LOCUS       sequence-349890-       11590 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  lentiviral vector expressing Cas9 and a sgRNA targeting COX4.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11590)
  AUTHORS   Spinelli JB, Rosen PC, Sprenger HG, Puszynska AM, Mann JL, Roessler 
            JM, Cangelosi AL, Henne A, Condon KJ, Zhang T, Kunchok T, Lewis CA, 
            Chandel NS, Sabatini DM
  TITLE     Fumarate is a terminal electron acceptor in the mammalian electron 
            transport chain.
  JOURNAL   Science. 2021 Dec 3;374(6572):1227-1237. doi: 
            10.1126/science.abi7495. Epub 2021 Dec 2.
  PUBMED    34855504
REFERENCE   2  (bases 1 to 11590)
  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"; doi: 
            "10.1126/science.abi7495"; journalName: "Science"; date: "2021-12-3-
            3"; volume: "374"; issue: "6572"; pages: "1227-1237"
COMMENT     Sequence Label: pLentiCRISPRv1_COX4
FEATURES             Location/Qualifiers
     source          1..11590
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    85..106
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(121..126,127..144,145..151)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    159..175
                     /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     164..186
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     183..199
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     183..199
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     218..238
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        220..238
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        266..492
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             493..673
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    720..845
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1338..1571
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1756..1800
                     /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"
     promoter        1978..2226
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1978..1998
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2149..2168
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        2248..2323
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     misc_feature    2371..2488
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2557..2768
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      2788..2797
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2797..2820
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2827..2847
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2872..6972
                     /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             2872..6972
                     /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             6973..7020
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             7036..7092
                     /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             7093..7692
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(7093..7112)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7589..7609
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    7708..8296
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(7761..7781)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8179..8190)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             8368..8601
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    8679..8800
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8716..8735)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8770..8789
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      8840..8975
