pENTR/D/TOPO (Invitrogen).

This was used in an LR reaction to transfer the insert to pCAG-DEST. This construct was used

for western blot validation of ZNF91 [1-11].

pCAG ZNF91 [1-24]. pCAG ZNF91 was digested with AleI and NotI and religated resulting in

the removal of the C-terminal 25-36 domains.

pCAG ZNF91 [1-30] .pCAG ZNF91 was digested with NsiI and NotI and religated resulting in

the removal of the C-terminal 31-36 domains.

pCAG ZNF91 [1-2; 23-36.]. SacI. present in the gene-optimized synthesized ZNF91

(Genscript) after it was ligated into pCAG.

RECONstruction ZNF91 and ZNF93 Ancestral proteins

pCAG ZNF91hominine HA. ZNF91 representing the LCA of HCG cDNA sequence was

synthesized as a codon-optimized C-terminal HA pUC57 insert flanked by EcoRI and NotI sites

from Genscript. This insert was EcoRI and NotI digested and inserted into the pCAG EN vector

at the EcoRI NotI.

pCAG ZNF91great ape HA. ZNF91 representing the LCA of HCGO cDNA sequence was

synthesized as a codon-optimized C-terminal HA pUC57 insert flanked by EcoRI and NotI sites

from Genscript. This insert was EcoRI and NotI digested and inserted into the pCAG EN vector

at the EcoRI NotI.

Page 5: Plasmids - static-content.springer.com10.1038...pCAG ZNF91 [1-11]. pCAG ZNF91 was digested with XmaI and NotI and religated resulting in the removal of the C-terminal 12-36 domains

pCAG ZNF91macaque HA. ZNF91 macaque cDNA sequence was synthesized as a codon-

optimized C-terminal HA pUC57 insert flanked by EcoRV sites from Genscript. This insert was

EcoRV digested and inserted into the pCAG EN vector at the EcoRV.

pCAG ZNF93 Human.

pCAG ZNF93great ape. ZNF93 representing the LCA of HCGO cDNA sequence was synthesized

as a codon-optimized C-terminal HA pUC57 insert flanked by EcoRI and NotI sites from

Genscript. This insert was EcoRI and NotI digested and inserted into the pCAG EN vector at the

EcoRI NotI.

pCAG ZNF93macaque. ZNF93 representing the macaque cDNA sequence was synthesized as a

codon-optimized C-terminal HA pUC57 insert flanked by EcoRI and NotI sites from Genscript.

This insert was EcoRI and NotI digested and inserted into the pCAG EN vector at the EcoRI

NotI.

Luciferase Reporter constructs

pGL4cp_SV40. This is a standard promega vector pGL4CP with SV40 minimal promoter

inserted before the firefly luciferase cDNA. This plasmid was a gift from the Privalsky

lab and the sequence details can be found here:

http://microbiology.ucdavis.edu/privalsky/sites/default/files/plasmids/pGL4CP_reporters/

pGL4CP-SV40.txt

pGL4CP-SV40-DEST. An rfB cassette (Invitrogen) was introduced at a blunted BglII

upstream of SV40 in pGL4CP-SV40 to give a GATEYWAY compatible destination

vector(Onodera et al., 2012) to screen KAP1+ repeat sequences.

Page 6: Plasmids - static-content.springer.com10.1038...pCAG ZNF91 [1-11]. pCAG ZNF91 was digested with XmaI and NotI and religated resulting in the removal of the C-terminal 12-36 domains

pGL4cp-OCT4Enh-SV40. ~1500bp OCT4 human enhancer region was PCR cloned from

hESC gDNA with primers and ligated upstream of SV40 at the EcoRI blunted site.

OCT4Enh F-CACCCCCTCCACTATGGAACCTGCAC,

OCT4Enh R-  AGCTGGCCTTGGCTGAAGTG

>OCT4Enh

CCCTCCACTATGGAACCTGCACATCAGGTTCCTTGCTCCCCTCTCAACCAAAACTC

AGACATCTAATACCACGGTAGGCCCCGTTCTCCCTCCCCCACCTCCCTGGCCCAG

GCCTCCAGCCCTAGGCCCTGGGTGGGGAAAACCAGGGGGTGGGGGGTGTGGAG

AAAAAATATCTGACTTCAGGTTCAAAGAAGCCTGGGAGGGACTGGGGGAAGGGGG

CAGGACAATGGCCTTGGCTGGACAATCCCGGTCCCCAGAGGGGGCAGCTCTAAC

CCTAAACAAGTGCTCAACCCTTGAATGGGCCTGGATGGCTCCCCTGGGGACTGCT

TCCTGCTCCCCAACCCCCCAGTCCCAATCCCCTCACACAGAATCCCCTTCAGAGAC

GCTAAAAGGAGCTCCAGCAACCCCCCTCTGCAATCCCCTCAAAGACTGAGCCTCA

GACGGGCACCAAGGGCCCCCCACAGGGACCTAGGTATCTAGTTCCTCCTTCCTCT

GGGGGACTCAGGCGTCCAGCTTCATCGTGCGTCCCTCCCCGAGCCTGGCAGATT

GAGGGATGTGCTTTGTTTAGTGGGGCTGGCTGGCAGAAAGACGCAGAGGAGGTG

GAGAGTGATTTGTGGAGGCGTGCAGGAAGGCTGCCCTAAGCTCCCCTTCAGGGTC

TGTTTTTCTGGGCCTGGCCTGAGTATCCTGAGGCTCATGCTGCTGGTCTAGTGCTT

GATTCTGTTTGCAAGAGAATAGCCAACGGAATGCCTGTCTGTGAGGGATGATGTTT

GTCTGTCTGCTCCCAAAACTTGATCTCAGTGGAGGGCCTGGGGTAAGTCTGGGGG

CTCCAGAGGGGGCTCTGGGCCAGGGCTCCCCACAGCTTCGAAGGCCAGAAGGCC

AGGTCTGGACTGGGCACGCTGACCTCTGTCGACTTAAGTAAGGCTTCTCATTGCAG

Page 7: Plasmids - static-content.springer.com10.1038...pCAG ZNF91 [1-11]. pCAG ZNF91 was digested with XmaI and NotI and religated resulting in the removal of the C-terminal 12-36 domains

GCTCCAGGCTCAGCCCTGCCTGGGCTTGTCTGCTGAGGTCAGTGGCTCTATCTGC

CTTCTAAGGGGATGGGTGTCCCGTGGCCAGCTGTCTTCATCTTGGTGGCATCCGT

GAGTCTTTTGAGACTTTTCCCCCACTCTTATGTTGCCTCTGTTCGTGTGCCCATCTC

CTGTCTGTGTAGACTTTTTGAGCCTAATTGTATGCGTGCATTTCAATACCTGCCACA

GGTCTGCCGGAAGGTCTACAAGGCAGTGGGGTTGCAGCTGTGTTCACTTCTCGGC

CTTTAACTGCCCAAAAGGCAGGTAGATTATGGGGCCTGGTGGGGGTAGGAGGAAC

ATGCTTCGGAACAGGAGGAGGCCCCTCCCCAGCCATCTCAATCCCCAGGACAGAA

CCATCACGGCACCTTTGTCATGCATCTCTCTGCTGTCTGCCAAGAAGACGGCCTCT

CAGAGGAGGGGGAGGGGCAGGCCTGGGATTTGGCTGGAATCTCCACACCAGTGT

TTCTCAGCTTGCCATCCTCCAGGTTCCCCAAAAGCGCTCTTCCCAAGCCAGTCCAG

AGAGTCCCTGCTGCCCATTTTCCTAGTGGCTCCTAAAACACCTTCCCCAATTTCCC

CACTCAACACCACCCTCTTGTTTTTAGATTATAATTTGTACTGTAGGTGGTGTATTTC

TGGCCTGGGCAAGAGGCCCATTCCCGAGAGGGACGCAGACAAGGGGTGGGTGC

CTGGGTCCCTGGCTGCCTTGTGGCTGGATATGAGCCCAGTCAGGGGTCAGCCTCC

TGCATGCCTAGACTCCTAGCCGGCCCCCTTCTGGGGTGCTCAGGGCTGATGGGA

GGTTGAGGCAGGCTTTCCTTCCTTCTCACTGTCCTGTTATGCCTGAAGGGTAGGTG

GCTTCACTTCAGCCAAGGCCAGCT

pRL-TK. This is a standard promega vector for Renilla with TK minimal promoter

inserted before the renilla luciferase cDNA(Onodera et al., 2012) used as a transfection

efficiency control at a ratio of 10 firefly : 1 renilla to normalize the firefly luciferase

activity in each well (ff/ren).

pGL4cp-SINE-R-SV40. SINE-R domain from SVA_D full length (~XcmI site through

polyA) was synthesized as a gBlock (IDT) and ligated into pENTR/D/TOPO

Page 8: Plasmids - static-content.springer.com10.1038...pCAG ZNF91 [1-11]. pCAG ZNF91 was digested with XmaI and NotI and religated resulting in the removal of the C-terminal 12-36 domains

(Invitrogen). This was moved to pGL4CP-SV40-DEST with the LR GATEWAY

reaction.

pGL4cp-SVA_D-SV40 (full length). Synthesized pUC57-SVA_D (Genescript) was

EcoRV digested and ligated directly to pGL4CP-SV40 at a blunted EcoRI, upstream of

SV40.

