LOCUS       sequence-352526-       12797 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  nuclease Cas expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 12797)
  AUTHORS   Sangree AK, Griffith AL, Szegletes ZM, Roy P, DeWeirdt PC, Hegde M, 
            McGee AV, Hanna RE, Doench JG
  TITLE     Benchmarking of SpCas9 variants enables deeper base editor screens 
            of BRCA1 and BCL2.
  JOURNAL   Nat Commun. 2022 Mar 14;13(1):1318. doi: 10.1038/s41467-022-28884-7.
  PUBMED    35288574
REFERENCE   2  (bases 1 to 12797)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2022-03-14"; volume: "13(1):1318. doi"; pages: " 
            10.1038/s41467-022-28884-7"
COMMENT     Sequence Label: pRDA_156
FEATURES             Location/Qualifiers
     source          1..12797
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    84..201
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        340..1518
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          571..1509
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1466..1486
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             1535..1555
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(1562..4960,4961..4963,4964..5209,5210..5212,5213..5560,
                     5561..5563,5564..5566,5567..5569,5570..5662)
                     /codon_start=1
                     /product="Streptococcus pyogenes Cas9 endonuclease with the
                     D1135V/G1218R/R1335E/T1337R mutations"
                     /label=Cas9 VRER
                     /note="has an altered recognition specificity for the 
                     protospacer adjacent motif (PAM) (Kleinstiver et al., 
                     2015)"
                     /translation="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFVSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASARELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKEYRSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             1562..5662
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFVSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASARELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKEYRSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             5669..5716
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             5729..5782
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             5789..6184
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     CDS             6185..6241
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             6248..6946
                     /codon_start=1
                     /product="monomeric far-red fluorescent protein (Shcherbo 
                     et al., 2009)"
                     /label=mKate2
                     /note="mammalian codon-optimized"
                     /translation="MVSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMRIK
                     AVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEASTETLYPADGGLEGRADMAL
                     KLVGGGHLICNLKTTYRSKKPAKNLKMPGVYYVDRRLERIKEADKETYVEQHEVAVARY
                     CDLPSKLGHR"
     misc_feature    6965..7553
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(7018..7038)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(7436..7447)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             7625..7858
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    7936..8057
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7973..7992)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8027..8046
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      8097..8232
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(8252..8271)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(8253..8271)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(8281..8298)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(8281..8297)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8290..8312)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      8439..8894
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(8526..8545)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     8736..8757
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        8920..9024