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(8995..9014)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(8996..9014)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(9024..9041)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(9024..9040)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(9033..9055)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      9182..9637
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9269..9288)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     9479..9500
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        9663..9767
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(9768..9836,9837..10628)
                     /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(9986..10005)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      10799..11387
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     11288..11307
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     11541..11558
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 tctccccgcg cgttggccga ttcattaatg cagctggcac gacaggtttc ccgactggaa
       61 agcgggcagt gagcgcaacg caattaatgt gagttagctc actcattagg caccccaggc
      121 tttacacttt atgcttccgg ctcgtatgtt gtgtggaatt gtgagcggat aacaatttca
      181 cacaggaaac agctatgacc atgattacgc caagcgcgca attaaccctc actaaaggga
      241 acaaaagctg gagctgcaag cttaatgtag tcttatgcaa tactcttgta gtcttgcaac
      301 atggtaacga tgagttagca acatgcctta caaggagaga aaaagcaccg tgcatgccga
      361 ttggtggaag taaggtggta cgatcgtgcc ttattaggaa ggcaacagac gggtctgaca
      421 tggattggac gaaccactga attgccgcat tgcagagata ttgtatttaa gtgcctagct
      481 cgatacataa acgggtctct ctggttagac cagatctgag cctgggagct ctctggctaa
      541 ctagggaacc cactgcttaa gcctcaataa agcttgcctt gagtgcttca agtagtgtgt
      601 gcccgtctgt tgtgtgactc tggtaactag agatccctca gaccctttta gtcagtgtgg
      661 aaaatctcta gcagtggcgc ccgaacaggg acttgaaagc gaaagggaaa ccagaggagc
      721 tctctcgacg caggactcgg cttgctgaag cgcgcacggc aagaggcgag gggcggcgac
      781 tggtgagtac gccaaaaatt ttgactagcg gaggctagaa ggagagagat gggtgcgaga
      841 gcgtcagtat taagcggggg agaattagat cgcgatggga aaaaattcgg ttaaggccag
      901 ggggaaagaa aaaatataaa ttaaaacata tagtatgggc aagcagggag ctagaacgat
      961 tcgcagttaa tcctggcctg ttagaaacat cagaaggctg tagacaaata ctgggacagc
     1021 tacaaccatc ccttcagaca ggatcagaag aacttagatc attatataat acagtagcaa
     1081 ccctctattg tgtgcatcaa aggatagaga taaaagacac caaggaagct ttagacaaga
     1141 tagaggaaga gcaaaacaaa agtaagacca ccgcacagca agcggccgct gatcttcaga
     1201 cctggaggag gagatatgag ggacaattgg agaagtgaat tatataaata taaagtagta
     1261 aaaattgaac cattaggagt agcacccacc aaggcaaaga gaagagtggt gcagagagaa
     1321 aaaagagcag tgggaatagg agctttgttc cttgggttct tgggagcagc aggaagcact
     1381 atgggcgcag cgtcaatgac gctgacggta caggccagac aattattgtc tggtatagtg
     1441 cagcagcaga acaatttgct gagggctatt gaggcgcaac agcatctgtt gcaactcaca
     1501 gtctggggca tcaagcagct ccaggcaaga atcctggctg tggaaagata cctaaaggat
     1561 caacagctcc tggggatttg gggttgctct ggaaaactca tttgcaccac tgctgtgcct
     1621 tggaatgcta gttggagtaa taaatctctg gaacagattt ggaatcacac gacctggatg
     1681 gagtgggaca gagaaattaa caattacaca agcttaatac actccttaat tgaagaatcg
     1741 caaaaccagc aagaaaagaa tgaacaagaa ttattggaat tagataaatg ggcaagtttg
     1801 tggaattggt ttaacataac aaattggctg tggtatataa aattattcat aatgatagta
     1861 ggaggcttgg taggtttaag aatagttttt gctgtacttt ctatagtgaa tagagttagg
     1921 cagggatatt caccattatc gtttcagacc cacctcccaa ccccgagggg acccagagag
     1981 ggcctatttc ccatgattcc ttcatatttg catatacgat acaaggctgt tagagagata
     2041 attagaatta atttgactgt aaacacaaag atattagtac aaaatacgtg acgtagaaag
     2101 taataatttc ttgggtagtt tgcagtttta aaattatgtt ttaaaatgga ctatcatatg
     2161 cttaccgtaa cttgaaagta tttcgatttc ttggctttat atatcttgtg gaaaggacga
     2221 aacaccggaa cttaatgcga tacactggtt ttagagctag aaatagcaag ttaaaataag
