LOCUS       sequence-307049-       13374 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Expresses dCas9-P2A-EGFP driven by human SYN promoter and empty 
            CRISPR Display sgRNA/accessory RNA from U6 promoter. .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13374)
  AUTHORS   Carullo NVN, Phillips Iii RA, Simon RC, Soto SAR, Hinds JE, 
            Salisbury AJ, Revanna JS, Bunner KD, Ianov L, Sultan FA, Savell KE, 
            Gersbach CA, Day JJ
  TITLE     Enhancer RNAs predict enhancer-gene regulatory links and are 
            critical for enhancer function in neuronal systems.
  JOURNAL   Nucleic Acids Res. 2020 Aug 18. pii: 5893972. doi: 
            10.1093/nar/gkaa671.
  PUBMED    32810208
REFERENCE   2  (bases 1 to 13374)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nucleic 
            Acids Res. 2020 Aug 18. pii: 5893972. doi: 10.1093/nar/gkaa671."
COMMENT     Sequence Label: lenti U6-sgRNA-acRNA SYN-dCas9-P2A-EGFP
FEATURES             Location/Qualifiers
     source          1..13374
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(47..66)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        238..617
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        619..817
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     772..792
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             835..1015
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1062..1187
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1680..1913
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2098..2142
                     /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    2440..2557
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     primer_bind     2610..2630
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        2612..2630
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     misc_RNA        complement(2672..2747)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        complement(2766..3014)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     complement(2824..2843)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(2994..3014)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     promoter        3092..3539
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     regulatory      3557..3566
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3563..3589,3590..3592,3593..6079,6080..6082,
                     6083..7666)
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=dCas9
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the D10A mutation in the RuvC 
                     catalytic domain and the H840A mutation in the HNH 
                     catalytic domain"
                     /translation="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             3563..7666
                     /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="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             7667..7714
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             7715..7738
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             7748..7804
                     /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             join(7808..7810,7811..7813,7814..8527)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(7808..7810,7811..7813,7814..8524)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(8114..8133)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     8461..8482
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    8552..9140
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8605..8625)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(9023..9034)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(9143..9159)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(9144..9160)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             9179..9359
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(9385..9402)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    9391..9615
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      9661..10089
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9748..9767)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     9958..9979
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(10098..10118)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        10103..10432
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      10283..10418
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     10345..10364
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        10480..10527
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             10546..10920
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    11050..11171
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(11087..11106)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     11141..11160
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(11220..11236)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(11220..11236)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(11233..11255)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    11244..11260
                     /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)."
     promoter        complement(join(11268..11274,11275..11292,11293..11298))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11313..11334
