LOCUS       sequence-178330-       12486 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral CRISPR-Cas9 delivery for SpCas9 and sgRNA (hU6), mNeon 
            coexpression, EFS Promoter driven.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 12486)
  AUTHORS   Reimer J, Knoess S, Labuhn M, Charpentier EM, Gohring G, 
            Schlegelberger B, Klusmann JH, Heckl D
  TITLE     CRISPR-Cas9-induced t(11;19)/MLL-ENL translocations initiate 
            leukemia in human hematopoietic progenitor cells in vivo.
  JOURNAL   Haematologica. 2017 Jun 1. pii: haematol.2017.164046. doi: 
            10.3324/haematol.2017.164046.
  PUBMED    28572162
REFERENCE   2  (bases 1 to 12486)
  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: 
            "Haematologica. 2017 Jun 1. pii: haematol.2017.164046. doi: 
            10.3324/haematol.2017.164046."
COMMENT     Sequence Label: L40C-CRISPR.EFS.mNeon
FEATURES             Location/Qualifiers
     source          1..12486
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     101..118
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    235..256
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(271..276,277..294,295..301)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    309..325
                     /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     314..336
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     333..349
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     333..349
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     368..388
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        370..388
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     enhancer        433..812
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     enhancer        869..1060
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(892..912)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        1079..1305
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             1306..1486
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1533..1658
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    2151..2384
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2569..2613
                     /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"
     misc_feature    2911..3028
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        3079..3327
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     3079..3099
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     3250..3269
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     promoter        3489..3700
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      3720..3729
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3729..3752
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             3759..3779
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3804..7904
                     /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
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             3804..7904
                     /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
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             7905..7952
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             7971..8027
                     /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"
     regulatory      8283..8292
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     misc_feature    8754..9342
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8807..8827)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(9225..9236)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             9414..9647
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    9725..9846
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(9762..9781)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     9816..9835
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      9886..10021