     2281 gctagtccgt tatcaacttg aaaaagtggc accgagtcgg tgcttttttg aattctagat
     2341 cttgagacaa atggcagtat tcatccacaa ttttaaaaga aaagggggga ttggggggta
     2401 cagtgcaggg gaaagaatag tagacataat agcaacagac atacaaacta aagaattaca
     2461 aaaacaaatt acaaaaattc aaaattttcg ggtttattac agggacagca gagatccact
     2521 ttggcgccgg ctcgagtggc tccggtgccc gtcagtgggc agagcgcaca tcgcccacag
     2581 tccccgagaa gttgggggga ggggtcggca attgaaccgg tgcctagaga aggtggcgcg
     2641 gggtaaactg ggaaagtgat gtcgtgtact ggctccgcct ttttcccgag ggtgggggag
     2701 aaccgtatat aagtgcagta gtcgccgtga acgttctttt tcgcaacggg tttgccgcca
     2761 gaacacaggt gtcgtgacgc gggatccgcc accatggatt acaaagacga tgacgataag
     2821 atggccccaa agaagaagcg gaaggtcggt atccacggag tcccagcagc cgacaagaag
     2881 tacagcatcg gcctggacat cggcaccaac tctgtgggct gggccgtgat caccgacgag
     2941 tacaaggtgc ccagcaagaa attcaaggtg ctgggcaaca ccgaccggca cagcatcaag
     3001 aagaacctga tcggagccct gctgttcgac agcggcgaaa cagccgaggc cacccggctg
     3061 aagagaaccg ccagaagaag atacaccaga cggaagaacc ggatctgcta tctgcaagag
     3121 atcttcagca acgagatggc caaggtggac gacagcttct tccacagact ggaagagtcc
     3181 ttcctggtgg aagaggataa gaagcacgag cggcacccca tcttcggcaa catcgtggac
     3241 gaggtggcct accacgagaa gtaccccacc atctaccacc tgagaaagaa actggtggac
     3301 agcaccgaca aggccgacct gcggctgatc tatctggccc tggcccacat gatcaagttc
     3361 cggggccact tcctgatcga gggcgacctg aaccccgaca acagcgacgt ggacaagctg
     3421 ttcatccagc tggtgcagac ctacaaccag ctgttcgagg aaaaccccat caacgccagc
     3481 ggcgtggacg ccaaggccat cctgtctgcc agactgagca agagcagacg gctggaaaat
     3541 ctgatcgccc agctgcccgg cgagaagaag aatggcctgt tcggaaacct gattgccctg
     3601 agcctgggcc tgacccccaa cttcaagagc aacttcgacc tggccgagga tgccaaactg
     3661 cagctgagca aggacaccta cgacgacgac ctggacaacc tgctggccca gatcggcgac
     3721 cagtacgccg acctgtttct ggccgccaag aacctgtccg acgccatcct gctgagcgac
     3781 atcctgagag tgaacaccga gatcaccaag gcccccctga gcgcctctat gatcaagaga
     3841 tacgacgagc accaccagga cctgaccctg ctgaaagctc tcgtgcggca gcagctgcct
     3901 gagaagtaca aagagatttt cttcgaccag agcaagaacg gctacgccgg ctacattgac
     3961 ggcggagcca gccaggaaga gttctacaag ttcatcaagc ccatcctgga aaagatggac
     4021 ggcaccgagg aactgctcgt gaagctgaac agagaggacc tgctgcggaa gcagcggacc
     4081 ttcgacaacg gcagcatccc ccaccagatc cacctgggag agctgcacgc cattctgcgg
     4141 cggcaggaag atttttaccc attcctgaag gacaaccggg aaaagatcga gaagatcctg
     4201 accttccgca tcccctacta cgtgggccct ctggccaggg gaaacagcag attcgcctgg
     4261 atgaccagaa agagcgagga aaccatcacc ccctggaact tcgaggaagt ggtggacaag
     4321 ggcgcttccg cccagagctt catcgagcgg atgaccaact tcgataagaa cctgcccaac
     4381 gagaaggtgc tgcccaagca cagcctgctg tacgagtact tcaccgtgta taacgagctg
     4441 accaaagtga aatacgtgac cgagggaatg agaaagcccg ccttcctgag cggcgagcag
     4501 aaaaaggcca tcgtggacct gctgttcaag accaaccgga aagtgaccgt gaagcagctg
     4561 aaagaggact acttcaagaa aatcgagtgc ttcgactccg tggaaatctc cggcgtggaa
     4621 gatcggttca acgcctccct gggcacatac cacgatctgc tgaaaattat caaggacaag
     4681 gacttcctgg acaatgagga aaacgaggac attctggaag atatcgtgct gaccctgaca
     4741 ctgtttgagg acagagagat gatcgaggaa cggctgaaaa cctatgccca cctgttcgac
     4801 gacaaagtga tgaagcagct gaagcggcgg agatacaccg gctggggcag gctgagccgg
     4861 aagctgatca acggcatccg ggacaagcag tccggcaaga caatcctgga tttcctgaag
     4921 tccgacggct tcgccaacag aaacttcatg cagctgatcc acgacgacag cctgaccttt
     4981 aaagaggaca tccagaaagc ccaggtgtcc ggccagggcg atagcctgca cgagcacatt
     5041 gccaatctgg ccggcagccc cgccattaag aagggcatcc tgcagacagt gaaggtggtg
     5101 gacgagctcg tgaaagtgat gggccggcac aagcccgaga acatcgtgat cgaaatggcc
     5161 agagagaacc agaccaccca gaagggacag aagaacagcc gcgagagaat gaagcggatc
     5221 gaagagggca tcaaagagct gggcagccag atcctgaaag aacaccccgt ggaaaacacc