> SVA_D-SV40 (full length)

TCTCCCTCTCCCTCTCTTTCCACGGTCTCCCTCTCCCTCTCTTTCCACGGTCTCCCT

CTCCCTCTCTTTCCACGGTCTTCCTCTGATGCCGAGCTGAAGCTGGACTGTACTGC

TGCCATCTCGGCTCACTGCAACCTCCCTGCCTGATTCTCCTGCCTCAGCCTGCCGA

GTGCCTGCGATTGCAGGCGCGCACTGCCACGCCTGACTGGTTTTCGTATTTTTTTG

GTGGAGACAGGGTTTTGCTGTGTTGGCCGGGCTGGTCTCCAGCTCCTAACCGCGA

GTGATCCGCCAGCCTCGGCCTCCCGAAGTGCTGGGATTGCAGACAGAGTCTCGTT

CACTCAGTGCTCAATGGTGCCCAGGCTGGAGTGCAGTGGCCTGATCTCGGCTCGC

TACAACCTCCACCTCCCAGCCGCCTGCCTTGGCCTCCCAAAGTGCCGAGATTGCA

GCCTCTGCCCGGCTGCCACGCCATCTGGGAAGTGAGGAGCGTCTCTGCCTGGCC

GCCCATCGTCTGGGATGTGAGGAGCCCCTCTGCCTGGCTGCCCAGTCTGGGAAG

TGAGGAGCGCCTCTTCCCAGCCGCCATCCCATCTAGGAAGTGAGGAGCGTCTCTG

CCCGGCTGCCCATCGTCTGAGATGTGGGGAGCGCCGCCGCCCTGTCTGGGACTT

GAGGAGCGCCTCTGCCCGGCCGCCCTGTCTGGGATGTGAGGAGCGCCTCTGCCC

GGCCGTGACCCTGTCTGGGAGGTGAGGAGCGTCTCTGCCCGGCCGCCCCGTCTG

AGAAGTGAGGAGCCCCTCCCCCCAGCAGCCGCCCCGTCTGAGAAGTGAGGAGCC

CCTCCGCCCGGCAGCCACCCGATCTGGGAAGTGAGGAGCGTCTCCGCCCGGCAG

Page 9: Plasmids - static-content.springer.com10.1038...pCAG ZNF91 [1-11]. pCAG ZNF91 was digested with XmaI and NotI and religated resulting in the removal of the C-terminal 12-36 domains

CCGCCCCGTCCGGGAGGGAGGTGGGGGGTCAGCCCCCGCCCGGCCAGCCGCC

CCGTCCGGGAGGTGGGGGCGCCTCTGCCCGGCCACCCCTTCTGGGAAGTGAGG

AGCCCCTCTGCCCGGCCACCACCCCGTCTGGAAGGTGTACCCAACAGCTCACTGA

GAACGGGCCATGATGACAATGGCGGTTTTGTGGAATAGAAAAGGGGGAAAGGTGG

GGAAAAGATTGAGAAATCGGATGGTTTCTGTGTCTGTGTAGAAAGAAGTAGACATG

GGAGACTTTTCATTTTGTTCTGTACTAAGAAAAATTCTTCTGCCTTGGGATCCTGTT

GATCTATGACCTTACCCCCAACCCTGTGCTCTCTGAAACATGTGCTGTGTCCACTC

AGGGTTAAATGGATTAAGGGCGGTGCAAGATGTGCTTTGTTAAACAGATGGTCGAA

GGCAGCATGCTCGTTAAGAGTCATCACCACTCCCTAATCTCAAGTACCCAGGGACA

CAAACACTGCGGAAGGCCACAGGGTCCTCTGCCTAGGAAAACCAGAGACCTTTGT

TCACTTGTTTATCTGCTGACCTTCCCTCCACTATTGTCCTATGACCCTGCCAAATCC

CCTCTGCGAGAAACACCCAAGAATGATCAATAAAAAAAAAAAAGAAAAGAA

no hex. pUC57 SVA_D was digested with EcoRV/BglI resulting in ‘ChopD.2’ a

fragment was 1419bp fragment lacking the Hexamer (CCCTCTn)domain. This was

ligated into pGL4CP-SV40, upstream of the SV40 promoter.

no hex/Alu. pUC57 SVA_D was digested with EcoRV/EagI resulting in ‘ChopC.1’ a 787

bp fragment consisting of the 3’ side of SVA and was ligated into pGL4CP-SV40,

upstream of the SV40 promoter.

partial VNTR. pGL4cp-SVA_D-SV40 (full length) was digested with Tth111I which

cuts twice liberating a 401bp fragment, and leaving 1136bp of the SVA_D which was

self ligated as ‘VNTR delta C’

Page 10: Plasmids - static-content.springer.com10.1038...pCAG ZNF91 [1-11]. pCAG ZNF91 was digested with XmaI and NotI and religated resulting in the removal of the C-terminal 12-36 domains

no VNTR. ‘E’. pGL4cp-SVA_D-SV40 (full length) was digested with Tth111I/XcmI

which removes 401bp+321bp fragments, and leaving 813bp of the SVA_D which

was self ligated as ‘VNTR delta D’.

no SINE-R. pUC57 SVA_D was digested with EcoRV/EagI resulting in ‘Chop C.3’ a s

686bp 5’ SVA fragment. This was ligated into pGL4CP-SV40, upstream of the SV40

promoter.

partial Sine-R. pGL4cp-SVA_D-SV40 (full length) was digested with PpuMI/XcmI

which removes a 350bp fragment, and leaving 1187bp of the SVA_D which was self

ligated as VNTR delta A.

Alu-VNTR (1-1.5). pUC57-SVA_D was digested with PfIMI/Tth111I  generating  a  180bp    Alu  domain  and  VNTR  domain  containing  fragment  including  63bp  of  the  VNTR  domain  or  about  1.5  VNTR  repeat  units.  This  was  cloned  upstream  of  OCT4Enh  at  EcoRI  blunt  in  pGL4cp-OCT4Enh-SV40 as  ‘VNTR  OCT4Enh  F'6’.      VNTR (8.5-15). pUC57-SVA_D was digested with Tth111I/XcmI   generating   a   323bp    VNTR   domain   containing   fragment   or   about   6.5   VNTR   repeat   units.   This   was   cloned  upstream  of  OCT4Enh  at  EcoRI  blunt  in  pGL4cp-OCT4Enh-SV40 as ‘VNTR  OCT4Enh  D3’.    

VNTR (1.5-8.5). pUC57-SVA_D was digested with Tth111I/PfIMI   generating   a   220bp  VNTR   domain   containing   fragment   or   about   7   VNTR   repeat   units.   This   was   cloned  upstream  of  OCT4Enh  at  EcoRI  blunt  in  pGL4cp-OCT4Enh-SV40 as ‘VNTR  OCT4Enh  F4’.     Alu-VNTR (1-7.5). pUC57-SVA_D was digested with Tth111I   generating   a   401bp  Alu  and  VNTR  domain  containing  fragment  or  about  7.5  VNTR  repeat  units.  This  was  cloned  upstream  of  OCT4Enh  at  EcoRI  blunt  in  pGL4cp-OCT4Enh-SV40 as ‘VNTR  OCT4Enh  C6’.    

Page 11: Plasmids - static-content.springer.com10.1038...pCAG ZNF91 [1-11]. pCAG ZNF91 was digested with XmaI and NotI and religated resulting in the removal of the C-terminal 12-36 domains

VNTR (1.5-15). pUC57-SVA_D was digested PfImI/XcmI with generating   a   544bp  

VNTR   domain   containing   fragment   or   about   13.5   VNTR   repeat   units.   This  was   cloned  

upstream  of  OCT4Enh  at  EcoRI  blunt   in  pGL4cp-OCT4Enh-SV40 as ‘VNTR OCT4Enh

E2’.

pGL4cp-L1PA4-5’UTR-SV40. Synthesized ~500bp L1PA4 (gBLOCk, IDT) was cloned

into pENTR-D/TOPO (Invitrogen) and transferred by GATEWAY reaction to pGL4CP-

SV40-DEST.