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(11451..11468)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(11622..12210)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(11702..11721)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(12381..13172,13173..13241))
                     /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     13004..13023
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(13242..13346)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg
       61 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
      121 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc
      181 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac
      241 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat
      301 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
      361 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
      421 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
      481 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
      541 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
      601 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt
      661 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc
      721 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg
      781 gcggtaggcg tgtacggtgg gaggtctata taagcagcgc gttttgcctg tactgggtct
      841 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt
      901 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac
      961 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc
     1021 gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc
     1081 ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa
     1141 ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg
     1201 ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata
     1261 aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc
     1321 tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga
     1381 caggatcaga agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc
     1441 aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca
     1501 aaagtaagac caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg
     1561 agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga accattagga
     1621 gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata
     1681 ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg
     1741 acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg
     1801 ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag
     1861 ctccaggcaa gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt
     1921 tggggttgct ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt
     1981 aataaatctc tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt
     2041 aacaattaca caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag
     2101 aatgaacaag aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata
     2161 acaaattggc tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta
     2221 agaatagttt ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta
     2281 tcgtttcaga cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa
     2341 gaaggtggag agagagacag agacagatcc attcgattag tgaacggatc ggcactgcgt
     2401 gcgccaattc tgcagacaaa tggcagtatt catccacaat tttaaaagaa aaggggggat
     2461 tggggggtac agtgcagggg aaagaatagt agacataata gcaacagaca tacaaactaa
     2521 agaattacaa aaacaaatta caaaaattca aaattttcgg gtttattaca gggacagcag
     2581 agatccagtt tggttaatta accaagcgcg caattaaccc tcactaaagg gaacaaaagc
     2641 tggagctcca ccgcggaaaa aaacagagac ggcaccgact cggtgccact ttttcaagtt
     2701 gataacggac tagccttatt ttaacttgct atttctagct ctaaaacagg tcttctcgaa
     2761 gacccggtgt ttcgtccttt ccacaagata tataaagcca agaaatcgaa atactttcaa
     2821 gttacggtaa gcatatgata gtccatttta aaacataatt ttaaaactgc aaactaccca
     2881 agaaattatt actttctacg tcacgtattt tgtactaata tctttgtgtt tacagtcaaa
     2941 ttaattccaa ttatctctct aacagccttg tatcgtatat gcaaatatga aggaatcatg
     3001 ggaaataggc cctcggtacc ttaacccgtg tcggctgcta gctaggtctt gaaaggagtg
     3061 ggaattggct ccggtgcccg tcagtttcga aagtgcaagt gggttttagg accaggatga
     3121 ggcggggtgg gggtgcctac ctgacgaccg accccgaccc actggacaag cacccaaccc
     3181 ccattcccca aattgcgcat cccctatcag agagggggag gggaaacagg atgcggcgag
     3241 gcgcgtgcgc actgccagct tcagcaccgc ggacagtgcc ttcgcccccg cctggcggcg
     3301 cgcgccaccg ccgcctcagc actgaaggcg cgctgacgtc actcgccggt cccccgcaaa
     3361 ctccccttcc cggccacctt ggtcgcgtcc gcgccgccgc cggcccagcc ggaccgcacc
     3421 acgcgaggcg cgagataggg gggcacgggc gcgaccatct gcgctgcggc gccggcgact
     3481 cagcgctgcc tcagtctgcg gtgggcagcg gaggagtcgt gtcgtgcctg agagcgcaga
     3541 ccggttctag agcgctgcca ccatggacaa gaagtacagc atcggcctgg ccatcggcac
     3601 caactctgtg ggctgggccg tgatcaccga cgagtacaag gtgcccagca agaaattcaa
     3661 ggtgctgggc aacaccgacc ggcacagcat caagaagaac ctgatcggag ccctgctgtt
     3721 cgacagcggc gaaacagccg aggccacccg gctgaagaga accgccagaa gaagatacac
     3781 cagacggaag aaccggatct gctatctgca agagatcttc agcaacgaga tggccaaggt
     3841 ggacgacagc ttcttccaca gactggaaga gtccttcctg gtggaagagg ataagaagca
     3901 cgagcggcac cccatcttcg gcaacatcgt ggacgaggtg gcctaccacg agaagtaccc
     3961 caccatctac cacctgagaa agaaactggt ggacagcacc gacaaggccg acctgcggct
     4021 gatctatctg gccctggccc acatgatcaa gttccggggc cacttcctga tcgagggcga
     4081 cctgaacccc gacaacagcg acgtggacaa gctgttcatc cagctggtgc agacctacaa
     4141 ccagctgttc gaggaaaacc ccatcaacgc cagcggcgtg gacgccaagg ccatcctgtc
     4201 tgccagactg agcaagagca gacggctgga aaatctgatc gcccagctgc ccggcgagaa