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(9025..9093,9094..9885)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(9243..9262)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      10056..10644
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     10545..10564
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     10798..10815
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    10932..10953
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(10968..10973,10974..10991,10992..10998)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11006..11022
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     11011..11033
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     11030..11046
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     11030..11046
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     11065..11085
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        11067..11085
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        11113..11339
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             11340..11520
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    11567..11692
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    12185..12418
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             12603..12647
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
ORIGIN
        1 cacctcccaa ccccgagggg accctggtac ccatgcatgc atgaattcta gatcttgaga
       61 caaatggcag tattcatcca caattttaaa agaaaagggg ggattggggg gtacagtgca
      121 ggggaaagaa tagtagacat aatagcaaca gacatacaaa ctaaagaatt acaaaaacaa
      181 attacaaaaa ttcaaaattt tcgggtttat tacagggaca gcagagatcc actttggcgc
      241 cggctcgagg gggcccgggt gcaaagatgg ataaagtttt aaacagagag gaatctttgc
      301 agctaatgga ccttctaggt cttgaaagga gtgggaattg gctccggtgc ccgtcagtgg
      361 gcagagcgca catcgcccac agtccccgag aagttggggg gaggggtcgg caattgaacc
      421 ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg atgtcgtgta ctggctccgc
      481 ctttttcccg agggtggggg agaaccgtat ataagtgcag tagtcgccgt gaacgttctt
      541 tttcgcaacg ggtttgccgc cagaacacag gtaagtgccg tgtgtggttc ccgcgggcct
      601 ggcctcttta cgggttatgg cccttgcgtg ccttgaatta cttccacctg gctgcagtac
      661 gtgattcttg atcccgagct tcgggttgga agtgggtggg agagttcgag gccttgcgct
      721 taaggagccc cttcgcctcg tgcttgagtt gaggcctggc ctgggcgctg gggccgccgc
      781 gtgcgaatct ggtggcacct tcgcgcctgt ctcgctgctt tcgataagtc tctagccatt
      841 taaaattttt gatgacctgc tgcgacgctt tttttctggc aagatagtct tgtaaatgcg
      901 ggccaagatc tgcacactgg tatttcggtt tttggggccg cgggcggcga cggggcccgt
      961 gcgtcccagc gcacatgttc ggcgaggcgg ggcctgcgag cgcggccacc gagaatcgga
     1021 cgggggtagt ctcaagctgg ccggcctgct ctggtgcctg gcctcgcgcc gccgtgtatc
     1081 gccccgccct gggcggcaag gctggcccgg tcggcaccag ttgcgtgagc ggaaagatgg
     1141 ccgcttcccg gccctgctgc agggagctca aaatggagga cgcggcgctc gggagagcgg
     1201 gcgggtgagt cacccacaca aaggaaaagg gcctttccgt cctcagccgt cgcttcatgt
     1261 gactccacgg agtaccgggc gccgtccagg cacctcgatt agttctcgag cttttggagt
     1321 acgtcgtctt taggttgggg ggaggggttt tatgcgatgg agtttcccca cactgagtgg
     1381 gtggagactg aagttaggcc agcttggcac ttgatgtaat tctccttgga atttgccctt
     1441 tttgagtttg gatcttggtt cattctcaag cctcagacag tggttcaaag tttttttctt
     1501 ccatttcagg tgtcgtgagg gatccaccat ggcaccaaag aagaagcgga aggtcgcagc
     1561 cgacaagaag tacagcatcg gcctggacat cggcaccaac tctgtgggct gggccgtgat
     1621 caccgacgag tacaaggtgc ccagcaagaa attcaaggtg ctgggcaaca ccgaccggca
     1681 cagcatcaag aagaacctga tcggagccct gctgttcgac agcggcgaaa cagccgaggc
     1741 cacccggctg aagagaaccg ccagaagaag atacaccaga cggaagaacc ggatctgcta
     1801 tctgcaagag atcttcagca acgagatggc caaggtggac gacagcttct tccacagact
     1861 ggaagagtcc ttcctggtgg aagaggataa gaagcacgag cggcacccca tcttcggcaa
     1921 catcgtggac gaggtggcct accacgagaa gtaccccacc atctaccacc tgagaaagaa
     1981 actggtggac agcaccgaca aggccgacct gcggctgatc tatctggccc tggcccacat
     2041 gatcaagttc cggggccact tcctgatcga gggcgacctg aaccccgaca acagcgacgt
     2101 ggacaagctg ttcatccagc tggtgcagac ctacaaccag ctgttcgagg aaaaccccat
     2161 caacgccagc ggcgtagacg ccaaggccat cctgtctgcc agactgagca agagcagacg
     2221 gctggaaaat ctgatcgccc agctgcccgg cgagaagaag aatggcctgt tcggaaacct