> L1PA4-5’UTR

GGGGGAGGAGCCAAGATGGCCGAATAGGAACAGCTCCGGTCTACAGCTCCCAGC

GTGAGTGACACAAAAGACGGGTGATTTCTGCATTTCCAACTGAGCTTTGAAGAGAG

TAGTGGTTCTTCCAGCATGCAGCTTGAGATCTGAGAACAGGCAGACTGCCTCCTCA

AGTGGGTCCCTTACCCCCGAGTAGCCTAACTAGGAGGCACCCCCGAGTAGGGGC

AGACTGACACCTCACATGGCAGGGTACTCCTCTGAGACAAAACTTCCAGAGGAAC

GATCAGGCAGCAGCATCTGCAGTTCACCAATATCTGCTGTTCTGCAGCCACTGCTG

CTGATACCCAGGCAAACAGGGTCTGGAGTGGACCTCTAGCAAACTCCAACAGACC

TGCAGCTGAGGGTCCTGTCTGTTAGAAGGAAAACTAACAAACAGAAAGGACATCCA

CACCAAAACCCACCTGTAC

129L1PA4. Synthesized 129bp L1PA4 (gBLOCk, IDT) with flanking sequence from L1HS

at the site of deletion (PvuII sites) was cloned into pENTR-D/TOPO (Invitrogen). This

was next PvuII digested and ligated into pGL4cp-OCT4Enh-SV40 at EcoRI blunt

upstream of OCT4Enh and SV40 promoter.

Page 12: Plasmids - static-content.springer.com10.1038...pCAG ZNF91 [1-11]. pCAG ZNF91 was digested with XmaI and NotI and religated resulting in the removal of the C-terminal 12-36 domains

129scramble. Same as 129L1PA4 except the 129bp element was scrambled before being

syntheized as a gBlock.

51L1PA4. Same as 129L1PA4 except a 51bp region, beginning at the Persikov predicted

18mer/ Zn Finger domains 8-13 was chosen as the anchor point such that 5’-3’ DNA

would be bound by a C-N terminal oriented protein where each finger binds to 3bp.

129Δ51L1PA4. Same as 129L1PA4 except a 51bp region in 51L1PA4 was deleted from the

129bp element at the gBlock synthesis step.

Retrotransposition reporter constructs

L1HS. A gift from the Kazazian lab (JHMI). Thanks to Dustin Hancks for sending plasmids plasmid map thanks to John Goodier. Hancks description: 99 RPS EGFP- hot L1, L1-RP (Kimberland et al 1999), cloned into pCEP4 (Invitrogen) lacking the CMV promoter (Moran et al 1996), L1 is driven by its own promoter in the 5’-UTR, puro selection, digest with NotI/Bstz17I, two bands 6kb (the L1) and 11 kb backbone + EGFP retrotransposition indicator cassette, (Ostertag et al 2000 NAR).

L1HS+129bp. L1HS plasmid was Digested with NotI/Bstz17I and ligated to pCAG EN

at NotI/EcoRV. Next, pCAG-L1HS was digested with PvuII, and an insert from the PvuII

fragment from 129L1PA4 was ligated. Finally, pCAG-L1HS-129L1PA4 was digested with

NotI/AleI and ligated back to the pCEP4 based vector that originally contained L1HS

insert. This final plasmid is meant to represent the L1PA4/3 ancestral state.

L1HS+129 scr. Same as L1HS+129bp, except the 129L1PA4 fragment is the 129scramble

insert.

Page 13: Plasmids - static-content.springer.com10.1038...pCAG ZNF91 [1-11]. pCAG ZNF91 was digested with XmaI and NotI and religated resulting in the removal of the C-terminal 12-36 domains

SVA_D and L1PA4 domain analysis was performed with a combination of restriction

enzymes and gBLOCK synthesized fragments.

VNTR-OCT4Enh-pGL4CP-SV40 was created by removing a 545bp, 13.5 repeat-

containing VNTR subunit fragment between PflMI and XcmI. Selected KRAB ZNFs

were obtained from cDNA, synthesized by Genscript, or subcloned from I.M.A.G.E.

vector through ATCC and ligated into pCAG_EN (Cepko Lab, Addgene) either directly

at EcoRI blunt site or through Gateway ENTRY cloning using pCAG_EN with an rfA

cassette integrated at the EcoRI site.

Extra  sequences  for  ZNF  paper    >ZNF91  Human  atgcctggaacacctgggagcctggagatgggcctgctgacctttcgggatgtcgctattgagtttagtcctgaagagtggcagtgcctggacaccgcccagcagaacctgtacaggaatgtgatgctggagaactatcgcaatctggccttcctgggaatcgctctgtccaagcctgacctgattacctacctggagcagggcaaggaaccctggaacatgaaacagcacgagatggtggatgaacccacaggcatctgcccccacttccctcaggacttttggccagagcagtcaatggaagatagcttccagaaggtcctgctgagaaagtacgagaaatgcgggcacgaaaatctgcagctgaggaagggatgtaaaagcgtggatgagtgcaaggtccacaaagaaggctataacaagctgaatcagtgtctgaccacagcccagtctaaagtgttccagtgcgggaagtacctgaaggtcttctataagttcctgaacagtaacaggcacacaatccgccatactggcaagaaatgcttcaagtgtaagaaatgcgtgaagagcttctgtatcaggctgcacaagacacagcataaatgcgtctacattactgagaagtcctgcaagtgtaaagagtgcgaaaaaacattccactggagctccactctgaccaaccacaaggagatccatactgaagacaagccttacaaatgtgaggaatgcggaaaggcctttaaacagctgtccaccctgactacccacaagatcatttgcgccaaggaaaaaatctataagtgtgaggaatgcggcaaggccttcctgtggtctagtacactgactaggcacaagcgcattcatacaggcgagaagccctataaatgcgaggaatgcgggaaggctttttctcactcaagcaccctggcaaagcacaaacgcatccatacaggagagaagccttataaatgtgaagagtgcggcaaggccttcagccgatcctctactctggctaagcacaaaagaatccacaccggggagaagccatacaagtgtaaagaatgcggaaaagcattttccaacagttcaaccctggccaatcacaagattacccatacagaggaaaaaccctataagtgtaaagaatgcgataaggccttcaaaaggctgagcactctgaccaagcacaaaatcattcatgcaggcgagaagctgtataagtgcgaagagtgcgggaaggccttcaaccgcagctccaatctgaccatccacaagttcatccacaccggcgaaaagccttataaatgtgaggagtgtggcaaggcttttaattggtctagttcactgactaagcacaaacgattccatacccgggagaagcccttcaagtgcaaggaatgcgggaaagcattcatctggagctccacactgactcgacacaagcggattcataccggggagaagccctataaatgtgaagaatgtggaaaggcctttaggcagtctagtaccctgacaaaacataaaatcattcataccggcgagaagccttataaatttgaagagtgtgggaaggcttttcgccagtccctgacactgaacaagcataagatcattcattctcgagaaaaaccttataaatgcaaagaatgcggcaaggcattcaaacagttttccaccctgacaactcacaagatcattcatgctggaaagaaactgtacaaatgcgaggaatgcggcaaagcattcaatcactcaagctccctgagtacacacaagatcattcatactggggagaagtcatataagtgcgaagaatgcggaaaggcctttctgtggtcctccacactgcggagacacaaaagaatccacactggagaaaagccctataaatgtgaggagtgcgggaaggctttctctcactcaagcgccctggccaaacacaaaagaatccatactggagagaagccttataagtgcaaggagtgtggcaaggccttcagcaacagctctacactggcaaatcacaagatcactcataccgaggaaaaaccatataaat