     5281 cagctgcaga acgagaagct gtacctgtac tacctgcaga atgggcggga tatgtacgtg
     5341 gaccaggaac tggacatcaa ccggctgtcc gactacgatg tggaccatat cgtgcctcag
     5401 agctttctga aggacgactc catcgacaac aaggtgctga ccagaagcga caagaaccgg
     5461 ggcaagagcg acaacgtgcc ctccgaagag gtcgtgaaga agatgaagaa ctactggcgg
     5521 cagctgctga acgccaagct gattacccag agaaagttcg acaatctgac caaggccgag
     5581 agaggcggcc tgagcgaact ggataaggcc ggcttcatca agagacagct ggtggaaacc
     5641 cggcagatca caaagcacgt ggcacagatc ctggactccc ggatgaacac taagtacgac
     5701 gagaatgaca agctgatccg ggaagtgaaa gtgatcaccc tgaagtccaa gctggtgtcc
     5761 gatttccgga aggatttcca gttttacaaa gtgcgcgaga tcaacaacta ccaccacgcc
     5821 cacgacgcct acctgaacgc cgtcgtggga accgccctga tcaaaaagta ccctaagctg
     5881 gaaagcgagt tcgtgtacgg cgactacaag gtgtacgacg tgcggaagat gatcgccaag
     5941 agcgagcagg aaatcggcaa ggctaccgcc aagtacttct tctacagcaa catcatgaac
     6001 tttttcaaga ccgagattac cctggccaac ggcgagatcc ggaagcggcc tctgatcgag
     6061 acaaacggcg aaaccgggga gatcgtgtgg gataagggcc gggattttgc caccgtgcgg
     6121 aaagtgctga gcatgcccca agtgaatatc gtgaaaaaga ccgaggtgca gacaggcggc
     6181 ttcagcaaag agtctatcct gcccaagagg aacagcgata agctgatcgc cagaaagaag
     6241 gactgggacc ctaagaagta cggcggcttc gacagcccca ccgtggccta ttctgtgctg
     6301 gtggtggcca aagtggaaaa gggcaagtcc aagaaactga agagtgtgaa agagctgctg
     6361 gggatcacca tcatggaaag aagcagcttc gagaagaatc ccatcgactt tctggaagcc
     6421 aagggctaca aagaagtgaa aaaggacctg atcatcaagc tgcctaagta ctccctgttc
     6481 gagctggaaa acggccggaa gagaatgctg gcctctgccg gcgaactgca gaagggaaac
     6541 gaactggccc tgccctccaa atatgtgaac ttcctgtacc tggccagcca ctatgagaag
     6601 ctgaagggct cccccgagga taatgagcag aaacagctgt ttgtggaaca gcacaagcac
     6661 tacctggacg agatcatcga gcagatcagc gagttctcca agagagtgat cctggccgac
     6721 gctaatctgg acaaagtgct gtccgcctac aacaagcacc gggataagcc catcagagag
     6781 caggccgaga atatcatcca cctgtttacc ctgaccaatc tgggagcccc tgccgccttc
     6841 aagtactttg acaccaccat cgaccggaag aggtacacca gcaccaaaga ggtgctggac
     6901 gccaccctga tccaccagag catcaccggc ctgtacgaga cacggatcga cctgtctcag
     6961 ctgggaggcg acaagcgtcc tgctgctact aagaaagctg gtcaagctaa gaaaaagaaa
     7021 gctagcggca gcggcgccac caacttcagc ctgctgaagc aggccggcga cgtggaggag
     7081 aaccccggcc ccatgaccga gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc
     7141 cccagggccg tacgcaccct cgccgccgcg ttcgccgact accccgccac gcgccacacc
     7201 gtcgatccgg accgccacat cgagcgggtc accgagctgc aagaactctt cctcacgcgc
     7261 gtcgggctcg acatcggcaa ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg
     7321 accacgccgg agagcgtcga agcgggggcg gtgttcgccg agatcggccc gcgcatggcc
     7381 gagttgagcg gttcccggct ggccgcgcag caacagatgg aaggcctcct ggcgccgcac
     7441 cggcccaagg agcccgcgtg gttcctggcc accgtcggag tctcgcccga ccaccagggc
     7501 aagggtctgg gcagcgccgt cgtgctcccc ggagtggagg cggccgagcg cgccggggtg
     7561 cccgccttcc tggagacctc cgcgccccgc aacctcccct tctacgagcg gctcggcttc
     7621 accgtcaccg ccgacgtcga ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag
     7681 cccggtgcct gaacgcgtta agtcgacaat caacctctgg attacaaaat ttgtgaaaga
     7741 ttgactggta ttcttaacta tgttgctcct tttacgctat gtggatacgc tgctttaatg
     7801 cctttgtatc atgctattgc ttcccgtatg gctttcattt tctcctcctt gtataaatcc
     7861 tggttgctgt ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc
     7921 actgtgtttg ctgacgcaac ccccactggt tggggcattg ccaccacctg tcagctcctt
     7981 tccgggactt tcgctttccc cctccctatt gccacggcgg aactcatcgc cgcctgcctt
     8041 gcccgctgct ggacaggggc tcggctgttg ggcactgaca attccgtggt gttgtcgggg
     8101 aaatcatcgt cctttccttg gctgctcgcc tgtgttgcca cctggattct gcgcgggacg
     8161 tccttctgct acgtcccttc ggccctcaat ccagcggacc ttccttcccg cggcctgctg
     8221 ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg gatctccctt