     2281 gattgccctg agcctgggcc tgacccccaa cttcaagagc aacttcgacc tggccgagga
     2341 tgccaaactg cagctgagca aggacaccta cgacgacgac ctggacaacc tgctggccca
     2401 gatcggcgac cagtacgccg acctgtttct ggccgccaag aacctgtccg acgccatcct
     2461 gctgagcgac atcctgagag tgaacaccga gatcaccaag gcccccctga gcgcctctat
     2521 gatcaagaga tacgacgagc accaccagga cctgaccctg ctgaaagctc tcgtgcggca
     2581 gcagctgcct gagaagtaca aagagatttt cttcgaccag agcaagaacg gctacgccgg
     2641 ctacattgac ggcggagcca gccaggaaga gttctacaag ttcatcaagc ccatcctgga
     2701 aaagatggac ggcaccgagg aactgctcgt gaagctgaac agagaggacc tgctgcggaa
     2761 gcagcgtacg ttcgacaacg gcagcatccc ccaccagatc cacctgggag agctgcacgc
     2821 cattctgcgg cggcaggaag atttttaccc attcctgaag gacaaccggg aaaagatcga
     2881 gaagatcctg accttccgca tcccctacta cgtgggccct ctggccaggg gaaacagcag
     2941 attcgcctgg atgaccagaa agagcgagga aaccatcacc ccctggaact tcgaggaagt
     3001 ggtggacaag ggcgctagcg cccagagctt catcgagcgg atgaccaact tcgataagaa
     3061 cctgcccaac gagaaggtgc tgcccaagca cagcctgctg tacgagtact tcaccgtgta
     3121 taacgagctg accaaagtga aatacgtgac cgagggaatg agaaagcccg ccttcctgag
     3181 cggcgagcag aaaaaggcca tcgtggacct gctgttcaag accaaccgga aagtgaccgt
     3241 gaagcagctg aaagaggact acttcaagaa aatcgagtgc ttcgactccg tggaaatctc
     3301 cggcgtggaa gatcggttca acgcctccct gggcacatac cacgatctgc tgaaaattat
     3361 caaggacaag gacttcctgg acaatgagga aaacgaggac attctggaag atatcgtgct
     3421 gaccctgaca ctgtttgagg acagagagat gatcgaggaa cggctgaaaa cctatgccca
     3481 cctgttcgac gacaaagtga tgaagcagct gaagcggcgg agatacaccg gctggggcag
     3541 gctgagccgg aagctgatca acggcatccg ggacaagcag tccggcaaga caatcctgga
     3601 tttcctgaag tccgacggct tcgccaacag aaacttcatg cagctgatcc acgacgacag
     3661 cctgaccttt aaagaggaca tccagaaagc ccaggtgtcc ggccagggcg atagcctgca
     3721 cgagcacatt gccaatctgg ccggcagccc cgccattaag aagggcatcc tgcagacagt
     3781 gaaggtggtg gacgagctcg tgaaagtgat gggccggcac aagcccgaga acatcgtgat
     3841 cgaaatggcc agagagaacc agaccaccca gaagggacag aagaacagcc gcgagagaat
     3901 gaagcggatc gaagagggca tcaaagagct gggcagccag atcctgaaag aacaccccgt
     3961 ggaaaacacc cagctgcaga acgagaagct gtacctgtac tacctgcaga atgggcggga
     4021 tatgtacgtg gaccaggaac tggacatcaa ccggctgtcc gactacgatg tggaccatat
     4081 cgtgcctcag agctttctga aggacgactc catcgacaac aaggtgctga ccagaagcga
     4141 caagaaccgg ggcaagagcg acaacgtgcc ctccgaagag gtcgtgaaga agatgaagaa
     4201 ctactggcgg cagctgctga acgccaagct gattacccag agaaagttcg acaatctgac
     4261 caaggccgag agaggcggcc tgagcgaact ggataaggcc ggcttcatca agagacagct
     4321 ggtggaaacc cggcagatca caaagcacgt ggcacagatc ctggactccc ggatgaacac
     4381 taagtacgac gagaatgaca agctgatccg ggaagtgaaa gtgatcaccc tgaagtccaa
     4441 gctggtgtcc gatttccgga aggatttcca gttttacaaa gtgcgcgaga tcaacaacta
     4501 ccaccacgcc cacgacgcct acctgaacgc cgtcgtggga accgccctga tcaaaaagta
     4561 ccctaagctg gaaagcgagt tcgtgtacgg cgactacaag gtgtacgacg tgcggaagat
     4621 gatcgccaag agcgagcagg aaatcggcaa ggctaccgcc aagtacttct tctacagcaa
     4681 catcatgaac tttttcaaga ccgagattac cctggccaac ggcgagatcc ggaagcggcc
     4741 tctgatcgag acaaacggcg aaaccgggga gatcgtgtgg gataagggcc gggattttgc
     4801 caccgtgcgg aaagtgctga gcatgcccca agtgaatatc gtgaaaaaga ccgaggtgca
     4861 gacaggcggc ttcagcaaag agtctatcct gcccaagagg aacagcgata agctgatcgc
     4921 cagaaagaag gactgggacc ctaagaagta cggcggcttc gtgagcccca ccgtggccta
     4981 ttctgtgctg gtggtggcca aagtggaaaa gggcaagtcc aagaaactga agagtgtgaa
     5041 agagctgctg gggatcacca tcatggaaag aagcagcttc gagaagaatc ccatcgactt
     5101 tctggaagcc aagggctaca aagaagtgaa aaaggacctg atcatcaagc tgcctaagta
     5161 ctccctgttc gagctggaaa acggccggaa gagaatgctg gcctctgcca gagaactgca
     5221 gaagggaaac gaactggccc tgccctccaa atatgtgaac ttcctgtacc tggccagcca
     5281 ctatgagaag ctgaagggct cccccgagga taatgagcag aaacagctgt ttgtggaaca
     5341 gcacaagcac tacctggacg agatcatcga gcagatcagc gagttctcca agagagtgat
     5401 cctggccgac gctaatctgg acaaagtgct gtccgcctac aacaagcacc gggataagcc
     5461 catcagagag caggccgaga atatcatcca cctgtttacc ctgaccaatc tgggagcccc
     5521 tgccgccttc aagtactttg acaccaccat cgaccggaag gagtacagga gcaccaaaga
     5581 ggtgctggac gccaccctga tccaccagag catcaccggc ctgtacgaga cacgaatcga
     5641 tttgtctcag ctgggaggcg acgccgcaaa aaggccggcg gccacgaaaa aggccggcca
