LOCUS       sequence-229088-       12480 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral CRISPR-Cas9 delivery for sgRNA (hU6), dTomato 
            coexpression, EFS Promoter driven.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 12480)
  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 12480)
  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.dTomato
FEATURES             Location/Qualifiers
     source          1..12480
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..249
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1..21
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     172..191
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     promoter        411..622
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      642..651
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             651..674
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             681..701
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             726..4826
                     /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             726..4826
                     /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             4827..4874
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             4893..4949
                     /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             4950..5651
                     /codon_start=1
                     /product="dimeric variant of DsRed fluorescent protein 
                     (Shaner et al., 2004)"
                     /label=dTomato
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFLYGMDELYK"
     primer_bind     complement(5083..5101)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     5351..5370
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     5545..5568
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     misc_feature    5670..6258
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5723..5743)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(6141..6152)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             6330..6563
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    6641..6762
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6678..6697)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6732..6751
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      6802..6937
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(6957..6976)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6958..6976)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6986..7003)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6986..7002)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6995..7017)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      7144..7599
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7231..7250)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7441..7462
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        7625..7729
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7730..7798,7799..8590)
                     /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(7948..7967)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8761..9349
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     9250..9269
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     9503..9520
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    9637..9658
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(9673..9678,9679..9696,9697..9703)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    9711..9727
                     /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     9716..9738
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     9735..9751
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     9735..9751
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     9770..9790
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        9772..9790
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     enhancer        9835..10214