     8281 tgggccgcct ccccgcgtcg actttaagac caatgactta caaggcagct gtagatctta
     8341 gccacttttt aaaagaaaag gggggactgg aagggctaat tcactcccaa cgaagacaag
     8401 atctgctttt tgcttgtact gggtctctct ggttagacca gatctgagcc tgggagctct
     8461 ctggctaact agggaaccca ctgcttaagc ctcaataaag cttgccttga gtgcttcaag
     8521 tagtgtgtgc ccgtctgttg tgtgactctg gtaactagag atccctcaga cccttttagt
     8581 cagtgtggaa aatctctagc agtacgtata gtagttcatg tcatcttatt attcagtatt
     8641 tataacttgc aaagaaatga atatcagaga gtgagaggaa cttgtttatt gcagcttata
     8701 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     8761 attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgg ctctagctat
     8821 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     8881 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg cctctgagct
     8941 attccagaag tagtgaggag gcttttttgg aggcctaggg acgtacccaa ttcgccctat
     9001 agtgagtcgt attacgcgcg ctcactggcc gtcgttttac aacgtcgtga ctgggaaaac
     9061 cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag ctggcgtaat
     9121 agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg
     9181 gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
     9241 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
     9301 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
     9361 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
     9421 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
     9481 ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta
     9541 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt
     9601 aacgcgaatt ttaacaaaat attaacgctt acaatttagg tggcactttt cggggaaatg
     9661 tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga
     9721 gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac
     9781 atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc
     9841 cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca
     9901 tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc
     9961 caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg
    10021 ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac
    10081 cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca
    10141 taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg
    10201 agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac
    10261 cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg
    10321 caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat
    10381 taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg
    10441 ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg
    10501 cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc
    10561 aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc
    10621 attggtaact gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt
    10681 tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt
    10741 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
    10801 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
    10861 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
    10921 gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca
    10981 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
    11041 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
    11101 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
    11161 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
    11221 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
    11281 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
    11341 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
    11401 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt
    11461 tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc
    11521 gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agcggaagag cgcccaatac
    11581 gcaaaccgcc
//