     5701 ggcaaaaaag aaaaaggcag gctcaggcga gggcagagga agtctgctaa catgcggtga
     5761 cgtcgaggag aatcctggcc caagcgctat ggccaagcct ttgtctcaag aagaatccac
     5821 cctcattgaa agagcaacgg ctacaatcaa cagcatcccc atctctgaag actacagcgt
     5881 cgccagcgca gctctctcta gcgacggccg catcttcact ggtgtcaatg tatatcattt
     5941 tactggggga ccttgtgcag aactcgtggt gctgggcact gctgctgctg cggcagctgg
     6001 caacctgact tgtatcgtcg cgatcggaaa tgagaacagg ggcatcttga gcccctgcgg
     6061 acggtgccga caggtgcttc tcgatctgca tcctgggatc aaagccatag tgaaggacag
     6121 tgatggacag ccgacggcag ttgggattcg tgaattgctg ccctctggtt atgtgtggga
     6181 gggcgcaaca aacttctctc tgctgaaaca agccggagat gtcgaagaga atcctggacc
     6241 gactagtatg gtgagcgagc tgattaagga gaacatgcac atgaagctat acatggaggg
     6301 caccgtgaac aaccaccact tcaagtgcac atccgagggc gaaggcaagc cctacgaggg
     6361 cacccagacc atgagaatca aggcggtcga gggcggccct ctccccttcg ccttcgacat
     6421 cctggctacc agcttcatgt acggcagcaa aaccttcatc aaccacaccc agggcatccc
     6481 cgacttcttt aagcagtcct tccccgaggg cttcacatgg gagagagtca ccacatacga
     6541 agacgggggc gtgctgaccg ctacccagga caccagcctc caggacggct gcctcatcta
     6601 caacgtcaag atcagagggg tgaacttccc atccaacggc cctgtgatgc agaagaaaac
     6661 actcggctgg gaggcctcca ccgagaccct gtaccccgct gacggcggcc tggaaggcag
     6721 agccgacatg gccctgaagc tcgtgggcgg gggccacctg atctgcaact tgaagaccac
     6781 atacagatcc aagaaacccg ctaagaacct caagatgccc ggcgtctact atgtggacag
     6841 aagactggaa agaatcaagg aggccgacaa agagacctac gtcgagcagc acgaggtggc
     6901 tgtggccaga tactgcgacc tccctagcaa actggggcac agatgataga cgcgttaagt
     6961 cgacaatcaa cctctggatt acaaaatttg tgaaagattg actggtattc ttaactatgt
     7021 tgctcctttt acgctatgtg gatacgctgc tttaatgcct ttgtatcatg ctattgcttc
     7081 ccgtatggct ttcattttct cctccttgta taaatcctgg ttgctgtctc tttatgagga
     7141 gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc
     7201 cactggttgg ggcattgcca ccacctgtca gctcctttcc gggactttcg ctttccccct
     7261 ccctattgcc acggcggaac tcatcgccgc ctgccttgcc cgctgctgga caggggctcg
     7321 gctgttgggc actgacaatt ccgtggtgtt gtcggggaaa tcatcgtcct ttccttggct
     7381 gctcgcctgt gttgccacct ggattctgcg cgggacgtcc ttctgctacg tcccttcggc
     7441 cctcaatcca gcggaccttc cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg
     7501 tcttcgcctt cgccctcaga cgagtcggat ctccctttgg gccgcctccc cgcgtcgact
     7561 ttaagaccaa tgacttacaa ggcagctgta gatcttagcc actttttaaa agaaaagggg
     7621 ggactggaag ggctaattca ctcccaacga agacaagatc tgctttttgc ttgtactggg
     7681 tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg
     7741 cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt
     7801 gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagt
     7861 acgtatagta gttcatgtca tcttattatt cagtatttat aacttgcaaa gaaatgaata
     7921 tcagagagtg agaggaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     7981 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     8041 ctcatcaatg tatcttatca tgtctggctc tagctatccc gcccctaact ccgcccatcc
     8101 cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta
     8161 tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag tgaggaggct
     8221 tttttggagg cctagggacg tacccaattc gccctatagt gagtcgtatt acgcgcgctc
     8281 actggccgtc gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg
     8341 ccttgcagca catccccctt tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg
     8401 cccttcccaa cagttgcgca gcctgaatgg cgaatgggac gcgccctgta gcggcgcatt
     8461 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     8521 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     8581 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     8641 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     8701 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     8761 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc
     8821 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaaatatt
     8881 aacgcttaca atttaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     8941 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     9001 caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
     9061 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