     4261 gaagaatggc ctgttcggaa acctgattgc cctgagcctg ggcctgaccc ccaacttcaa
     4321 gagcaacttc gacctggccg aggatgccaa actgcagctg agcaaggaca cctacgacga
     4381 cgacctggac aacctgctgg cccagatcgg cgaccagtac gccgacctgt ttctggccgc
     4441 caagaacctg tccgacgcca tcctgctgag cgacatcctg agagtgaaca ccgagatcac
     4501 caaggccccc ctgagcgcct ctatgatcaa gagatacgac gagcaccacc aggacctgac
     4561 cctgctgaaa gctctcgtgc ggcagcagct gcctgagaag tacaaagaga ttttcttcga
     4621 ccagagcaag aacggctacg ccggctacat tgacggcgga gccagccagg aagagttcta
     4681 caagttcatc aagcccatcc tggaaaagat ggacggcacc gaggaactgc tcgtgaagct
     4741 gaacagagag gacctgctgc ggaagcagcg gaccttcgac aacggcagca tcccccacca
     4801 gatccacctg ggagagctgc acgccattct gcggcggcag gaagattttt acccattcct
     4861 gaaggacaac cgggaaaaga tcgagaagat cctgaccttc cgcatcccct actacgtggg
     4921 ccctctggcc aggggaaaca gcagattcgc ctggatgacc agaaagagcg aggaaaccat
     4981 caccccctgg aacttcgagg aagtggtgga caagggcgct tccgcccaga gcttcatcga
     5041 gcggatgacc aacttcgata agaacctgcc caacgagaag gtgctgccca agcacagcct
     5101 gctgtacgag tacttcaccg tgtataacga gctgaccaaa gtgaaatacg tgaccgaggg
     5161 aatgagaaag cccgccttcc tgagcggcga gcagaaaaag gccatcgtgg acctgctgtt
     5221 caagaccaac cggaaagtga ccgtgaagca gctgaaagag gactacttca agaaaatcga
     5281 gtgcttcgac tccgtggaaa tctccggcgt ggaagatcgg ttcaacgcct ccctgggcac
     5341 ataccacgat ctgctgaaaa ttatcaagga caaggacttc ctggacaatg aggaaaacga
     5401 ggacattctg gaagatatcg tgctgaccct gacactgttt gaggacagag agatgatcga
     5461 ggaacggctg aaaacctatg cccacctgtt cgacgacaaa gtgatgaagc agctgaagcg
     5521 gcggagatac accggctggg gcaggctgag ccggaagctg atcaacggca tccgggacaa
     5581 gcagtccggc aagacaatcc tggatttcct gaagtccgac ggcttcgcca acagaaactt
     5641 catgcagctg atccacgacg acagcctgac ctttaaagag gacatccaga aagcccaggt
     5701 gtccggccag ggcgatagcc tgcacgagca cattgccaat ctggccggca gccccgccat
     5761 taagaagggc atcctgcaga cagtgaaggt ggtggacgag ctcgtgaaag tgatgggccg
     5821 gcacaagccc gagaacatcg tgatcgaaat ggccagagag aaccagacca cccagaaggg
     5881 acagaagaac agccgcgaga gaatgaagcg gatcgaagag ggcatcaaag agctgggcag
     5941 ccagatcctg aaagaacacc ccgtggaaaa cacccagctg cagaacgaga agctgtacct
     6001 gtactacctg cagaatgggc gggatatgta cgtggaccag gaactggaca tcaaccggct
     6061 gtccgactac gatgtggacg ctatcgtgcc tcagagcttt ctgaaggacg actccatcga
     6121 caacaaggtg ctgaccagaa gcgacaagaa ccggggcaag agcgacaacg tgccctccga
     6181 agaggtcgtg aagaagatga agaactactg gcggcagctg ctgaacgcca agctgattac
     6241 ccagagaaag ttcgacaatc tgaccaaggc cgagagaggc ggcctgagcg aactggataa
     6301 ggccggcttc atcaagagac agctggtgga aacccggcag atcacaaagc acgtggcaca
     6361 gatcctggac tcccggatga acactaagta cgacgagaat gacaagctga tccgggaagt
     6421 gaaagtgatc accctgaagt ccaagctggt gtccgatttc cggaaggatt tccagtttta
     6481 caaagtgcgc gagatcaaca actaccacca cgcccacgac gcctacctga acgccgtcgt
     6541 gggaaccgcc ctgatcaaaa agtaccctaa gctggaaagc gagttcgtgt acggcgacta
     6601 caaggtgtac gacgtgcgga agatgatcgc caagagcgag caggaaatcg gcaaggctac
     6661 cgccaagtac ttcttctaca gcaacatcat gaactttttc aagaccgaga ttaccctggc
     6721 caacggcgag atccggaagc ggcctctgat cgagacaaac ggcgaaaccg gggagatcgt
     6781 gtgggataag ggccgggatt ttgccaccgt gcggaaagtg ctgagcatgc cccaagtgaa
     6841 tatcgtgaaa aagaccgagg tgcagacagg cggcttcagc aaagagtcta tcctgcccaa
     6901 gaggaacagc gataagctga tcgccagaaa gaaggactgg gaccctaaga agtacggcgg
     6961 cttcgacagc cccaccgtgg cctattctgt gctggtggtg gccaaagtgg aaaagggcaa
     7021 gtccaagaaa ctgaagagtg tgaaagagct gctggggatc accatcatgg aaagaagcag
     7081 cttcgagaag aatcccatcg actttctgga agccaagggc tacaaagaag tgaaaaagga
     7141 cctgatcatc aagctgccta agtactccct gttcgagctg gaaaacggcc ggaagagaat
     7201 gctggcctct gccggcgaac tgcagaaggg aaacgaactg gccctgccct ccaaatatgt
     7261 gaacttcctg tacctggcca gccactatga gaagctgaag ggctcccccg aggataatga
     7321 gcagaaacag ctgtttgtgg aacagcacaa gcactacctg gacgagatca tcgagcagat
     7381 cagcgagttc tccaagagag tgatcctggc cgacgctaat ctggacaaag tgctgtccgc
     7441 ctacaacaag caccgggata agcccatcag agagcaggcc gagaatatca tccacctgtt
     7501 taccctgacc aatctgggag cccctgccgc cttcaagtac tttgacacca ccatcgaccg
     7561 gaagaggtac accagcacca aagaggtgct ggacgccacc ctgatccacc agagcatcac
     7621 cggcctgtac gagacacgga tcgacctgtc tcagctggga ggcgacaagc gacctgccgc
     7681 cacaaagaag gctggacagg ctaagaagaa gaaagattac aaagacgatg acgataaggg
     7741 atccggcgca acaaacttct ctctgctgaa acaagccgga gatgtcgaag agaatcctgg
     7801 accgaccatg gtgagcaaag gcgaggagct gttcaccggg gtggtgccca tcctggtcga
     7861 gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc
     7921 cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg
     7981 gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct accccgacca
     8041 catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac
     8101 catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga
     8161 caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct
     8221 ggggcacaag ctggagtaca actacaacag ccacaacgtc tatatcatgg ccgacaagca
     8281 gaagaacggc atcaaggtga acttcaagat ccgccacaac atcgaggacg gcagcgtgca
     8341 gctcgccgac cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga
     8401 caaccactac ctgagcaccc agtccgccct gagcaaagac cccaacgaga agcgcgatca
     8461 catggtcctg ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta
     8521 caagtaagaa ttcgatatca agcttatcga taatcaacct ctggattaca aaatttgtga
     8581 aagattgact ggtattctta actatgttgc tccttttacg ctatgtggat acgctgcttt