Page 14: Plasmids - static-content.springer.com10.1038...pCAG ZNF91 [1-11]. pCAG ZNF91 was digested with XmaI and NotI and religated resulting in the removal of the C-terminal 12-36 domains

gcaaggaatgcgacaagaccttcaaaaggctgtccacactgactaaacataagatcattcatgccggggagaagctgtataagtgtgaagaatgcggaaaggctttcaaccgcagttcaaatctgacaatccacaagttcatccatactggggaaaaaccctataaatgtgaggaatgcgggaaggcattcaactggagctcctctctgactaaacataagagaatccatacaagggagaagcctttcaagtgcaaagagtgcggcaaagcctttatctggagttcaaccctgacaagacacaagaggattcataccggagagaagccatataagtgcgaggagtgcggcaaggcttttagccggagcagcaccctgaccaagcacaaaactatccatactggcgaaaaaccttacaaatgcaaagaatgcggaaaggcattcaaacactctagtgcactggctaaacacaagattatccacgcaggcgagaagctgtacaagtgcgaagaatgtgggaaggcctttaaccagtcaagcaatctgaccacacacaaaatcattcataccaaggagaaacccagcaagtccgaggaatgcgataaagccttcatctggtcctctacactgactgaacacaagcgcattcatacccgagagaagacatacaagtgcgaggaatgcggcaaggctttttctcagcctagtcacctgactacccataaacggatgcacaccggagagaagccatacaaatgcgaagagtgcggcaaggcattctcccagagttcaaccctgacaactcataagatcattcacacaggggaaaaaccttacaagtgcgaagagtgtggaaaggcctttagaaaaagctccaccctgacagaacataagatcattcacaccggcgagaagccatataagtgtgaggagtgtgggaaggctttctctcagtctagtactctgacccgccatactcgaatgcatactggagagaaaccttataagtgcgaagagtgtggcaaggcctttaacagatcaagcaagctgaccacacataaaatcattcacaccggggagaagccctacaagtgtgaggagtgtggaaaggctttcatctcctctagtactctgaatgggcataagcggattcacaccagagagaagccatataaatgcgaagagtgcggaaaggcctttagccagtcaagcacactgactcggcataagagactgcacaccggcgagaagccctacaaatgtggcgagtgtggcaaggcattcaaagagtcctctgccctgactaagcacaagattatccacaccggcgagaagccttacaaatgtgaaaagtgcgggaaagcctttaaccagagttcaatcctgacaaatcacaagaaaatccacaccattacaccagtgattcccctgctgtgggaggctgaagcaggcgggtcaaggggacaggagatggagactattctggctaacaccgtgaaacctctgctgtactga    >ZNF91  HCG  GAATTCATGCCTGGGACACCTGGTAGCCTGGAAATGGGCCTCCTGACTTTTAGGGATGTGGCAATCGAGTTCTCACCTGAAGAGTGGCAGTGCCTGGACACCGCCCAGCAGAACCTGTACCGGAATGTGATGCTGGAGAACTATCGCAATCTGGCTTTCCTCGGCATCGCACTCAGCAAGCCTGACCTGATTACATACCTGGAGCAGGGAAAGGAACCCTGGAACATGAAACAGCACGAGATGGTGGATGAACCCACTGGCATCTGTCCCCACTTCCCTCAGGACTTTTGGCCAGAGCAGTCAATGGAAGATAGCTTTCAGAAGGTCCTGCTCCGAAAGTACGAGAAATGCGGGCACGAAAACCTCCAGCTCAGGAAGGGTTGTAAATCCGTGGATGAGTGCAAGGTCCACAAAGAAGGATATAACAAGCTGAATCAGTGTCTCACCACAGCTCAGTCTAAGGTGTTCCAGTGCGGGAAGTACCTGAAGGTCTTCTATAAGTTTCTCAACAGTAACCGGCACACAATCCGCCATACTGGAAAGAAATGCTTCAAGTGTAAGAAATGCGTGAAGAGCTTCTGTATCCGGCTGCACAAGACACAGCATAAATGCGTCTACATTACTGAGAAGTCCTATAAGTGTAAAGAGTGCGAAAAAACCTTCCACTGGAGCAGCACCCTGACCAACCACAAGGAGATCCATACAGAAGACAAGCCCTACAAATGTGAGGAATGCGGCAAGGCCTTTAAACAGCTGTCCACCCTCACTACCCACAAGATCATTTGTGCCAAGGAGAAAATCTATAAGTGCGAAGAATGCGGCAAGGCCTTCCTGTGGTCTAGTACACTCACTCGGCACAAGCGCATTCATACAGGCGAGAAACCTTATAAGTGTGAGGAATGCGGAAAGGCATTTTCTCACTCAAGCACCCTGGCCAAGCACAAACGCATCCATACAGGGGAGAAGCCTTATAAATGCGAAGAGTGTGGCAAGGCCTTCAGCCACTCCTCTACTCTCGCAAAGCACAAACGAATCCATACCGGCGAGAAGCCATACAAGTGTAAAGAATGCGGAAAAGCCTTTTCCAACAGTTCAACCCTGGCTAATCACAAGATTACCCATACAGAGGAAAAACCCTATAAGTGTAAAGAATGCGATAAGGCCTTCAAACGGCTGAGCACTCTCACCAAGCACAAAATCATTCATGCAGGGGAGAAGCTGTATAAGTGCGAGGAGTGTGGCAAGGCCTTCAACCGCAGCTCCAATCTCACCATCCACAAGTTTATTCATACAGGCGAGAAGCCTTATAAATGTGAAGAGTGCGGAAAGGCCTTCAACTGGTCTAGTTCACTCACTAAACATAAACGAATCCATACCAGGGAGAAGCCATTCAAGTGCAAGGAGTGCGGCAAGGCCTTCATCTGGAGCAGCACCCTGACGCGACACAAGAGGATTCATACCGGGGAGAAACCTTATAAATGCGAAGAGTGTGGAAAAGCATTCAGGCAGTCTAGTACCCTGACAAAACATAAAATCATTCATACTGGTGAAAAGCCTTATAAATTCGAGGAATGCGGAAAGGCCTTTAGACAGTC

Page 15: Plasmids - static-content.springer.com10.1038...pCAG ZNF91 [1-11]. pCAG ZNF91 was digested with XmaI and NotI and religated resulting in the removal of the C-terminal 12-36 domains

CCTGACCCTCAACAAGCATAAGATCATTCATTCTCGGGAGAAACCTTACAAATGCAAAGAATGCGGGAAGGCTTTCAAACAGTTTTCCACCCTGACAACTCACAAGATCATTCATGCTGGGAAGAAACTCTACAAGTGCGAGGAATGTGGCAAGGCTTTTAATCAGTCAAGCTCCCTGAGTACACACAAGATCATTCATACTGGCGAGAAGTCATATAAGTGCGAGGAGTGCGGAAAGGCTTTCCTGTGGTCTAGTACACTCAGGAGACACAAGAGAATCCACACTGGGGAGAAGCCCTATAAATGCGAGGAGTGCGGAAAGGCCTTTAGCCACTCAAGCACCCTGGCCAAACACAAAAGAATCCATACTGGTGAGAAGCCTTACAAATGTAAGGAATGCGGCAAGGCTTTCTCTAACAGCTCCACCCTGGCAAATCATAAGATCACTCATACCGAGGAAAAACCATATAAATGCAAAGAATGCGACAAGGCCTTTAAACGGCTGTCTACACTCACTAAACATAAGATCATTCATGCCGGGGAGAAGCTGTATAAGTGTGAGGAATGCGGAAAGGCCTTCAACCGCAGTTCAAATCTCACAATTCATAAGTTTATTCACACAGGCGAGAAGCCCTATAAATGTGAAGAGTGTGGAAAAGCATTCAACTGGAGCAGCAGCCTGACCAAGCACAAAAGAATCCATACACGGGAGAAGCCTTTCAAGTGCAAGGAATGCGGCAAAGCCTTTATCTGGAGTTCAACCCTGACAAGACACAAGCGGATTCATACAGGCGAAAAACCATATAAGTGCGAGGAGTGCGGCAAGGCATTCAGCCACAGCTCCACTCTGACCAAGCACAAAACTATTCACACCGGCGAAAAACCTTACAAATGTAAAGAGTGCGGCAAGGCCTTCAAGCACTCTAGTGCCTTAGCTAAACATAAAATTATCCACGCAGGGGAGAAGCTCTATAAGTGTGAAGAATGTGGAAAGGCTTTTAACCAGTCAAGCAATCTGACCACCCATAAGATCATTCATACCAAGGAGAAACCCAGCAAGTCCGAGGAATGCGATAAAGCCTTTATCTGGTCCTCTACACTGACTGAACACAAGCGCATTCATACCCGAGAGAAGACATATAAGTGCGAGGAGTGTGGAAAGGCTTTCAGCCAGCCTAGTCACCTGACTACCCATAAGAGGATGCACACAGGCGAGAAGCCATATAAATGTGAAGAATGCGGAAAGGCATTTTCCCAGAGTTCAACTCTGACAACTCATAAGATCATTCACACCGGTGAGAAACCTTATAAATGTGAGGAGTGTGGCAAGGCCTTCAGGAAAAGCTCCACCCTGACAGAACACAAAATTATCCACACTGGTGAGAAGCCATACAAATGCGAAGAATGCGGAAAGGCTTTTTCTCAGTCTAGTACTCTGACCCGCCATACTCGAATGCACACCGGTGAAAAGCCCTATAAATGTGAAGAATGTGGTAAAGCATTCAACAGGTCAAGCAAGCTGACCACACATAAAATCATCCATACTGGCGAGAAGCCTTACAAGTGCGAGGAATGCGGAAAGGCTTTTATCTCCTCTAGTACACTGAATGGGCATAAGAGGATTCACACTAGAGAGAAGCCATACAAATGTGAAGAGTGCGGCAAGGCCTTCAGCCAGTCAAGCACACTGACTAGGCATAAGAGACTCCACACCGGGGAGAAGCCCTACAAATGTGGCGAGTGTGGAAAGGCCTTTAAAGAGTCCTCTGCTCTGACTAAGCACAAAATCATTCACACCGGGGAGAAGCCTTACAAATGTGAAAAGTGCGGCAAGGCCTTCAACCAGAGTTCAATCCTGACAAATCACAAGAAAATCCACACCATTACACCAGTGATTCCCGCACTCTGGGAGGCAGAAGCCGGCGGGAGCAGGGGGCAGGAGATTGAAACCATTCTCGCAAATACCGTGAAACCCAGACTCTATTACCCATACGATGTTCCAGATTACGCTTGAGCGGCCGC    >ZNF91  HCGO  GAATTCATGCCCGGAACCCCCGGAAGCCTGGAGATGGGCCTCCTGACATTTCGAGACGTCGCTATTGAGTTTAGTCCTGAAGAGTGGCAGTGTCTGGACACCGCCCAGCAGAACCTGTACCGCAATGTGATGCTGGAGAACTATCGAAATCTGGCCTTCCTCGGAATCGCTCTCTCCAAGCCTGACCTGATTACATACCTGGAGCAGGGGAAAGAACCCTGGAACATGAAGCAGCACGAGATGGTGGATGAACCAACTGGCATCTGTCCCCACTTCGCTCAGGACTTTTGGCCCGAGCAGAGTATGGAAGATTCATTCCAGAAGGTCCTGCTCAGGAAGTACGAGAAATGCGGGCACGAAAACCTCCAGCTCAGAAAGGGTTGTAAATCTGTGGATGAGTGCAAGGTCCACAAAGAAGGGTATAACAAACTGAATCAGTGTCTCACCACAGCCCAGTCCAAGGTGTTCCAGTGCGGCAAGTACCTGAAGGTCTTCTATAAGTTCCTCAACTCTAACAGGCACACCATCAGACATACAGGAAAGAAATGCTTCAAGTGTAAGAAATGCGTGAAGAGTTTTTGTATCCGGCTGCACAAAACTCAGCATAAGTGCGTCTACATTACCGAGAAATCCTATAAGTGTAAAGAGTGCGAAAAGACCTTCCACTGGAGCAGCACCCTGACTAACCACAAGGA