     9121 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     9181 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     9241 ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc
     9301 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     9361 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     9421 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     9481 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     9541 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
     9601 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     9661 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     9721 tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag
     9781 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     9841 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     9901 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     9961 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
    10021 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
    10081 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
    10141 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
    10201 gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
    10261 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
    10321 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
    10381 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
    10441 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
    10501 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
    10561 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
    10621 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
    10681 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
    10741 cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc
    10801 gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt
    10861 tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag
    10921 cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt acactttatg
    10981 cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac aggaaacagc
    11041 tatgaccatg attacgccaa gcgcgcaatt aaccctcact aaagggaaca aaagctggag
    11101 ctgcaagctt aatgtagtct tatgcaatac tcttgtagtc ttgcaacatg gtaacgatga
    11161 gttagcaaca tgccttacaa ggagagaaaa agcaccgtgc atgccgattg gtggaagtaa
    11221 ggtggtacga tcgtgcctta ttaggaaggc aacagacggg tctgacatgg attggacgaa
    11281 ccactgaatt gccgcattgc agagatattg tatttaagtg cctagctcga tacataaacg
    11341 ggtctctctg gttagaccag atctgagcct gggagctctc tggctaacta gggaacccac
    11401 tgcttaagcc tcaataaagc ttgccttgag tgcttcaagt agtgtgtgcc cgtctgttgt
    11461 gtgactctgg taactagaga tccctcagac ccttttagtc agtgtggaaa atctctagca
    11521 gtggcgcccg aacagggact tgaaagcgaa agggaaacca gaggagctct ctcgacgcag
    11581 gactcggctt gctgaagcgc gcacggcaag aggcgagggg cggcgactgg tgagtacgcc
    11641 aaaaattttg actagcggag gctagaagga gagagatggg tgcgagagcg tcagtattaa
    11701 gcgggggaga attagatcgc gatgggaaaa aattcggtta aggccagggg gaaagaaaaa
    11761 atataaatta aaacatatag tatgggcaag cagggagcta gaacgattcg cagttaatcc
    11821 tggcctgtta gaaacatcag aaggctgtag acaaatactg ggacagctac aaccatccct
    11881 tcagacagga tcagaagaac ttagatcatt atataataca gtagcaaccc tctattgtgt
    11941 gcatcaaagg atagagataa aagacaccaa ggaagcttta gacaagatag aggaagagca
    12001 aaacaaaagt aagaccaccg cacagcaagc ggccgctgat cttcagacct ggaggaggag
    12061 atatgaggga caattggaga agtgaattat ataaatataa agtagtaaaa attgaaccat
    12121 taggagtagc acccaccaag gcaaagagaa gagtggtgca gagagaaaaa agagcagtgg
    12181 gaataggagc tttgttcctt gggttcttgg gagcagcagg aagcactatg ggcgcagcgt
    12241 caatgacgct gacggtacag gccagacaat tattgtctgg tatagtgcag cagcagaaca
    12301 atttgctgag ggctattgag gcgcaacagc atctgttgca actcacagtc tggggcatca
    12361 agcagctcca ggcaagaatc ctggctgtgg aaagatacct aaaggatcaa cagctcctgg
    12421 ggatttgggg ttgctctgga aaactcattt gcaccactgc tgtgccttgg aatgctagtt
    12481 ggagtaataa atctctggaa cagatttgga atcacacgac ctggatggag tgggacagag
    12541 aaattaacaa ttacacaagc ttaatacact ccttaattga agaatcgcaa aaccagcaag
    12601 aaaagaatga acaagaatta ttggaattag ataaatgggc aagtttgtgg aattggttta
    12661 acataacaaa ttggctgtgg tatataaaat tattcataat gatagtagga ggcttggtag
    12721 gtttaagaat agtttttgct gtactttcta tagtgaatag agttaggcag ggatattcac
    12781 cattatcgtt tcagacc
//