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(10041..10060)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(10042..10060)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(10070..10087)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(10070..10086)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(10079..10101)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      10228..10683
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(10315..10334)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     10525..10546
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        10709..10813
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(10814..10882,10883..11674)
                     /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(11032..11051)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      11845..12433
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     12334..12353
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga
       61 gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga
      121 agcggaagag cgcccaatac gcaaaccgcc tctccccgcg cgttggccga ttcattaatg
      181 cagctggcac gacaggtttc ccgactggaa agcgggcagt gagcgcaacg caattaatgt
      241 gagttagctc actcattagg caccccaggc tttacacttt atgcttccgg ctcgtatgtt
      301 gtgtggaatt gtgagcggat aacaatttca cacaggaaac agctatgacc atgattacgc
      361 caagcgcgca attaaccctc actaaaggga acaaaagctg gagctgcaag cggtaccgtc
      421 gacctgcagg ttgacattga ttattgacta gttattaata gtaatcaatt acggggtcat
      481 tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg
      541 gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa
      601 cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact
      661 tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta
      721 aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt
      781 acatctacgt attagtcatc gctattacca tggcgcgcca tgcagggcct gaaataacct
      841 ctgaaagagg aacttggtta ggtaccttct gaggcggaaa gaaccagctg tggaatgtgt
      901 gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
      961 atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta
     1021 tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc ttaagaggcg cgccttaatg
     1081 tagtcttatg caatactctt gtagtcttgc aacatggtaa cgatgagtta gcaacatgcc
     1141 ttacaaggag agaaaaagca ccgtgcatgc cgattggtgg aagtaaggtg gtacgatcgt
     1201 gccttattag gaaggcaaca gacgggtctg acatggattg gacgaaccac tgaattgccg
     1261 cattgcagag atattgtatt taagtgccta gctcgataca taaacgggtc tctctggtta
     1321 gaccagatct gagcctggga gctctctggc taactaggga acccactgct taagcctcaa
     1381 taaagcttgc cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac
     1441 tagagatccc tcagaccctt ttagtcagtg tggaaaatct ctagcagtgg cgcccgaaca
     1501 gggacttgaa agcgaaaggg aaaccagagg agctctctcg acgcaggact cggcttgctg
     1561 aagcgcgcac ggcaagaggc gaggggcggc gactggtgag tacgccaaaa attttgacta
     1621 gcggaggcta gaaggagaga gatgggtgcg agagcgtcag tattaagcgg gggagaatta
     1681 gatcgcgatg ggaaaaaatt cggttaaggc cagggggaaa gaaaaaatat aaattaaaac
     1741 atatagtatg ggcaagcagg gagctagaac gattcgcagt taatcctggc ctgttagaaa
     1801 catcagaagg ctgtagacaa atactgggac agctacaacc atcccttcag acaggatcag
     1861 aagaacttag atcattatat aatacagtag caaccctcta ttgtgtgcat caaaggatag
     1921 agataaaaga caccaaggaa gctttagaca agatagagga agagcaaaac aaaagtaaga
     1981 ccaccgcaca gcaagcggcc gctgatcttc agacctggag gaggagatat gagggacaat
     2041 tggagaagtg aattatataa atataaagta gtaaaaattg aaccattagg agtagcaccc
     2101 accaaggcaa agagaagagt ggtgcagaga gaaaaaagag cagtgggaat aggagctttg
     2161 ttccttgggt tcttgggagc agcaggaagc actatgggcg cagcgtcaat gacgctgacg
     2221 gtacaggcca gacaattatt gtctggtata gtgcagcagc agaacaattt gctgagggct
     2281 attgaggcgc aacagcatct gttgcaactc acagtctggg gcatcaagca gctccaggca
     2341 agaatcctgg ctgtggaaag atacctaaag gatcaacagc tcctggggat ttggggttgc
     2401 tctggaaaac tcatttgcac cactgctgtg ccttggaatg ctagttggag taataaatct
     2461 ctggaacaga tttggaatca cacgacctgg atggagtggg acagagaaat taacaattac
     2521 acaagcttaa tacactcctt aattgaagaa tcgcaaaacc agcaagaaaa gaatgaacaa
     2581 gaattattgg aattagataa atgggcaagt ttgtggaatt ggtttaacat aacaaattgg
