LOCUS       sequence-376609-       11246 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Lentiviral vector to co-express an spsgRNA with NLS-tagBFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11246)
  AUTHORS   Sekiguchi R, Mehlferber MM, Matsumoto K, Wang S
  TITLE     Efficient Gene Knockout in Salivary Gland Epithelial Explant 
            Cultures.
  JOURNAL   J Dent Res. 2023 Feb;102(2):197-206. doi: 10.1177/00220345221128201.
            Epub 2022 Nov 10.
  PUBMED    36366748
REFERENCE   2  (bases 1 to 11246)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1177/00220345221128201"; journalName: "J Dent Res"; date: 
            "2023-02"; volume: "102"; issue: "2"; pages: "197-206"
COMMENT     Sequence Label: 
            pW299-lenti-spsgRNA-Esp3I-2kb-filler-pEF1s-NLS-tagBFP-P2A-BlastR
FEATURES             Location/Qualifiers
     source          1..11246
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     153..173
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             216..396
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    443..568
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1061..1294
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1479..1523
                     /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    1821..1938
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        1989..2237
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1989..2009
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2160..2179
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        4119..4194
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        4256..4467
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     CDS             4495..4542
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             4543..5238
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="SELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKVV
                     EGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTAT
                     QDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALKL
                     VGGSHLIANIKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYCD
                     LPSKLGHKLN"
     CDS             5248..5304
                     /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             5305..5703
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     misc_feature    5719..6307
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5772..5792)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(6190..6201)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             6379..6612
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(6638..6655)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6644..6868
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      6914..7342
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7001..7020)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7211..7232
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(7351..7371)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        7356..7685
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      7536..7671
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     7598..7617
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        7733..7780
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             7799..8173
                     /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    8303..8424
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8340..8359)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8394..8413
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(8473..8489)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8473..8489)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(8486..8508)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    8497..8513
                     /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(8521..8527,8528..8545,8546..8551))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8566..8587
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(8704..8721)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(8875..9463)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(8955..8974)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(9634..10425,10426..10494))
                     /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     10257..10276