Page 16: Plasmids - static-content.springer.com10.1038...pCAG ZNF91 [1-11]. pCAG ZNF91 was digested with XmaI and NotI and religated resulting in the removal of the C-terminal 12-36 domains

GATCCATACAGAAGACAAGCCTTACAAATGTGAGGAATGCGGAAAGGCCTTTAAACAGCTGTCCACTCTCACTACCCACAAGATCATTTGTGCTGAGGAAAAAATCTACAAGTGCGAGGAATGCGGGAAAGCATTCCTGTGGTCTAGTACCCTCACAAGGCACAAGAGAATCCACACAGGCGAGAAGCCCTATAAGTGCGAAGAATGCGGAAAGGCATTTCGCCAGTCAAGCACTCTGACCAAGCACAAAATCATTCATACTGGGGAGAAGCCTTATAAATTCGAGGAATGCGGCAAGGCCTTCAGGCAGTCTCTGACCCTCAACAAGCATAAGATCATTCATAGTCGCGAGAAGCCATACAAGTGTAAAGAATGCGGCAAGGCTTTCAAACAGTTTAGCACCCTGACAACTCACAAGATCATTCATGCCGGCAAGAAACTCTATAACTGCGAAGAGTGCGGAAAGGCTTTCAATCAGTCCTCTAGTCTGTCAACTCATAAAATCATTCATACCGGGGAGAAGAGCTATAAGTGTGAAGAGTGCGGCAAGGCCTTCCTGTGGTCAAGCACACTCTCTCGGCACAAGCGCATCCATACTGGCGAGAAGCCCTGCAAATGTGAGGAATGTGGAAAGGCCTTCAGCCACTCCTCTACCCTGGCTAAGCACAAAAGAATCCACACAGGGGAGAAACCTTATAAATGCAAAGAATGTGGCAAGGCCTTCAGCAACAGCAGCACCCTCGCCAATCACAAAATTACACATACTGAGGAAAAACCTTATAAATGTAAGGAGTGTGGCAAGGCCTTCAAACGACTGTCTACCCTCACAAAACATAAAATCATTCATGCAGGCGAGAAGCTGTATAAGTGCGAGGAGTGTGGAAAGGCCTTCAACAGAAGCTCCAATCTCACCACACACAAGTTTATCCATACCGGGGAGAAGCCCTATAAGTGTGAAGAATGTGGCAAGGCCTTCAACTGGTCTAGTTCACTGACAAAACATAAAAGAATCCACACTCGCGAGAAACCTTTCAAGTGCAAGGAGTGCGGCAAGGCCTTCATCTGGAGCAGCACCCTCACACGACACAAGAGGATTCATACAGGCGAGAAGCCATATAAGTGCGAGGAATGCGGAAAGGCTTTTAGCCACTCTAGTACACTGACTAAGCACAAAACAATCCATACTGGGGAGAAGCCCTACAAATGCAAAGAATGCGGCAAGGCCTTCAAGCACCCTTCCACCCTCGCCAAACATAAGATTATCCACGCAGGCGAGAAGCTCTATAAGTGTGAGGAGTGCGGAAAGGCCTTTAACCAGTCAAGCAATCTGACAACACACAAGATCATTCATACAAAGGAGAAACCATCAAAAAGCGAGGAATGCGACAAGGCTTTCATCTGGTCCTCTACCCTGACAGAACACAAGAGAATCCACACTAGGGAGAAGACCTATAAGTGTGAGGAATGTGGCAAGGCATTTTCCCAGCCATCTCACCTGACAACTCATAAGAGGATGCACACAGGTGAGAAGCCCTATAAATGCGAAGAATGCGGCAAGGCCTTCAGCCAGAGTTCAACTCTGACCACACATAAGATCATTCACACCGGCGAGAAGCCCTATAAATGTGAAGAATGCGGCAAGGCCTTCAGGAAGAGCAGCACCCTGACCGAGCACAAGATCATTCACACAGGCGAAAAGCCTTATAAATGCGAAGAATGCGGCAAGGCCTTCAGCCAGTCTAGTACACTGACTAGACATACACGGATGCACACTGGCGAGAAGCCATATAAATGCGAGGAGTGCGGAAAGGCTTTTAACAGATCAAGCAAACTGACTACCCATAAAATTATTCACACAGGAGAAAAACCCTATAAATGCGAGGAATGTGGCAAGGCCTTCATCAGCAGCAGCACCCTGAACGGCCATAAGAGAATCCACACTCGGGAGAAACCATATAAGTGTGAAGAATGCGGAAAGGCTTTTTCCCAGTCAAGCACCCTGACACGCCATAAGCGACTCCACACCGGTGAAAAGCCATATAAATGTGGCGAATGCGGAAAGGCATTCAAAGAGTCCTCTGCCCTGACCAAGCATAAAATCATCCATACCGGCGAGAAGCCTTACAAATGTGAAAAGTGCGGCAAGGCCTTCAACCAGAGTTCAATCCTGACCAATCATAAGAAAATTCACACCGCAAACCCATGTACATGGGGCGGATGCGGGGGTAGCTTCAATAGGAGCAGCACCTTCACCGATCATGAAGTGATCCACACAGGGGTCAAGCTGTATAAATGTGAAGAATGCGGCAAGAGCTTCTTCTGGAGCAGCGCCCTCTCCCGCCACAAGAAAATCCACCCCGGCGTGCAGCCTTACAAGCAGGAGAAATTCGGCAAGGCTTTTAACCAGTTTTCCCACCTGAGCACAGATAAGATTACCCATATTGGAGAGAAGTCATACAAATGCGAGTACCCATACGATGTTCCAGATTACGCTTGAGCGGCCGC    >ZNF91  rhesus  macaque  GATATCATGGGCCTCCTCACCTTTCGGGATGTGGCAATCGAGTTTAGCCTGGAAGAGTGGCAGAGACTGGACGCCGCTCAGCAGAACCTGTATCGCAACGTGATGCTGGAGAACTACCGAAATCTGGCATTCCTCGGAATCGCCGTCAGCAAGCCTGACCTGATTACCTGCCTGGAGCAGGGGAAAGAACCCTGGAATATGAAGAGA