     2641 ctgtggtata taaaattatt cataatgata gtaggaggct tggtaggttt aagaatagtt
     2701 tttgctgtac tttctatagt gaatagagtt aggcagggat attcaccatt atcgtttcag
     2761 acccacctcc caaccccgag gggacccgac aggcccgaag gaatagaaga agaaggtgga
     2821 gagagagaca gagacagatc cattcgatta gtgaacggat cggcactgcg tgcgccaatt
     2881 ctgcagacaa atggcagtat tcatccacaa ttttaaaaga aaagggggga ttggggggta
     2941 cagtgcaggg gaaagaatag tagacataat agcaacagac atacaaacta aagaattaca
     3001 aaaacaaatt acaaaaattc aaaattttcg ggtttattac agggacagca gagatccagt
     3061 ttggttaatt aaggtaccga gggcctattt cccatgattc cttcatattt gcatatacga
     3121 tacaaggctg ttagagagat aattagaatt aatttgactg taaacacaaa gatattagta
     3181 caaaatacgt gacgtagaaa gtaataattt cttgggtagt ttgcagtttt aaaattatgt
     3241 tttaaaatgg actatcatat gcttaccgta acttgaaagt atttcgattt cttggcttta
     3301 tatatcttgt ggaaaggacg aaacaccgga gacgtatgtc gacaactcgt ctctgtttaa
     3361 gagctatgct ggaaacagca tagcaagttt aaataaggct agtccgttat caacttgaaa
     3421 aagtggcacc gagtcggtgc ttttttgaat tctcctctag aactcgagtg gctccggtgc
     3481 ccgtcagtgg gcagagcgca catcgcccac agtccccgag aagttggggg gaggggtcgg
     3541 caattgaacc ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg atgtcgtgta
     3601 ctggctccgc ctttttcccg agggtggggg agaaccgtat ataagtgcag tagtcgccgt
     3661 gaacgttctt tttcgcaacg ggtttgccgc cagaacacag gtgtcgtgac gcgggatccg
     3721 ccaccatgga ttacaaagac gatgacgata agatggcccc aaagaagaag cggaaggtcg
     3781 gtatccacgg agtcccagca gccgacaaga agtacagcat cggcctggac atcggcacca
     3841 actctgtggg ctgggccgtg atcaccgacg agtacaaggt gcccagcaag aaattcaagg
     3901 tgctgggcaa caccgaccgg cacagcatca agaagaacct gatcggagcc ctgctgttcg
     3961 acagcggcga aacagccgag gccacccggc tgaagagaac cgccagaaga agatacacca
     4021 gacggaagaa ccggatctgc tatctgcaag agatcttcag caacgagatg gccaaggtgg
     4081 acgacagctt cttccacaga ctggaagagt ccttcctggt ggaagaggat aagaagcacg
     4141 agcggcaccc catcttcggc aacatcgtgg acgaggtggc ctaccacgag aagtacccca
     4201 ccatctacca cctgagaaag aaactggtgg acagcaccga caaggccgac ctgcggctga
     4261 tctatctggc cctggcccac atgatcaagt tccggggcca cttcctgatc gagggcgacc
     4321 tgaaccccga caacagcgac gtggacaagc tgttcatcca gctggtgcag acctacaacc
     4381 agctgttcga ggaaaacccc atcaacgcca gcggcgtgga cgccaaggcc atcctgtctg
     4441 ccagactgag caagagcaga cggctggaaa atctgatcgc ccagctgccc ggcgagaaga
     4501 agaatggcct gttcggaaac ctgattgccc tgagcctggg cctgaccccc aacttcaaga
     4561 gcaacttcga cctggccgag gatgccaaac tgcagctgag caaggacacc tacgacgacg
     4621 acctggacaa cctgctggcc cagatcggcg accagtacgc cgacctgttt ctggccgcca
     4681 agaacctgtc cgacgccatc ctgctgagcg acatcctgag agtgaacacc gagatcacca
     4741 aggcccccct gagcgcctct atgatcaaga gatacgacga gcaccaccag gacctgaccc
     4801 tgctgaaagc tctcgtgcgg cagcagctgc ctgagaagta caaagagatt ttcttcgacc
     4861 agagcaagaa cggctacgcc ggctacattg acggcggagc cagccaggaa gagttctaca
     4921 agttcatcaa gcccatcctg gaaaagatgg acggcaccga ggaactgctc gtgaagctga
     4981 acagagagga cctgctgcgg aagcagcgga ccttcgacaa cggcagcatc ccccaccaga
     5041 tccacctggg agagctgcac gccattctgc ggcggcagga agatttttac ccattcctga
     5101 aggacaaccg ggaaaagatc gagaagatcc tgaccttccg catcccctac tacgtgggcc
     5161 ctctggccag gggaaacagc agattcgcct ggatgaccag aaagagcgag gaaaccatca
     5221 ccccctggaa cttcgaggaa gtggtggaca agggcgcttc cgcccagagc ttcatcgagc
     5281 ggatgaccaa cttcgataag aacctgccca acgagaaggt gctgcccaag cacagcctgc
     5341 tgtacgagta cttcaccgtg tataacgagc tgaccaaagt gaaatacgtg accgagggaa
     5401 tgagaaagcc cgccttcctg agcggcgagc agaaaaaggc catcgtggac ctgctgttca
     5461 agaccaaccg gaaagtgacc gtgaagcagc tgaaagagga ctacttcaag aaaatcgagt
     5521 gcttcgactc cgtggaaatc tccggcgtgg aagatcggtt caacgcctcc ctgggcacat
     5581 accacgatct gctgaaaatt atcaaggaca aggacttcct ggacaatgag gaaaacgagg
     5641 acattctgga agatatcgtg ctgaccctga cactgtttga ggacagagag atgatcgagg
     5701 aacggctgaa aacctatgcc cacctgttcg acgacaaagt gatgaagcag ctgaagcggc
     5761 ggagatacac cggctggggc aggctgagcc ggaagctgat caacggcatc cgggacaagc