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     enhancer        10271..10462
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(10294..10314)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        10481..10707
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             10708..10888
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    10935..11060
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    11553..11786
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             11971..12015
                     /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    12313..12430
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
ORIGIN
        1 gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag
       61 ataattagaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga
      121 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat
      181 atgcttaccg taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga
      241 cgaaacaccg gagacgtatg tcgacaactc gtctctgttt aagagctatg ctggaaacag
      301 catagcaagt ttaaataagg ctagtccgtt atcaacttga aaaagtggca ccgagtcggt
      361 gcttttttga attctcctct agaactcgag tggctccggt gcccgtcagt gggcagagcg
      421 cacatcgccc acagtccccg agaagttggg gggaggggtc ggcaattgaa ccggtgccta
      481 gagaaggtgg cgcggggtaa actgggaaag tgatgtcgtg tactggctcc gcctttttcc
      541 cgagggtggg ggagaaccgt atataagtgc agtagtcgcc gtgaacgttc tttttcgcaa
      601 cgggtttgcc gccagaacac aggtgtcgtg acgcgggatc cgccaccatg gattacaaag
      661 acgatgacga taagatggcc ccaaagaaga agcggaaggt cggtatccac ggagtcccag
      721 cagccgacaa gaagtacagc atcggcctgg acatcggcac caactctgtg ggctgggccg
      781 tgatcaccga cgagtacaag gtgcccagca agaaattcaa ggtgctgggc aacaccgacc
      841 ggcacagcat caagaagaac ctgatcggag ccctgctgtt cgacagcggc gaaacagccg
      901 aggccacccg gctgaagaga accgccagaa gaagatacac cagacggaag aaccggatct
      961 gctatctgca agagatcttc agcaacgaga tggccaaggt ggacgacagc ttcttccaca
     1021 gactggaaga gtccttcctg gtggaagagg ataagaagca cgagcggcac cccatcttcg
     1081 gcaacatcgt ggacgaggtg gcctaccacg agaagtaccc caccatctac cacctgagaa
     1141 agaaactggt ggacagcacc gacaaggccg acctgcggct gatctatctg gccctggccc
     1201 acatgatcaa gttccggggc cacttcctga tcgagggcga cctgaacccc gacaacagcg
     1261 acgtggacaa gctgttcatc cagctggtgc agacctacaa ccagctgttc gaggaaaacc
     1321 ccatcaacgc cagcggcgtg gacgccaagg ccatcctgtc tgccagactg agcaagagca
     1381 gacggctgga aaatctgatc gcccagctgc ccggcgagaa gaagaatggc ctgttcggaa
     1441 acctgattgc cctgagcctg ggcctgaccc ccaacttcaa gagcaacttc gacctggccg
     1501 aggatgccaa actgcagctg agcaaggaca cctacgacga cgacctggac aacctgctgg
     1561 cccagatcgg cgaccagtac gccgacctgt ttctggccgc caagaacctg tccgacgcca
     1621 tcctgctgag cgacatcctg agagtgaaca ccgagatcac caaggccccc ctgagcgcct
     1681 ctatgatcaa gagatacgac gagcaccacc aggacctgac cctgctgaaa gctctcgtgc
     1741 ggcagcagct gcctgagaag tacaaagaga ttttcttcga ccagagcaag aacggctacg
     1801 ccggctacat tgacggcgga gccagccagg aagagttcta caagttcatc aagcccatcc
     1861 tggaaaagat ggacggcacc gaggaactgc tcgtgaagct gaacagagag gacctgctgc
     1921 ggaagcagcg gaccttcgac aacggcagca tcccccacca gatccacctg ggagagctgc
     1981 acgccattct gcggcggcag gaagattttt acccattcct gaaggacaac cgggaaaaga
     2041 tcgagaagat cctgaccttc cgcatcccct actacgtggg ccctctggcc aggggaaaca
     2101 gcagattcgc ctggatgacc agaaagagcg aggaaaccat caccccctgg aacttcgagg
     2161 aagtggtgga caagggcgct tccgcccaga gcttcatcga gcggatgacc aacttcgata
     2221 agaacctgcc caacgagaag gtgctgccca agcacagcct gctgtacgag tacttcaccg
     2281 tgtataacga gctgaccaaa gtgaaatacg tgaccgaggg aatgagaaag cccgccttcc
     2341 tgagcggcga gcagaaaaag gccatcgtgg acctgctgtt caagaccaac cggaaagtga
     2401 ccgtgaagca gctgaaagag gactacttca agaaaatcga gtgcttcgac tccgtggaaa
     2461 tctccggcgt ggaagatcgg ttcaacgcct ccctgggcac ataccacgat ctgctgaaaa
     2521 ttatcaagga caaggacttc ctggacaatg aggaaaacga ggacattctg gaagatatcg