Page 17: Plasmids - static-content.springer.com10.1038...pCAG ZNF91 [1-11]. pCAG ZNF91 was digested with XmaI and NotI and religated resulting in the removal of the C-terminal 12-36 domains

CACGAGATGGTGGATAAGGCCCCCGCTATCTGCCCTCACTTCGCCCAGGACTTTTGGCCCGAGCAGGGCATGGAAGATAGCTTCCAGAAGGTGCTGCTCCGGAAGTACGAGAAATGCGGACACGAAAACCTCCAGCTCCGCAAGTCCTGTAAATCTGTGGACGAGTGCAAAGTCCACCGAGAAGGATACTCTAAGCTGAATCAGTGTTTCACCACAACTCGGAGTAAGGTGTTTCAGTGCGGGAAGTATCTGAAAGTCTTCCATAAGTTTCTCAACTCCAATCGCCACATGATCAGCATTACCGAGAAATCCTACAAGTGCAAAGAGTGTGAAAAGATTTTCCACTGGAGCAGCACCCTGACAAACCACAAGAAAATTCATACAGAGGATAAGCCATACAAGTGCGAGGAATGTGGCAAGGCCTTCAAGCAGCTGTCTACCCTCATCACACACAAGATTACATGTGCCGAGGAAAAAATCTACAAATGCGAGGAATGTGGAAAAACTTTCAACCAGTCTAGTAATCTGACCACACACAAGATCATTCATAGTGGGCAGAAGCCCTATAAATGCGAGCAGTGTGGCAAGGCCTTCATCTGGCTGAGCACTCTCACCGGTCATAAGAAAATTCATACCAGGGGCAAGCCATGTAAATACGAGGAATGCGGCAAGGCCTTCATCTGCTCCCCCACACTGACTAGGAGAAAGCGGATTCACACCGAGGAAAAGCTCTATAAGTGTGAGGAATGTGGAAAAGCATTTCTGTGGTCAAGCACCCTCACACGGCACAAGCGCATCCATACAGGCGAGAAGCCTTATAAGTGCGAGGAATGTGGAAAGGCCTTCAGGCAGTCCTCTACTCTGACCAAGCACAAAAGAATCCATACTGGGGAGAAGCCATATAAATGTGAAGAGTGCGGCAAGGCCTTCAGGCAGTCACTGACACTCACTAAGCACAAAATCATTCATAGCGGCGAGAAGCACTACAAGTGCAAAGAATGTGGAAAGACATTCAAACAGTTTTCTACTCTGACTACCCACAAGATCATTCATGCCGGGAAGAAATGCTATAAATGTGAAGAATGTGGCAAGGCCTTCAACCAGAGTTCAAGCCTCTCCACCCATAAAATCATTCATACAGGCGAGAAGTCTTACAAATGTGAGGAATGCGGAAAGGCTTTTATCTGGTCCTCTACCCTGACACGACACAAGAGGATTCATACCGGGGAGAAGCCCTGCAAATGTGAGGAGTGCGGCAAGGCCTTCAGCCACAGTTCAACACTCGCTAAACACAAAAGAATCCACACTGGCGAGAAACCTTATAAGTGTAAAGAATGCGGAAAGGCCTTTTCTAACAGCTCCACTCTGACCAATCACAAAATCACACATACTGAGGAAAAGCCATACAAGTGTAAAGAGTGCGGCAAGGCTTTCAAACGCCTGTCCACCCTCACAAAACATAAAATCATTCATGCAGGGGAAAAGCTGTATAAATGTGAAGAATGCGGCAAGGCCTTCAACAGGTCTAGTAATCTCACAGCCCATAAGATCATTCATACTGGCGAGAAGCCATACAAGTGTGAAGAGTGCGGAAAGGCATTCATCTGGTCAAGCTCCCTGACCAAGCACAAACGGATTCATACACGCGAGAAACTCTTCAAGTGTAAAGAATGCGACAAGGCCTTTATCTGGTCTAGTACTCTCACCAGACACAAGCGGATTCATACAGGCGAAAAACCCTATAAATGTGAGGAATGCGGCAAGGCCTTCAACCACTCAAGCACACTGACTAAGCACAAAACTATCCATACCGGCGAGAAGCCTTACAAATGCAAAGAATGCGGAAAGCCATTCAAAATCCCCTCCACCCTGGCCAAGCACAAAATCATCTACGCTGGGGAGAAGCTCTACAAGTGTGAGGAATGCGGCAAGGCCCTGAACCAGTCCTCTAATCTCACAACTCACAAAGTGATCAACACAAAGGAGAAACCAAACAAGTCAGAAGAGTGTGGCAAGGCCTTTGTCTGGAGTTCAACCCTGACAGAGCACAAGCGAATCCATACCAGGGAAAAGCCTTATAAGTGTGAAGAGTGTGGCAAGGCTTTCAGTCAGCCTTCACACCTGACCACACATAAGAGAATGCACACAGGCGAGAAGCCATACAAATGCGAAGAATGCGGCAAGGCCTTCAGCCAGAGCAGCACCCTGACTACCCATAAGATCATTCACACAGGGGAGAAGCCTTATAAGTGTGAGGAGTGCGGTAAAGCCTTCCGGAAGTCTAGTACTCTGACCGAGCATAAGATCATTCACACCGGCGAAAAGCCATATAAGTGCGAAGAGTGCGGAAAGGCTTTTAGCCAGTCAAGCACACTGACTCGCCATAAGCGAATGCACACAGGTGAAAAGCCATATAAATGCGAAGAATGCGGCAAGGCCTTCAACAGATCCTCTAAACTGACAACTCACAAAATCATTCACACCGGCGAGAAGCCCTACAAATGTGAAGAGTGTGGAAAGGCTTTTATCAGTTCAAGCACCCTGACAGGGCATAAGAGAATCCACACACGGGAGAAGCCTTATAAATGCGAGGAGTGTGGCAAGGCCTTCAGCCAGTCCTCTACTCTCACAAGGCATAAGAGAATGCACACCGGGGAGAAGCCATACAAATGTGAAGAGTGCGGCAAGGCCTTCAAGGAAAGTTCAGCACTGACTAAGCATAAAATTATCCACACAGGTGAAAAACCTTATAAATGTGAAAAGTGCGGAAAAGCCTTCAACCAGAGCAGCATCCTGACAAATCATAAGAAAATTCACACTAGGGAGAAGCCTTACACCTGTAAAGAATGCGGCAAGAGCTTCAATAGGAGCAGCACCTTCACCAAGCATAAAGTGATCCACACCGGGGTCAAGC

Page 18: Plasmids - static-content.springer.com10.1038...pCAG ZNF91 [1-11]. pCAG ZNF91 was digested with XmaI and NotI and religated resulting in the removal of the C-terminal 12-36 domains

TGTATAAGTGCGAAGAATGCGGCAAGAGCTTCTTCTGGAGCAGCGCCCTCATCCGGCACAAGAAAATTCATACTGGGCAGCAGCCTTACAAGCAGGAGAAATTCGGCAAGACCTTCAACCAGTTTTCACACCTGACAACCGACAAGATTACACGGATTGGGGAGAAGTCCTACCCATACGATGTTCCAGATTACGCTTAGGATATC    >129L1PA4  

CTCCTCTGAGACAAAACTTCCAGAGGAACGATCAGGCAGCAGCATTTGCGGTTCACCAATATCCGCTGTTCTGCAGCCACCGCTGCTGATACCCAGGCAAACAGGGTCTGGAGTGGACCTCCAGCAAAC  >129L1PA4  scr  