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(10495..10599)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(10674..10693)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        10865..11244
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        join(11246,1..198)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
ORIGIN
        1 gatgcggttt tggcagtaca tcaatgggcg tggatagcgg tttgactcac ggggatttcc
       61 aagtctccac cccattgacg tcaatgggag tttgttttgg caccaaaatc aacgggactt
      121 tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg
      181 ggaggtctat ataagcagcg cgttttgcct gtactgggtc tctctggtta gaccagatct
      241 gagcctggga gctctctggc taactaggga acccactgct taagcctcaa taaagcttgc
      301 cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc
      361 tcagaccctt ttagtcagtg tggaaaatct ctagcagtgg cgcccgaaca gggacttgaa
      421 agcgaaaggg aaaccagagg agctctctcg acgcaggact cggcttgctg aagcgcgcac
      481 ggcaagaggc gaggggcggc gactggtgag tacgccaaaa attttgacta gcggaggcta
      541 gaaggagaga gatgggtgcg agagcgtcag tattaagcgg gggagaatta gatcgcgatg
      601 ggaaaaaatt cggttaaggc cagggggaaa gaaaaaatat aaattaaaac atatagtatg
      661 ggcaagcagg gagctagaac gattcgcagt taatcctggc ctgttagaaa catcagaagg
      721 ctgtagacaa atactgggac agctacaacc atcccttcag acaggatcag aagaacttag
      781 atcattatat aatacagtag caaccctcta ttgtgtgcat caaaggatag agataaaaga
      841 caccaaggaa gctttagaca agatagagga agagcaaaac aaaagtaaga ccaccgcaca
      901 gcaagcggcc gctgatcttc agacctggag gaggagatat gagggacaat tggagaagtg
      961 aattatataa atataaagta gtaaaaattg aaccattagg agtagcaccc accaaggcaa
     1021 agagaagagt ggtgcagaga gaaaaaagag cagtgggaat aggagctttg ttccttgggt
     1081 tcttgggagc agcaggaagc actatgggcg cagcgtcaat gacgctgacg gtacaggcca
     1141 gacaattatt gtctggtata gtgcagcagc agaacaattt gctgagggct attgaggcgc
     1201 aacagcatct gttgcaactc acagtctggg gcatcaagca gctccaggca agaatcctgg
     1261 ctgtggaaag atacctaaag gatcaacagc tcctggggat ttggggttgc tctggaaaac
     1321 tcatttgcac cactgctgtg ccttggaatg ctagttggag taataaatct ctggaacaga
     1381 tttggaatca cacgacctgg atggagtggg acagagaaat taacaattac acaagcttaa
     1441 tacactcctt aattgaagaa tcgcaaaacc agcaagaaaa gaatgaacaa gaattattgg
     1501 aattagataa atgggcaagt ttgtggaatt ggtttaacat aacaaattgg ctgtggtata
     1561 taaaattatt cataatgata gtaggaggct tggtaggttt aagaatagtt tttgctgtac
     1621 tttctatagt gaatagagtt aggcagggat attcaccatt atcgtttcag acccacctcc
     1681 caaccccgag gggacccgac aggcccgaag gaatagaaga agaaggtgga gagagagaca
     1741 gagacagatc cattcgatta gtgaacggat cggcactgcg tgcgccaatt ctgcagacaa
     1801 atggcagtat tcatccacaa ttttaaaaga aaagggggga ttggggggta cagtgcaggg
     1861 gaaagaatag tagacataat agcaacagac atacaaacta aagaattaca aaaacaaatt
     1921 acaaaaattc aaaattttcg ggtttattac agggacagca gagatccagt ttggttaatt
     1981 aaggtaccga gggcctattt cccatgattc cttcatattt gcatatacga tacaaggctg
     2041 ttagagagat aattagaatt aatttgactg taaacacaaa gatattagta caaaatacgt
     2101 gacgtagaaa gtaataattt cttgggtagt ttgcagtttt aaaattatgt tttaaaatgg
     2161 actatcatat gcttaccgta acttgaaagt atttcgattt cttggcttta tatatcttgt
     2221 ggaaaggacg aaacaccgga gacggttgta aatgagcaca caaaatacac atgctaaaat
     2281 attatattct atgaccttta taaaatcaac caaaatcttc tttttaataa ctttagtatc
     2341 aataattaga atttttatgt tcctttttgc aaacttttaa taaaaatgag caaaataaaa
     2401 aaacgctagt tttagtaact cgcgttgttt tcttcacctt taataatagc tactccacca
     2461 cttgttccta agcggtcagc tcctgcttca atcatttttt gagcatcttc aaatgttcta
     2521 actccaccag ctgctttaac taaagcattg tctttaacaa ctgacttcat tagtttaaca
     2581 tcttcaaatg ttgcacctga ttttgaaaat cctgttgatg ttttaacaaa ttctaatcca
     2641 gcttcaacag ctatttcaca agctttcatg atttcttctt ttgttaataa acaattttcc
     2701 ataatacatt taacaacatg tgatccagct gcttttttta cagctttcat gtcttctaaa
     2761 actaattcat aatttttgtc ttttaatgca ccaatattta ataccatatc aatttctgtt
     2821 gcaccatctt taattgcttc agaaacttcg aatgcttttg tagctgttgt gcatgcacct
     2881 agaggaaaac ctacaacatt tgttattcct acatttgtgc cttttaataa ttctttacaa
     2941 tagcttgttc aatatgaatt aacacaaact gttgcaaaat caaattcaat tgcttcatca
     3001 cataattgtt taatttcagc tttcgtagca tcttgtttta ataatgtgtg atctatatat
     3061 ttgtttagtt tcattttttc tcctatatat tcatttttaa ttttaattct ttaataattt
     3121 cgtctacttt aactttagcg ttttgaacag attcaccaac acctataaaa taaattttta