     2581 tgctgaccct gacactgttt gaggacagag agatgatcga ggaacggctg aaaacctatg
     2641 cccacctgtt cgacgacaaa gtgatgaagc agctgaagcg gcggagatac accggctggg
     2701 gcaggctgag ccggaagctg atcaacggca tccgggacaa gcagtccggc aagacaatcc
     2761 tggatttcct gaagtccgac ggcttcgcca acagaaactt catgcagctg atccacgacg
     2821 acagcctgac ctttaaagag gacatccaga aagcccaggt gtccggccag ggcgatagcc
     2881 tgcacgagca cattgccaat ctggccggca gccccgccat taagaagggc atcctgcaga
     2941 cagtgaaggt ggtggacgag ctcgtgaaag tgatgggccg gcacaagccc gagaacatcg
     3001 tgatcgaaat ggccagagag aaccagacca cccagaaggg acagaagaac agccgcgaga
     3061 gaatgaagcg gatcgaagag ggcatcaaag agctgggcag ccagatcctg aaagaacacc
     3121 ccgtggaaaa cacccagctg cagaacgaga agctgtacct gtactacctg cagaatgggc
     3181 gggatatgta cgtggaccag gaactggaca tcaaccggct gtccgactac gatgtggacc
     3241 atatcgtgcc tcagagcttt ctgaaggacg actccatcga caacaaggtg ctgaccagaa
     3301 gcgacaagaa ccggggcaag agcgacaacg tgccctccga agaggtcgtg aagaagatga
     3361 agaactactg gcggcagctg ctgaacgcca agctgattac ccagagaaag ttcgacaatc
     3421 tgaccaaggc cgagagaggc ggcctgagcg aactggataa ggccggcttc atcaagagac
     3481 agctggtgga aacccggcag atcacaaagc acgtggcaca gatcctggac tcccggatga
     3541 acactaagta cgacgagaat gacaagctga tccgggaagt gaaagtgatc accctgaagt
     3601 ccaagctggt gtccgatttc cggaaggatt tccagtttta caaagtgcgc gagatcaaca
     3661 actaccacca cgcccacgac gcctacctga acgccgtcgt gggaaccgcc ctgatcaaaa
     3721 agtaccctaa gctggaaagc gagttcgtgt acggcgacta caaggtgtac gacgtgcgga
     3781 agatgatcgc caagagcgag caggaaatcg gcaaggctac cgccaagtac ttcttctaca
     3841 gcaacatcat gaactttttc aagaccgaga ttaccctggc caacggcgag atccggaagc
     3901 ggcctctgat cgagacaaac ggcgaaaccg gggagatcgt gtgggataag ggccgggatt
     3961 ttgccaccgt gcggaaagtg ctgagcatgc cccaagtgaa tatcgtgaaa aagaccgagg
     4021 tgcagacagg cggcttcagc aaagagtcta tcctgcccaa gaggaacagc gataagctga
     4081 tcgccagaaa gaaggactgg gaccctaaga agtacggcgg cttcgacagc cccaccgtgg
     4141 cctattctgt gctggtggtg gccaaagtgg aaaagggcaa gtccaagaaa ctgaagagtg
     4201 tgaaagagct gctggggatc accatcatgg aaagaagcag cttcgagaag aatcccatcg
     4261 actttctgga agccaagggc tacaaagaag tgaaaaagga cctgatcatc aagctgccta
     4321 agtactccct gttcgagctg gaaaacggcc ggaagagaat gctggcctct gccggcgaac
     4381 tgcagaaggg aaacgaactg gccctgccct ccaaatatgt gaacttcctg tacctggcca
     4441 gccactatga gaagctgaag ggctcccccg aggataatga gcagaaacag ctgtttgtgg
     4501 aacagcacaa gcactacctg gacgagatca tcgagcagat cagcgagttc tccaagagag
     4561 tgatcctggc cgacgctaat ctggacaaag tgctgtccgc ctacaacaag caccgggata
     4621 agcccatcag agagcaggcc gagaatatca tccacctgtt taccctgacc aatctgggag
     4681 cccctgccgc cttcaagtac tttgacacca ccatcgaccg gaagaggtac accagcacca
     4741 aagaggtgct ggacgccacc ctgatccacc agagcatcac cggcctgtac gagacacgga
     4801 tcgacctgtc tcagctggga ggcgacaagc gtcctgctgc tactaagaaa gctggtcaag
     4861 ctaagaaaaa gaaagctagc ggaggaagcg gcgccaccaa tttcagcctg ctgaaacagg
     4921 ccggcgacgt ggaagagaac cctggcccta tggtgagcaa gggcgaggag gtcatcaaag
     4981 agttcatgcg cttcaaggtg cgcatggagg gctccatgaa cggccacgag ttcgagatcg
     5041 agggcgaggg cgagggccgc ccctacgagg gcacccagac cgccaagctg aaggtgacca
     5101 agggcggccc cctgcccttc gcctgggaca tcctgtcccc ccagttcatg tacggctcca
     5161 aggcgtacgt gaagcacccc gccgacatcc ccgattacaa gaagctgtcc ttccccgagg
     5221 gcttcaagtg ggagcgcgtg atgaacttcg aggacggcgg tctggtgacc gtgacccagg
     5281 actcctccct gcaggacggc acgctgatct acaaggtgaa gatgcgcggc accaacttcc
     5341 cccccgacgg ccccgtaatg cagaagaaga ccatgggctg ggaggcctcc accgagcgcc
     5401 tgtacccccg cgacggcgtg ctgaagggcg agatccacca ggccctgaag ctgaaggacg
     5461 gcggccacta cctggtggag ttcaagacca tctacatggc caagaagccc gtgcaactgc
     5521 ccggctacta ctacgtggac accaagctgg acatcacctc ccacaacgag gactacacca
     5581 tcgtggaaca gtacgagcgc tccgagggcc gccaccacct gttcctgtac ggcatggacg
     5641 agctgtacaa gtaaacgcgt taagtcgaca atcaacctct ggattacaaa atttgtgaaa
     5701 gattgactgg tattcttaac tatgttgctc cttttacgct atgtggatac gctgctttaa