     8641 aatgcctttg tatcatgcta ttgcttcccg tatggctttc attttctcct ccttgtataa
     8701 atcctggttg ctgtctcttt atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt
     8761 gtgcactgtg tttgctgacg caacccccac tggttggggc attgccacca cctgtcagct
     8821 cctttccggg actttcgctt tccccctccc tattgccacg gcggaactca tcgccgcctg
     8881 ccttgcccgc tgctggacag gggctcggct gttgggcact gacaattccg tggtgttgtc
     8941 ggggaaatca tcgtcctttc cttggctgct cgcctgtgtt gccacctgga ttctgcgcgg
     9001 gacgtccttc tgctacgtcc cttcggccct caatccagcg gaccttcctt cccgcggcct
     9061 gctgccggct ctgcggcctc ttccgcctct tcgccttcgc cctcagacga gtcggatctc
     9121 cctttgggcc gcctccccgc atcgataccg tcgacctcga gaccttccgg actgtactgg
     9181 gtctctctgg ttagaccaga tctgagcctg ggagctctct ggctaactag ggaacccact
     9241 gcttaagcct caataaagct tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg
     9301 tgactctggt aactagagat ccctcagacc cttttagtca gtgtggaaaa tctctagcag
     9361 ggcccgttta aacccgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg
     9421 tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct
     9481 aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg
     9541 gggtggggca ggacagcaag ggggaggatt gggaagagaa tagcaggcat gctggggatg
     9601 cggtgggctc tatggcttct gaggcggaaa gaaccagctg gggctctagg gggtatcccc
     9661 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     9721 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     9781 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     9841 gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc
     9901 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
     9961 gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct tttgatttat
    10021 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
    10081 acgcgaatta attctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc
    10141 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc
    10201 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat
    10261 agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc
    10321 gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct ctgcctctga
    10381 gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctccc
    10441 gggagcttgt atatccattt tcggatctga tcagcacgtg ttgacaatta atcatcggca
    10501 tagtatatcg gcatagtata atacgacaag gtgaggaact aaaccatggc caagttgacc
    10561 agtgccgttc cggtgctcac cgcgcgcgac gtcgccggag cggtcgagtt ctggaccgac
    10621 cggctcgggt tctcccggga cttcgtggag gacgacttcg ccggtgtggt ccgggacgac
    10681 gtgaccctgt tcatcagcgc ggtccaggac caggtggtgc cggacaacac cctggcctgg
    10741 gtgtgggtgc gcggcctgga cgagctgtac gccgagtggt cggaggtcgt gtccacgaac
    10801 ttccgggacg cctccgggcc ggccatgacc gagatcggcg agcagccgtg ggggcgggag
    10861 ttcgccctgc gcgacccggc cggcaactgc gtgcacttcg tggccgagga gcaggactga
    10921 cacgtgctac gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc
    10981 gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc
    11041 gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca
    11101 aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc
    11161 aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg
    11221 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc
    11281 ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg
    11341 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
    11401 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
    11461 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
    11521 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
    11581 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
    11641 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
    11701 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
    11761 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
    11821 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
    11881 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
    11941 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
    12001 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
    12061 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
    12121 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
    12181 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
    12241 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
    12301 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
    12361 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
    12421 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
    12481 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
    12541 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
    12601 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
    12661 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
    12721 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
    12781 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
    12841 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
    12901 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
    12961 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
    13021 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
    13081 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
    13141 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
    13201 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
    13261 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
    13321 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgac
//
