LOCUS       sequence-151115-       13600 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fusion protein Cas9+GFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13600)
  AUTHORS   Pulido-Quetglas C, Aparicio-Prat E, Arnan C, Polidori T, Hermoso T, 
            Palumbo E, Ponomarenko J, Guigo R, Johnson R
  TITLE     Scalable Design of Paired CRISPR Guide RNAs for Genomic Deletion.
  JOURNAL   PLoS Comput Biol. 2017 Mar 2;13(3):e1005341. doi: 
            10.1371/journal.pcbi.1005341. eCollection 2017 Mar.
  PUBMED    28253259
REFERENCE   2  (bases 1 to 13600)
  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: "PLoS Comput
            Biol."; date: "2017-03-2"; volume: "13(3):e1005341. doi"; pages: " 
            10.1371/journal.pcbi.1005341. eCollection 2017 Mar"
COMMENT     Sequence Label: pLentiCas9-GFP
FEATURES             Location/Qualifiers
     source          1..13600
                     /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"
     promoter        2650..2861
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      2880..2889
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2886..6989
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             2886..6989
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             6990..7037
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             7038..7061
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             join(7068..7070,7071..7073,7074..7784)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKLSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLGTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(7374..7393)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     7721..7742
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             7785..7802
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             7812..7868
                     /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             7869..8267
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     misc_feature    8292..8880
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8345..8365)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8763..8774)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(8883..8899)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8884..8900)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             9405..9585
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(9611..9628)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    9617..9841
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      9887..10315
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9974..9993)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     10184..10205
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(10324..10344)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        10329..10658
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      10509..10644
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     10571..10590
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        10706..10753
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             10772..11146
                     /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    11276..11397
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(11313..11332)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     11367..11386
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(11446..11462)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(11446..11462)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(11459..11481)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    11470..11486
                     /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(11494..11500,11501..11518,11519..11524))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11539..11560
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(11677..11694)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(11848..12436)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(11928..11947)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(12607..13398,13399..13467))
                     /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     13230..13249
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(13468..13572)
                     /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 gctagctagg tcttgaaagg agtgggaatt ggctccggtg