     5761 tgcctttgta tcatgctatt gcttcccgta tggctttcat tttctcctcc ttgtataaat
     5821 cctggttgct gtctctttat gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt
     5881 gcactgtgtt tgctgacgca acccccactg gttggggcat tgccaccacc tgtcagctcc
     5941 tttccgggac tttcgctttc cccctcccta ttgccacggc ggaactcatc gccgcctgcc
     6001 ttgcccgctg ctggacaggg gctcggctgt tgggcactga caattccgtg gtgttgtcgg
     6061 ggaaatcatc gtcctttcct tggctgctcg cctgtgttgc cacctggatt ctgcgcggga
     6121 cgtccttctg ctacgtccct tcggccctca atccagcgga ccttccttcc cgcggcctgc
     6181 tgccggctct gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt cggatctccc
     6241 tttgggccgc ctccccgcgt cgactttaag accaatgact tacaaggcag ctgtagatct
     6301 tagccacttt ttaaaagaaa aggggggact ggaagggcta attcactccc aacgaagaca
     6361 agatctgctt tttgcttgta ctgggtctct ctggttagac cagatctgag cctgggagct
     6421 ctctggctaa ctagggaacc cactgcttaa gcctcaataa agcttgcctt gagtgcttca
     6481 agtagtgtgt gcccgtctgt tgtgtgactc tggtaactag agatccctca gaccctttta
     6541 gtcagtgtgg aaaatctcta gcagtacgta tagtagttca tgtcatctta ttattcagta
     6601 tttataactt gcaaagaaat gaatatcaga gagtgagagg aacttgttta ttgcagctta
     6661 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     6721 gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct ggctctagct
     6781 atcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
     6841 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc ggcctctgag
     6901 ctattccaga agtagtgagg aggctttttt ggaggcctag ggacgtaccc aattcgccct
     6961 atagtgagtc gtattacgcg cgctcactgg ccgtcgtttt acaacgtcgt gactgggaaa
     7021 accctggcgt tacccaactt aatcgccttg cagcacatcc ccctttcgcc agctggcgta
     7081 atagcgaaga ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg aatggcgaat
     7141 gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
     7201 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
     7261 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
     7321 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg
     7381 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
     7441 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt
     7501 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
     7561 ttaacgcgaa ttttaacaaa atattaacgc ttacaattta ggtggcactt ttcggggaaa
     7621 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     7681 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     7741 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     7801 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     7861 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     7921 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc
     7981 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     8041 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     8101 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     8161 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     8221 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat
     8281 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     8341 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     8401 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     8461 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     8521 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     8581 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     8641 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     8701 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     8761 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     8821 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     8881 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