     5821 agtccggcaa gacaatcctg gatttcctga agtccgacgg cttcgccaac agaaacttca
     5881 tgcagctgat ccacgacgac agcctgacct ttaaagagga catccagaaa gcccaggtgt
     5941 ccggccaggg cgatagcctg cacgagcaca ttgccaatct ggccggcagc cccgccatta
     6001 agaagggcat cctgcagaca gtgaaggtgg tggacgagct cgtgaaagtg atgggccggc
     6061 acaagcccga gaacatcgtg atcgaaatgg ccagagagaa ccagaccacc cagaagggac
     6121 agaagaacag ccgcgagaga atgaagcgga tcgaagaggg catcaaagag ctgggcagcc
     6181 agatcctgaa agaacacccc gtggaaaaca cccagctgca gaacgagaag ctgtacctgt
     6241 actacctgca gaatgggcgg gatatgtacg tggaccagga actggacatc aaccggctgt
     6301 ccgactacga tgtggaccat atcgtgcctc agagctttct gaaggacgac tccatcgaca
     6361 acaaggtgct gaccagaagc gacaagaacc ggggcaagag cgacaacgtg ccctccgaag
     6421 aggtcgtgaa gaagatgaag aactactggc ggcagctgct gaacgccaag ctgattaccc
     6481 agagaaagtt cgacaatctg accaaggccg agagaggcgg cctgagcgaa ctggataagg
     6541 ccggcttcat caagagacag ctggtggaaa cccggcagat cacaaagcac gtggcacaga
     6601 tcctggactc ccggatgaac actaagtacg acgagaatga caagctgatc cgggaagtga
     6661 aagtgatcac cctgaagtcc aagctggtgt ccgatttccg gaaggatttc cagttttaca
     6721 aagtgcgcga gatcaacaac taccaccacg cccacgacgc ctacctgaac gccgtcgtgg
     6781 gaaccgccct gatcaaaaag taccctaagc tggaaagcga gttcgtgtac ggcgactaca
     6841 aggtgtacga cgtgcggaag atgatcgcca agagcgagca ggaaatcggc aaggctaccg
     6901 ccaagtactt cttctacagc aacatcatga actttttcaa gaccgagatt accctggcca
     6961 acggcgagat ccggaagcgg cctctgatcg agacaaacgg cgaaaccggg gagatcgtgt
     7021 gggataaggg ccgggatttt gccaccgtgc ggaaagtgct gagcatgccc caagtgaata
     7081 tcgtgaaaaa gaccgaggtg cagacaggcg gcttcagcaa agagtctatc ctgcccaaga
     7141 ggaacagcga taagctgatc gccagaaaga aggactggga ccctaagaag tacggcggct
     7201 tcgacagccc caccgtggcc tattctgtgc tggtggtggc caaagtggaa aagggcaagt
     7261 ccaagaaact gaagagtgtg aaagagctgc tggggatcac catcatggaa agaagcagct
     7321 tcgagaagaa tcccatcgac tttctggaag ccaagggcta caaagaagtg aaaaaggacc
     7381 tgatcatcaa gctgcctaag tactccctgt tcgagctgga aaacggccgg aagagaatgc
     7441 tggcctctgc cggcgaactg cagaagggaa acgaactggc cctgccctcc aaatatgtga
     7501 acttcctgta cctggccagc cactatgaga agctgaaggg ctcccccgag gataatgagc
     7561 agaaacagct gtttgtggaa cagcacaagc actacctgga cgagatcatc gagcagatca
     7621 gcgagttctc caagagagtg atcctggccg acgctaatct ggacaaagtg ctgtccgcct
     7681 acaacaagca ccgggataag cccatcagag agcaggccga gaatatcatc cacctgttta
     7741 ccctgaccaa tctgggagcc cctgccgcct tcaagtactt tgacaccacc atcgaccgga
     7801 agaggtacac cagcaccaaa gaggtgctgg acgccaccct gatccaccag agcatcaccg
     7861 gcctgtacga gacacggatc gacctgtctc agctgggagg cgacaagcgt cctgctgcta
     7921 ctaagaaagc tggtcaagct aagaaaaaga aagctagcgg aggaagcggc gccaccaatt
     7981 tcagcctgct gaaacaggcc ggcgacgtgg aagagaaccc tggccctatg gtgtccaagg
     8041 gcgaagagga caacatggcc agcctgcctg ccacccacga gctgcacatc ttcggcagca
     8101 tcaacggcgt ggacttcgac atggtgggac agggcaccgg caaccccaac gacggctacg
     8161 aggaactgaa cctgaagtcc acaaagggcg acctgcagtt cagcccctgg attctggtgc
     8221 cccacatcgg ctacggcttc caccagtacc tgccctaccc cgacggcatg agccctttcc
     8281 aagccgccat ggtggatggc agcggctacc aggtgcaccg gaccatgcag tttgaggacg
     8341 gcgccagcct gaccgtgaac tacaggtaca catacgaggg cagccacatc aagggcgagg
     8401 cccaagtgaa gggcacaggc tttccagccg acggccccgt gatgaccaat agcctgacag
     8461 ccgccgactg gtgcagaagc aagaaaacct accccaatga caagaccatc atcagcacct
     8521 tcaagtggtc ctacaccacc ggcaatggca agcggtacag aagcaccgcc cggaccacct
     8581 acaccttcgc caaacccatg gccgccaact acctgaagaa ccagcctatg tacgtgttcc
     8641 gcaagaccga gctgaagcac tccaagacag aactgaactt caaagagtgg cagaaagcct
     8701 tcaccgacgt gatgggcatg gacgagctgt acaagtgaac gcgttaagtc gacaatcaac
     8761 ctctggatta caaaatttgt gaaagattga ctggtattct taactatgtt gctcctttta
     8821 cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt
     8881 tcattttctc ctccttgtat aaatcctggt tgctgtctct ttatgaggag ttgtggcccg
     8941 ttgtcaggca acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc actggttggg