     2641 cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg
     2701 gcaattgatc cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt
     2761 actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg
     2821 tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggaccggttc tagagcgctg
     2881 ccaccatgga caagaagtac agcatcggcc tggacatcgg caccaactct gtgggctggg
     2941 ccgtgatcac cgacgagtac aaggtgccca gcaagaaatt caaggtgctg ggcaacaccg
     3001 accggcacag catcaagaag aacctgatcg gagccctgct gttcgacagc ggcgaaacag
     3061 ccgaggccac ccggctgaag agaaccgcca gaagaagata caccagacgg aagaaccgga
     3121 tctgctatct gcaagagatc ttcagcaacg agatggccaa ggtggacgac agcttcttcc
     3181 acagactgga agagtccttc ctggtggaag aggataagaa gcacgagcgg caccccatct
     3241 tcggcaacat cgtggacgag gtggcctacc acgagaagta ccccaccatc taccacctga
     3301 gaaagaaact ggtggacagc accgacaagg ccgacctgcg gctgatctat ctggccctgg
     3361 cccacatgat caagttccgg ggccacttcc tgatcgaggg cgacctgaac cccgacaaca
     3421 gcgacgtgga caagctgttc atccagctgg tgcagaccta caaccagctg ttcgaggaaa
     3481 accccatcaa cgccagcggc gtggacgcca aggccatcct gtctgccaga ctgagcaaga
     3541 gcagacggct ggaaaatctg atcgcccagc tgcccggcga gaagaagaat ggcctgttcg
     3601 gaaacctgat tgccctgagc ctgggcctga cccccaactt caagagcaac ttcgacctgg
     3661 ccgaggatgc caaactgcag ctgagcaagg acacctacga cgacgacctg gacaacctgc
     3721 tggcccagat cggcgaccag tacgccgacc tgtttctggc cgccaagaac ctgtccgacg
     3781 ccatcctgct gagcgacatc ctgagagtga acaccgagat caccaaggcc cccctgagcg
     3841 cctctatgat caagagatac gacgagcacc accaggacct gaccctgctg aaagctctcg
     3901 tgcggcagca gctgcctgag aagtacaaag agattttctt cgaccagagc aagaacggct
     3961 acgccggcta cattgacggc ggagccagcc aggaagagtt ctacaagttc atcaagccca
     4021 tcctggaaaa gatggacggc accgaggaac tgctcgtgaa gctgaacaga gaggacctgc
     4081 tgcggaagca gcggaccttc gacaacggca gcatccccca ccagatccac ctgggagagc
     4141 tgcacgccat tctgcggcgg caggaagatt tttacccatt cctgaaggac aaccgggaaa
     4201 agatcgagaa gatcctgacc ttccgcatcc cctactacgt gggccctctg gccaggggaa
     4261 acagcagatt cgcctggatg accagaaaga gcgaggaaac catcaccccc tggaacttcg
     4321 aggaagtggt ggacaagggc gcttccgccc agagcttcat cgagcggatg accaacttcg
     4381 ataagaacct gcccaacgag aaggtgctgc ccaagcacag cctgctgtac gagtacttca
     4441 ccgtgtataa cgagctgacc aaagtgaaat acgtgaccga gggaatgaga aagcccgcct
     4501 tcctgagcgg cgagcagaaa aaggccatcg tggacctgct gttcaagacc aaccggaaag
     4561 tgaccgtgaa gcagctgaaa gaggactact tcaagaaaat cgagtgcttc gactccgtgg
     4621 aaatctccgg cgtggaagat cggttcaacg cctccctggg cacataccac gatctgctga
     4681 aaattatcaa ggacaaggac ttcctggaca atgaggaaaa cgaggacatt ctggaagata
     4741 tcgtgctgac cctgacactg tttgaggaca gagagatgat cgaggaacgg ctgaaaacct
     4801 atgcccacct gttcgacgac aaagtgatga agcagctgaa gcggcggaga tacaccggct
     4861 ggggcaggct gagccggaag ctgatcaacg gcatccggga caagcagtcc ggcaagacaa
     4921 tcctggattt cctgaagtcc gacggcttcg ccaacagaaa cttcatgcag ctgatccacg
     4981 acgacagcct gacctttaaa gaggacatcc agaaagccca ggtgtccggc cagggcgata
     5041 gcctgcacga gcacattgcc aatctggccg gcagccccgc cattaagaag ggcatcctgc
     5101 agacagtgaa ggtggtggac gagctcgtga aagtgatggg ccggcacaag cccgagaaca
     5161 tcgtgatcga aatggccaga gagaaccaga ccacccagaa gggacagaag aacagccgcg
     5221 agagaatgaa gcggatcgaa gagggcatca aagagctggg cagccagatc ctgaaagaac
     5281 accccgtgga aaacacccag ctgcagaacg agaagctgta cctgtactac ctgcagaatg
     5341 ggcgggatat gtacgtggac caggaactgg acatcaaccg gctgtccgac tacgatgtgg
     5401 accatatcgt gcctcagagc tttctgaagg acgactccat cgacaacaag gtgctgacca
     5461 gaagcgacaa gaaccggggc aagagcgaca acgtgccctc cgaagaggtc gtgaagaaga
     5521 tgaagaacta ctggcggcag ctgctgaacg ccaagctgat tacccagaga aagttcgaca
     5581 atctgaccaa ggccgagaga ggcggcctga gcgaactgga taaggccggc ttcatcaaga
     5641 gacagctggt ggaaacccgg cagatcacaa agcacgtggc acagatcctg gactcccgga
     5701 tgaacactaa gtacgacgag aatgacaagc tgatccggga agtgaaagtg atcaccctga
     5761 agtccaagct ggtgtccgat ttccggaagg atttccagtt ttacaaagtg cgcgagatca
     5821 acaactacca ccacgcccac gacgcctacc tgaacgccgt cgtgggaacc gccctgatca
     5881 aaaagtaccc taagctggaa agcgagttcg tgtacggcga ctacaaggtg tacgacgtgc
     5941 ggaagatgat cgccaagagc gagcaggaaa tcggcaaggc taccgccaag tacttcttct
     6001 acagcaacat catgaacttt ttcaagaccg agattaccct ggccaacggc gagatccgga
     6061 agcggcctct gatcgagaca aacggcgaaa ccggggagat cgtgtgggat aagggccggg
     6121 attttgccac cgtgcggaaa gtgctgagca tgccccaagt gaatatcgtg aaaaagaccg