     8941 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     9001 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     9061 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     9121 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     9181 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     9241 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     9301 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     9361 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     9421 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
     9481 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
     9541 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
     9601 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
     9661 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
     9721 ataacaattt cacacaggaa acagctatga ccatgattac gccaagcgcg caattaaccc
     9781 tcactaaagg gaacaaaagc tggagctgca agcggtaccg tcgacctgca ggttgacatt
     9841 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
     9901 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
     9961 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
    10021 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
    10081 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
    10141 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
    10201 tcgctattac catggcgcgc catgcagggc ctgaaataac ctctgaaaga ggaacttggt
    10261 taggtacctt ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa
    10321 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
    10381 ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
    10441 attagtcagc aaccatagtc ccttaagagg cgcgccttaa tgtagtctta tgcaatactc
    10501 ttgtagtctt gcaacatggt aacgatgagt tagcaacatg ccttacaagg agagaaaaag
    10561 caccgtgcat gccgattggt ggaagtaagg tggtacgatc gtgccttatt aggaaggcaa
    10621 cagacgggtc tgacatggat tggacgaacc actgaattgc cgcattgcag agatattgta
    10681 tttaagtgcc tagctcgata cataaacggg tctctctggt tagaccagat ctgagcctgg
    10741 gagctctctg gctaactagg gaacccactg cttaagcctc aataaagctt gccttgagtg
    10801 cttcaagtag tgtgtgcccg tctgttgtgt gactctggta actagagatc cctcagaccc
    10861 ttttagtcag tgtggaaaat ctctagcagt ggcgcccgaa cagggacttg aaagcgaaag
    10921 ggaaaccaga ggagctctct cgacgcagga ctcggcttgc tgaagcgcgc acggcaagag
    10981 gcgaggggcg gcgactggtg agtacgccaa aaattttgac tagcggaggc tagaaggaga
    11041 gagatgggtg cgagagcgtc agtattaagc gggggagaat tagatcgcga tgggaaaaaa
    11101 ttcggttaag gccaggggga aagaaaaaat ataaattaaa acatatagta tgggcaagca
    11161 gggagctaga acgattcgca gttaatcctg gcctgttaga aacatcagaa ggctgtagac
    11221 aaatactggg acagctacaa ccatcccttc agacaggatc agaagaactt agatcattat
    11281 ataatacagt agcaaccctc tattgtgtgc atcaaaggat agagataaaa gacaccaagg
    11341 aagctttaga caagatagag gaagagcaaa acaaaagtaa gaccaccgca cagcaagcgg
    11401 ccgctgatct tcagacctgg aggaggagat atgagggaca attggagaag tgaattatat
    11461 aaatataaag tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc aaagagaaga
    11521 gtggtgcaga gagaaaaaag agcagtggga ataggagctt tgttccttgg gttcttggga
    11581 gcagcaggaa gcactatggg cgcagcgtca atgacgctga cggtacaggc cagacaatta
    11641 ttgtctggta tagtgcagca gcagaacaat ttgctgaggg ctattgaggc gcaacagcat
    11701 ctgttgcaac tcacagtctg gggcatcaag cagctccagg caagaatcct ggctgtggaa
    11761 agatacctaa aggatcaaca gctcctgggg atttggggtt gctctggaaa actcatttgc
    11821 accactgctg tgccttggaa tgctagttgg agtaataaat ctctggaaca gatttggaat
    11881 cacacgacct ggatggagtg ggacagagaa attaacaatt acacaagctt aatacactcc
    11941 ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt ggaattagat
    12001 aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt ggctgtggta tataaaatta
    12061 ttcataatga tagtaggagg cttggtaggt ttaagaatag tttttgctgt actttctata
    12121 gtgaatagag ttaggcaggg atattcacca ttatcgtttc agacccacct cccaaccccg
    12181 aggggacccg acaggcccga aggaatagaa gaagaaggtg gagagagaga cagagacaga
    12241 tccattcgat tagtgaacgg atcggcactg cgtgcgccaa ttctgcagac aaatggcagt
    12301 attcatccac aattttaaaa gaaaaggggg gattgggggg tacagtgcag gggaaagaat
    12361 agtagacata atagcaacag acatacaaac taaagaatta caaaaacaaa ttacaaaaat
    12421 tcaaaatttt cgggtttatt acagggacag cagagatcca gtttggttaa ttaaggtacc
//