CCGAGACACCCCCCCGATCATGCACAGCTTCGACGGTTTTGGACTCTTATGCGCGGTCGGCGTAATCCTCCTACGAAGATAAGTACTTCGAAGGGGATATACCAACCCGGAGATGACACCCGAGAAACC    >51L1PA4  ATATCCGCTGTTCTGCAGCCACCGCTGCTGATACCCAGGCAAACAGGGTCT    >ZNF93  HCGO  GAATTCATGGGACCCCTCCAGTTCCGCGATGTGGCTATTGAGTTCAGCCTCGAAGAATGGCATTGTCTCGATACCGCACAGCGAAACCTCTACCGGAATGTGATGCTGGAGAACTACTCAAATCTGGTCTTCCTCGGAATCGTGGTCAGCAAGCCCGACCTGATTGCTCACCTGGAGCAGGGGAAGAAACCTCTGACAATGAAGCGCCACGAAATGGTGGCTAACCCAAGCGTCATCTGCTCCCACTTCGCACAGGACCTGTGGCCAGAGCAGAATATCAAGGATAGCTTTCAGAAAGTGATTCTCAGGAGATACGAAAAGAGGGGGCACGGTAACCTCCAGCTCATCAAGAGATGCGAGTCCGTGGACGAATGTAAAGTCCACACCGGCGGATACAACGGCCTGAATCAGTGCTCTACCACAACTCAGAGTAAGGTGTTCCAGTGTGATAAGTATGGAAAAGTCTTCCACAAGTTTAGTAACTCAAATAGACACAACATCCGGCATACAGAGAAGAAACCCTTTAAGTGCATTGAATGTGGGAAAGCCTTCAATCAGTTTAGCACACTGATCACTCACAAGAAAATTCATACTGGCGAGAAGCCTTACATCTGCGAGGAATGTGGAAAGGCATTCAAATATAGCTCCGCCCTCAATACCCACAAGAGAATCCATACAGGAGAGAAGCCATACAAATGCGACAAGTGTGATAAAGCCTTTATTGCTTCTAGTACTCTGTCCAAGCACGAAATCATTCATACCGGGAAGAAACCCTATAAGTGCGAGGAATGTGGTAAAGCCTTCAACCAGTCAAGCACACTCACTAAGCACAAGAAAATCCATACCGGCGAGAAGCCTTACAAATGTGAAGAATGTGGAAAGGCTTTTAATCAGTCCTCTACCCTGACAAAGCACAAGAAAATTCATACAGGGGAGAAACCATACGTGTGCGAGGAATGTGGCAAGGCCTTCAAGTATTCTCGCACACTCACCACACACAAGCGAATCCATACTGGCGAGAAGCCCTATAAATGCAACAAGTGTGGCAAGGCCTTCATCGCCAGTTCAACACTGAGCCGACACGAAATCATTCACATGGGGAAGAAACACTACAAATGTGAGGAGTGCGGCAAGGCCTTCATCTGGAGCAGCGTGCTCACTCGACACAAGAGGGTGCACACAGGTGAGAAGCCTTACAAATGCGAAGAGTGTGGAAAGGCTTTTAAATATTCTAGTGTGCTGTCAAGCCACAAGCGCAGTCATACCGGGGAGAAGCCATATAAATGTGAAGAGTGCGGCAAGGCCTTCGTGGCATCCTCTACCCTGAGCAAACACGAAATCATTCATACAGGCAAGAAACCCTACAAATGTGAGGAATGCGGCAAGGCCTTCAACCAGAGTTCAAGCCTGACCAAACACAAGAAAATCCATACTGGGGAGAAGCCTTATAAATGTGAAGAGTGTGGCAAGGCCTTCAACCAGTCCTCTAGTCTGACAAAGCATAAGAAAATTCATACAGGCGAAAAACCATACAAATGTGAGGAGTGTGGAAAGGCTTTTAACCAGTCAAGCACTCTGACCAAGCATAAGAAAATCCACACTCGGGAGAAACCTTATAAATGTGAAGAATGTGGCAAGGCCTTCCACCTGTCTACTCATCTCACTACCCACAAGATTCTGCATACCGGGGAGAAACCTTACCGGTGCCGCGAATGTGGCAAGGCCTTCAACCACTCCGCTACCCTCTCCTCTCACAAGAAAATCCATTCTGGCGAGAAGTACCCATACGATGTTCCAGATTACGCTTGAGCGGCCGC  

Page 19: Plasmids - static-content.springer.com10.1038...pCAG ZNF91 [1-11]. pCAG ZNF91 was digested with XmaI and NotI and religated resulting in the removal of the C-terminal 12-36 domains

 >ZNF93  SerF  GAATTCATGGGTCCTCTCCAGTTTAGGGATGTGGCTATTGAGTTTAGTCTCGAAGAATGGCATTGTCTCGATACAGCTCAGCGAAATCTCTATCGGAATGTGATGCTGGAGAACTACTCCAATCTGGTCTTCCTCGGCATCGTGGTCTCTAAGCCCGACCTGATTGCTCACCTGGAGCAGGGAAAGAAACCTCTGACCATGAAGCGCCACGAAATGGTGGCCAACCCATCCGTCATCTGCTCTCACTTCGCTCAGGACCTGTGGCCTGAGCAGAACATCAAGGATTCTTTCCAGAAAGTGATTCTCAGGAGATACGAAAAGCGCGGGCACGGTAACCTCCAGCTCATCAAGCGATGCGAGAGCGTGGACGAATGTAAAGTCCACACCGGCGGATACAACGGGCTGAATCAGTGCAGTACCACAACTCAGTCAAAGGTGTTCCAGTGTGATAAGTATGGAAAAGTCTTCCACAAGTTTTCAAACAGCAATCGACACAACATCAGGCATACCGAGAAGAAACCCTTCAAGTGCATTGAATGTGGCAAAGCCTTCAATCAGTTTAGCACTCTGATCACCCACAAGAAAATTCATACAGGCGAGAAGCCCTACATCTGCGAGGAATGTGGAAAGGCTTTCAAATATAGCTCCGCACTCAATACACACAAGAGAATCCATACTGGCGAGAAGCCTTACAAATGCGACAAGTGTGATAAAGCCTTTATTGCTTCTAGTACCCTGTCCAAGCACGAAATCATTCATACAGGGAAGAAACCATATAAGTGCGAGGAATGTGGTAAAGCATTCAACCAGTCAAGCACACTCACTAAGCACAAGAAAATCCACACAGGTGAAAAGCCCTACAAATGTGAAGAATGTGGAAAGGCCTTTAATCAGTCCTCTACCCTGACAAAGCACAAGAAAATTCATACCGGGGAGAAACCTTACGTGTGCGAGGAATGTGGCAAGGCCTTCAAGTATAGCAGAATCCTCACCACACACAAGCGGATTCATACCGGTGAGAAGCCATATAAATGCAACAAGTGTGGAAAAGCATTCATCAGTTCAAGCTCCCTGTCTAGTCACGAGTTCATCCACATGGGGAAGAAACACTACAAATGTGAGGAATGTGGTAAAGCCTTTATCTCAAGCTCCTCTCTCACCAGCCACAAGAGGGTGCACACCGGCGAGAAGCCCTATAAATGTGAGGAGTGCGGAAAGGCTTTCAAAAGTTCAAGCTCCCTGTCTAGTCATAAGAGATCCCATACTGGGGAAAAACCTTACAAATGCGAAGAGTGTGGCAAGGCCTTCGTGTCAAGCTCCTCTCTGAGTTCACATGAGATCATTCATACAGGCAAGAAACCTTACAAGTGCGAGGAATGTGGAAAAGCCTTCAACAGCTCCTCTAGTCTGACTTCCCACAAGAAAATCCATACCGGGGAGAAGCCCTACAAATGCGAAGAATGCGGCAAGGCCTTCAACAGCAGCAGCAGCCTGACATCTCACAAGAAAATTCACACAGGCGAGAAGCCTTATAAATGTGAGGAGTGTGGAAAGGCATTCAACCAGAGTTCAACTCTGATCAAACATAAGAAAATTCACACCCGAGAGAAGCCATACAAATGCGAAGAATGTGGCAAGGCCTTTCACCTGTCCACACATCTCACTACCCACAAGATCCTGCATACTGGGGAGAAACCCTATCGGTGCCGCGAGTGCGGCAAGGCCTTCAACCACAGCGCCACCCTCAGCTCCCATAAGAAAATCCACTCCGGAGAGAAGCCCTACGAATGCGATAAGTGTGGCAAGGCCTTCATCTCTCCTTCTAGTCTGAGTCGCCACGAGATTATCCATACAGGCGAAAAGCCATACCCATACGATGTTCCAGATTACGCTTGAGCGGCCGC    >ZNF93  rhesus  macaque  GAATTCATGGGACCTCTCCAGTTCCGGGACGTGGCTATTGAGTTTAGTCTCGAAGAGTGGCATTGTCTCGATACCGCACAGCGCAACCTCTACAGGAACGTGATGCTGGAGAACTACTCAAATCTGGTCTTCCTCGGCATCGTGGTCAGCAAGCCCGACCTGATTGCCCACCTGGAGCAGGGAAAGAAACCTCTGACAATGAAGAGACACGAAATGGTGGCTAACCCATCTGTCATGTGCAGTCACTTCGCACAGGACCTCTGGCCCGAGCAGAATATGAAGGATAGCTTTCAGAAAATGATCCCTAGGAGATACGAGAAGCGGGGCCACGAAAACGTGCAGCTGATTAAGTGGCGCGAGAGCATGGACGAATGCAAGGTCCACACCGGCGGATATAACGGGCCAAATCAGTGCTGTACCACAACTCAGTCTAAGATGTTCCAGTGTGATAAGTACGGCAAAGTGTTCCACAAGTTTAGCAACTCCAATAGACACAAAATCCGGCATACCGAGAAGAAACCCTTCAAGTGCATTGAATGTGGAAAAGCATTCAATCTGTTTAGCACACTCACTCGACACAAGAGAATCCACACAGGGGAGAAACCCTACATTTGCGAGGAATGTGGCAAGGCTTTCAAGTATAGCTCCACACTGACCACCCATAAGAGAATCCACACTGGGGAGAAGCCTTATA