     9001 gcattgccac cacctgtcag ctcctttccg ggactttcgc tttccccctc cctattgcca
     9061 cggcggaact catcgccgcc tgccttgccc gctgctggac aggggctcgg ctgttgggca
     9121 ctgacaattc cgtggtgttg tcggggaaat catcgtcctt tccttggctg ctcgcctgtg
     9181 ttgccacctg gattctgcgc gggacgtcct tctgctacgt cccttcggcc ctcaatccag
     9241 cggaccttcc ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc
     9301 gccctcagac gagtcggatc tccctttggg ccgcctcccc gcgtcgactt taagaccaat
     9361 gacttacaag gcagctgtag atcttagcca ctttttaaaa gaaaaggggg gactggaagg
     9421 gctaattcac tcccaacgaa gacaagatct gctttttgct tgtactgggt ctctctggtt
     9481 agaccagatc tgagcctggg agctctctgg ctaactaggg aacccactgc ttaagcctca
     9541 ataaagcttg ccttgagtgc ttcaagtagt gtgtgcccgt ctgttgtgtg actctggtaa
     9601 ctagagatcc ctcagaccct tttagtcagt gtggaaaatc tctagcagta cgtatagtag
     9661 ttcatgtcat cttattattc agtatttata acttgcaaag aaatgaatat cagagagtga
     9721 gaggaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
     9781 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
     9841 atcttatcat gtctggctct agctatcccg cccctaactc cgcccatccc gcccctaact
     9901 ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag
     9961 gccgaggccg cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc
    10021 ctagggacgt acccaattcg ccctatagtg agtcgtatta cgcgcgctca ctggccgtcg
    10081 ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc cttgcagcac
    10141 atcccccttt cgccagctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac
    10201 agttgcgcag cctgaatggc gaatgggacg cgccctgtag cggcgcatta agcgcggcgg
    10261 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt
    10321 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
    10381 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
    10441 attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga
    10501 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc
    10561 ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa
    10621 aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta acgcttacaa
    10681 tttaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
    10741 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg
    10801 aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc
    10861 attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga
    10921 tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga
    10981 gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg
    11041 cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc
    11101 tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac
    11161 agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact
    11221 tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca
    11281 tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg
    11341 tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact
    11401 acttactcta gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg
    11461 accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg
    11521 tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat
    11581 cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc
    11641 tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat
    11701 actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt
    11761 tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
    11821 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
    11881 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
    11941 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt
    12001 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
    12061 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
    12121 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
    12181 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
    12241 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
    12301 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
    12361 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
    12421 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
    12481 ttttgc
//