     6181 aggtgcagac aggcggcttc agcaaagagt ctatcctgcc caagaggaac agcgataagc
     6241 tgatcgccag aaagaaggac tgggacccta agaagtacgg cggcttcgac agccccaccg
     6301 tggcctattc tgtgctggtg gtggccaaag tggaaaaggg caagtccaag aaactgaaga
     6361 gtgtgaaaga gctactgggg atcaccatca tggaaagaag cagcttcgag aagaatccca
     6421 tcgactttct ggaagccaag ggctacaaag aagtgaaaaa ggacctgatc atcaagctgc
     6481 ctaagtactc cctgttcgag ctggaaaacg gccggaagag aatgctggcc tctgccggcg
     6541 aactgcagaa gggaaacgaa ctggccctgc cctccaaata tgtgaacttc ctgtacctgg
     6601 ccagccacta tgagaagctg aagggctccc ccgaggataa tgagcagaaa cagctgtttg
     6661 tggaacagca caagcactac ctggacgaga tcatcgagca gatcagcgag ttctccaaga
     6721 gagtgatcct ggccgacgct aatctggaca aagtgctgtc cgcctacaac aagcaccggg
     6781 ataagcccat cagagagcag gccgagaata tcatccacct gtttaccctg accaatctgg
     6841 gagcccctgc cgccttcaag tactttgaca ccaccatcga ccggaagagg tacaccagca
     6901 ccaaagaggt gctggacgcc accctgatcc accagagcat caccggcctg tacgagacac
     6961 ggatcgacct gtctcagctg ggaggcgaca agcgacctgc cgccacaaag aaggctggac
     7021 aggctaagaa gaagaaagat tacaaagacg atgacgataa gggatccatg gtgagcaagg
     7081 gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc gacgtaaacg
     7141 gccacaagct cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc aagctgaccc
     7201 tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc
     7261 tgacctacgg cgtgcagtgc ttcagccgct accccgacca catgaagcag cacgacttct
     7321 tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc aaggacgacg
     7381 gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg aaccgcatcg
     7441 agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag ctggagtaca
     7501 actacaacag ccacaacgtc tatatcatgg ccgacaagca gaagaacggc atcaaggtga
     7561 acttcaagat ccgccacaac atcgaggacg gcagcgtgca gctcgccgac cactaccagc
     7621 agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac ctgggcaccc
     7681 agtccgccct gagcaaagac cccaacgaga agcgcgatca catggtcctg ctggagttcg
     7741 tgaccgccgc cgggatcact ctcggcatgg acgagctgta caagcatcat caccatcacc
     7801 acggatccgg cgcaacaaac ttctctctgc tgaaacaagc cggagatgtc gaagagaatc
     7861 ctggaccgat ggccaagcct ttgtctcaag aagaatccac cctcattgaa agagcaacgg
     7921 ctacaatcaa cagcatcccc atctctgaag actacagcgt cgccagcgca gctctctcta
     7981 gcgacggccg catcttcact ggtgtcaatg tatatcattt tactggggga ccttgtgcag
     8041 aactcgtggt gctgggcact gctgctgctg cggcagctgg caacctgact tgtatcgtcg
     8101 cgatcggaaa tgagaacagg ggcatcttga gcccctgcgg acggtgccga caggtgcttc
     8161 tcgatctgca tcctgggatc aaagccatag tgaaggacag tgatggacag ccgacggcag
     8221 ttgggattcg tgaattgctg ccctctggtt atgtgtggga gggctaagaa ttcgatatca
     8281 agcttatcgg taatcaacct ctggattaca aaatttgtga aagattgact ggtattctta
     8341 actatgttgc tccttttacg ctatgtggat acgctgcttt aatgcctttg tatcatgcta
     8401 ttgcttcccg tatggctttc attttctcct ccttgtataa atcctggttg ctgtctcttt
     8461 atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg
     8521 caacccccac tggttggggc attgccacca cctgtcagct cctttccggg actttcgctt
     8581 tccccctccc tattgccacg gcggaactca tcgccgcctg ccttgcccgc tgctggacag
     8641 gggctcggct gttgggcact gacaattccg tggtgttgtc ggggaaatca tcgtcctttc
     8701 cttggctgct cgcctgtgtt gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc
     8761 cttcggccct caatccagcg gaccttcctt cccgcggcct gctgccggct ctgcggcctc
     8821 ttccgcgtct tcgccttcgc cctcagacga gtcggatctc cctttgggcc gcctccccgc
     8881 atcgataccg tcgacctcga gacctagaaa aacatggagc aatcacaagt agcaatacag
     8941 cagctaccaa tgctgattgt gcctggctag aagcacaaga ggaggaggag gtgggttttc
     9001 cagtcacacc tcaggtacct ttaagaccaa tgacttacaa ggcagctgta gatcttagcc
     9061 actttttaaa agaaaagggg ggactggaag ggctaattca ctcccaacga agacaagata
     9121 tccttgatct gtggatctac cacacacaag gctacttccc tgattggcag aactacacac
     9181 cagggccagg gatcagatat ccactgacct ttggatggtg ctacaagcta gtaccagttg
     9241 agcaagagaa ggtagaagaa gccaatgaag gagagaacac ccgcttgtta caccctgtga
     9301 gcctgcatgg gatggatgac ccggagagag aagtattaga gtggaggttt gacagccgcc
     9361 tagcatttca tcacatggcc cgagagctgc atccggactg tactgggtct ctctggttag
     9421 accagatctg agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat
     9481 aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact
     9541 agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagggcc cgtttaaacc
     9601 cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc
     9661 gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa
     9721 attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac
     9781 agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt gggctctatg
     9841 gcttctgagg cggaaagaac cagctggggc tctagggggt atccccacgc gccctgtagc
     9901 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     9961 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
    10021 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
    10081 ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag
    10141 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
    10201 actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg
    10261 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattaattc
    10321 tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta
    10381 tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag
    10441 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa
    10501 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
    10561 taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta ttccagaagt
    10621 agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccggga gcttgtatat
    10681 ccattttcgg atctgatcag cacgtgttga caattaatca tcggcatagt atatcggcat
    10741 agtataatac gacaaggtga ggaactaaac catggccaag ttgaccagtg ccgttccggt
    10801 gctcaccgcg cgcgacgtcg ccggagcggt cgagttctgg accgaccggc tcgggttctc
    10861 ccgggacttc gtggaggacg acttcgccgg tgtggtccgg gacgacgtga ccctgttcat
    10921 cagcgcggtc caggaccagg tggtgccgga caacaccctg gcctgggtgt gggtgcgcgg
    10981 cctggacgag ctgtacgccg agtggtcgga ggtcgtgtcc acgaacttcc gggacgcctc
    11041 cgggccggcc atgaccgaga tcggcgagca gccgtggggg cgggagttcg ccctgcgcga
    11101 cccggccggc aactgcgtgc acttcgtggc cgaggagcag gactgacacg tgctacgaga
    11161 tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc
    11221 cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc accccaactt
    11281 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
    11341 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca
    11401 tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat agctgtttcc
    11461 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg
    11521 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc
    11581 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
    11641 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
    11701 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
    11761 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
    11821 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
    11881 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
    11941 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
    12001 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
    12061 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
    12121 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
    12181 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
    12241 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
    12301 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
    12361 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
    12421 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
    12481 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
    12541 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
    12601 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
    12661 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
    12721 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
    12781 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
    12841 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
    12901 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
    12961 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
    13021 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
    13081 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
    13141 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
    13201 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
    13261 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
    13321 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
    13381 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
    13441 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
    13501 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
    13561 aggggttccg cgcacatttc cccgaaaagt gccacctgac
//