Page 20: Plasmids - static-content.springer.com10.1038...pCAG ZNF91 [1-11]. pCAG ZNF91 was digested with XmaI and NotI and religated resulting in the removal of the C-terminal 12-36 domains

AATGCGACAAGTGTGATAAAGCCTTTATTGCTTCTAGTACCCTCTCCAAGCACGAAATCATTCATACAGGCAAGAAACCATACAAGTGTGAAGAGTGCGGCAAGGCCTTCAACAGGTCAAGCATCCTGACTACCCACAAGAGAATCCACACCGGGGAGAAGCCCTATAAATGCGATAAGTGTGGCAAGGCCTTCATCACTTCCTCTACCCTCTCTAAACATGAGATTATCCACACAGGCAAGAAACCTTACAAGTGCGAGGAATGTGGAAAAGCCTTCTTTCAGAGTTCAATCCTGACTCGGCACAAGATCATTCATAGCGGGGAGAAGCCATACAAATGTGAAGAGTGTGGAAAAGCCTTTAAATATTCTTCCGTGCTCGCCACCCACAAGAAAAGTCATACAGAGGAAAAGAGGTACAAATGTGAAGAATGTGGCAAGACCTTCAAGCAGTCTAGTTATCTGTCAACTCACAAGATCATTCATACCGGCGAGAAGCCCTACAAATGCGAGGAATGTGGAAAGGCTTTCAACCAGTCAAGCACCCTCACAAAGCACAAGAAAATCCATACCGGGGAGAAGCCTTACGTGTGCGAAGAGTGTGGCAAGGCCTTCAAGTACTCTTGTACTCTGACAACTCACAAGAGAATCCATACCGGCGAGAAGCCATACAAGTGCAATAAGTGTGGAAAAGCATTCATTGCCTCCTCTACACTCAGTAAGCACGAAATCATTCACATGGGGAAGAAACACTATAAATGTGAGGAGTGTGGCAAGGCCTTCGTGTGGAGCAGCGTGCTGACTCGACATAAAATTATCCACACAGGTGAAAAACCTTATAAGTGTGAAGAATGTGGAAAGGCCTTCAAATATAGCTCCGTGCTGTCTAGTCACAAGCGCAGCCACACAGGCGAGAAGCCTTACAAATGTGAAGAATGCGGCAAGGCGTTCAAGTACAGCAGCGCCCTGAGCACCCACAAGATCATTCATACTGGCGAGAAGCCATATAAGTGTGAGGAGTGTGGACGCGTGTTCAAGCGATCCTCTGTCCTGTCCAAACACAAGAAAATCCATACAGCCGAGAAGACTTATAAGTGTGAGGAATGCGGAAAGGCTTTTAACCAGAGTTCAACTCTGACCACCCATAAGAAAATCCACACAGGGAAGAAACCCTATACATGTGAGGAATGCGGCAAGGCCTTCCACCTGTCTGCACATCTCACTACCCACAAGATCCTGCATACCGGAGAGAAACCTTACCGGTGCCGCGAATGTGGGGAGGCATTCAATAGATCAGCTACACTGTTCTCCCATAAAAAGATTCATTCAGGCGAAAATTACCCATACGATGTTCCAGATTACGCTTGAGCGGCCGC    >Consensus  monkey  ZNF93  (not  in  manuscript)  GAATTCATGGGTCCCCTCCAGTTCAGAGATGTGGCTATTGAGTTTTCCCTCGAAGAATGGCACTGCCTCGATACCGCTCAGAGAAACCTGTACAGAAACGTGATGCTGGAGAACTACAGCAATCTGGTCTTCCTCGGAATCGTGGTCTCCAAGCCCGACCTGATTGCCCACCTGGAGCAGGGGAAGAAACCTCTGACAATGAAGCGGCACGAAATGGTGGCCAACCCATCAGTCATGTGCAGCCACTTCGCTCAGGACCTCTGGCCCGAGCAGAATATGAAGGATAGCTTTCAGAAAATGATCCCTAGGAGATACGAAAAGCGGGGGCACGGTAACGTGCAGCTGATTAAGTGGCGCGAGTCTATGGACGAATGCAAGGTCCACACCGGCGGATATAACGGCCCCAATCAGTGCTGTACCACAACTCAGAGTAAGATGTTCCAGTGTGATAAGTACGGAAAAGTGTTCCACAAGTTTTCTAACAGTAACAGACACAAGATCCGGCATACTGAGAAGAAACCTTTTAAGTGCATTGAATGTGGCAAGGCCTTCAACCTGTTCAGCACCCTGACTCGACACAAGAGAATCCACACCGGCGAGAAACCATACATTTGCGAGGAATGTGGAAAGGCATTCAAATATAGCTCCACACTGACCACCCATAAGAGAATCCACACTGGAGAGAAGCCCTATAAATGCGACAAGTGTGATAAAGCCTTTATTGCTTCTAGTACCCTCTCCAAGCACGAAATCATTCATACAGGGAAGAAACCTTACAAGTGCGAGGAATGTGGTAAAGCCTTCAATAGGTCAAGCATCCTGACTACCCACAAAAGAATCCACACAGGCGAGAAGCCATATAAATGCGATAAGTGTGGCAAGGCCTTCATCACTTCCTCTACCCTCTCTAAACATGAAATCATTCATACCGGGAAGAAACCCTATAAATGTGAAGAATGTGGTAAAGCATTCTTTCAGAGTTCAATCCTGACACGCCACAAGATCATTCATAGCGGCGAGAAGCCTTACAAATGTGAAGAGTGTGGCAAGGCTTTCAAATATAGCTCCGTGCTCGCAACCCACAAGAAATCACATACAGAGGAAAAGCGATACAAATGCGAGGAATGTGGCAAGACCTTCAAGCAGTCTAGTTATCTGTCCACTCACAAGATCATTCATACCGGGGAGAAGCCTTATAAGTGTGAGGAGTGCGGCAAGGCCTTCAACCAGTCAAGCACCCTCACAAAGCACAAGAAAATCCATACTGGCGAAAAGCCCTACGTGTGCGAAGAGTGTGGAAAGGCCTTTAAATATAGCTGTACTCTGACAACTCATAAAAGAATCCACACCGGGGAGAAGCCTTACAAGTGCAATAAGTGTGGTAAAGCA

Page 21: Plasmids - static-content.springer.com10.1038...pCAG ZNF91 [1-11]. pCAG ZNF91 was digested with XmaI and NotI and religated resulting in the removal of the C-terminal 12-36 domains

TTCATTGCCTCCTCTACACTGTCTAAACACGAAATCATTCACATGGGCAAGAAACACTATAAGTGCGAGGAATGCGGCAAGGCCTTCGTGTGGAGCAGCGTGCTGACTCGACATAAAATTATCCATACCGGGGAGAAGCCCTACAAGTGTGAAGAATGCGGCAAGGCCTTCAAGTATAGCTCCGTGCTGTCTAGTCACAAGAGGTCTCATACCGGCGAGAAGCCCTACAAGTGCGAAGAGTGTGGCAAGGCCTTCAAGTATTCAAGCGCCCTGAGTACACACAAGATCATTCATACTGGGGAGAAGCCTTATAAATGCGAGGAATGTGGTCGCGTGTTCAAGCGATCCTCTGTCCTGTCCAAACACAAGAAAATCCATACAGCAGAGAAGACTTACAAATGTGAGGAGTGTGGCAAGGCCTTTAACCAGAGTTCAACCCTGACCACCCATAAGAAAATCCATACCGGCAAGAAACCATATACATGTGAGGAGTGTGGAAAGGCTTTCCACCTGTCCGCACATCTCACTACCCACAAGATCCTGCATACAGGGGAGAAACCCTACCGGTGCCGCGAGTGTGGTGAAGCCTTCAACAGATCCGCTACTCTCTTTTCTCACAAGAAAATCCACTCTGGGGAGAATAAGCCATACAAATGCGACAAGTGTGGCAAAATCTTCATTAGTCCCAGCTCCTTTTCAAGACACGAGATTATTCATACCGGAGAAAAGTCATACCCCTACGATGTGCCAGATTACGCATACCCATACGATGTTCCAGATTACGCTTGAGCGGCCGC