     3181 gtttaggttc agttccactt gggcgaacag caaatcatga cttatcttct aaataaaatt
     3241 ttagtaagtc ttgtcctggc atattataca ttccatcgat gtagtcttca acattaacaa
     3301 ctttaagtcc agcaatttga gttaagggtg ttgctctcaa tgatttcatt aatggttcaa
     3361 tttttaattt cttttcttct ggtttaaaat tcaagtttaa agtgaaagtg taatatgcac
     3421 ccatttcttt aaataaatct tctaaatagt ctactaatgt tttattttgt tttttataaa
     3481 atcaagcagc ctctgctatt aatatagaag cttgtattcc atctttatct ctagctgagt
     3541 catcaattac atatccataa ctttcttcat aagcaaaaac aaaatttaat ccgttatctt
     3601 cttctttagc aatttctcta cccattcatt taaatccagt taaagttttt acaatattaa
     3661 ctccatattt ttcatgagcg attctatcac ccaaatcact tgttacaaaa cttgaatata
     3721 gagccggatt ttttggaatg ctatttaagc gttttagatt tgataatttt caatcaatta
     3781 aaattggtcc tgtttgattt ccatctaatc ttacaaaatg accatcatgt tttattgcca
     3841 ttccaaatct gtcagcatct gggtcattca taataataat atctgcatca tgtttaatac
     3901 catattcaag cggtattttt catgcaggat caaattctgg atttggattt acaacatttt
     3961 taaatgtttc atcttcaaat gcatgctctt caacctcaat aacgttatat cctgattcac
     4021 gtaatatttt tggggtaaat ttagttcctg ttccattaac tgcgctaaaa ataattttta
     4081 aatctttttt agcttcttgc tcttttttgt acgtctctgt tttagagcta gaaatagcaa
     4141 gttaaaataa ggctagtccg ttatcaactt gaaaaagtgg caccgagtcg gtgctttttt
     4201 gaattcgcta gctaggtctt gaaaggagtg ggaattggct ccggtgcccg tcagtgggca
     4261 gagcgcacat cgcccacagt ccccgagaag ttggggggag gggtcggcaa ttgatccggt
     4321 gcctagagaa ggtggcgcgg ggtaaactgg gaaagtgatg tcgtgtactg gctccgcctt
     4381 tttcccgagg gtgggggaga accgtatata agtgcagtag tcgccgtgaa cgttcttttt
     4441 cgcaacgggt ttgccgccag aacacaggac cggttctaga gcgctgccac catgaagcga
     4501 cctgccgcca caaagaaggc tggacaggct aagaagaaga aaagcgagct gattaaggag
     4561 aacatgcaca tgaagctgta catggagggc accgtggaca accatcactt caagtgcaca
     4621 tccgagggcg aaggcaagcc ctacgagggc acccagacca tgagaatcaa ggtggtcgag
     4681 ggcggccctc tccccttcgc cttcgacatc ctggctacta gcttcctcta cggcagcaag
     4741 accttcatca accacaccca gggcatcccc gacttcttca agcagtcctt ccctgagggc
     4801 ttcacatggg agagagtcac cacatacgaa gacgggggcg tgctgaccgc tacccaggac
     4861 accagcctcc aggacggctg cctcatctac aacgtcaaga tcagaggggt gaacttcaca
     4921 tccaacggcc ctgtgatgca gaagaaaaca ctcggctggg aggccttcac cgaaactctg
     4981 taccccgctg acggcggcct ggaaggcaga aacgacatgg ccctgaagct cgtgggcggg
     5041 agccatctga tcgcaaacat caagaccaca tatagatcca agaaacccgc taagaacctc
     5101 aagatgcctg gcgtctacta tgtggactac agactggaaa gaatcaagga ggccaacaac
     5161 gagacctacg tcgagcagca cgaggtggca gtggccagat actgcgacct ccctagcaaa
     5221 ctggggcaca agcttaatgg atccggcgca acaaacttct ctctgctgaa acaagccgga
     5281 gatgtcgaag agaatcctgg accgatggcc aagcctttgt ctcaagaaga atccaccctc
     5341 attgaaagag caacggctac aatcaacagc atccccatct ctgaagacta cagcgtcgcc
     5401 agcgcagctc tctctagcga cggccgcatc ttcactggtg tcaatgtata tcattttact
     5461 gggggacctt gtgcagaact cgtggtgctg ggcactgctg ctgctgcggc agctggcaac
     5521 ctgacttgta tcgtcgcgat cggaaatgag aacaggggca tcttgagccc ctgcggacgg
     5581 tgccgacagg tgcttctcga tctgcatcct gggatcaaag ccatagtgaa ggacagtgat
     5641 ggacagccga cggcagttgg gattcgtgaa ttgctgccct ctggttatgt gtgggagggc
     5701 taaacgcgtt aagtcgacaa tcaacctctg gattacaaaa tttgtgaaag attgactggt
     5761 attcttaact atgttgctcc ttttacgcta tgtggatacg ctgctttaat gcctttgtat
     5821 catgctattg cttcccgtat ggctttcatt ttctcctcct tgtataaatc ctggttgctg
     5881 tctctttatg aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt
     5941 gctgacgcaa cccccactgg ttggggcatt gccaccacct gtcagctcct ttccgggact
     6001 ttcgctttcc ccctccctat tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc
     6061 tggacagggg ctcggctgtt gggcactgac aattccgtgg tgttgtcggg gaaatcatcg
     6121 tcctttcctt ggctgctcgc ctgtgttgcc acctggattc tgcgcgggac gtccttctgc
     6181 tacgtccctt cggccctcaa tccagcggac cttccttccc gcggcctgct gccggctctg
     6241 cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc ggatctccct ttgggccgcc
     6301 tccccgcgtc gactttaaga ccaatgactt acaaggcagc tgtagatctt agccactttt
     6361 taaaagaaaa ggggggactg gaagggctaa ttcactccca acgaagacaa gatctgcttt
     6421 ttgcttgtac tgggtctctc tggttagacc agatctgagc ctgggagctc tctggctaac
     6481 tagggaaccc actgcttaag cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg
     6541 cccgtctgtt gtgtgactct ggtaactaga gatccctcag acccttttag tcagtgtgga
     6601 aaatctctag cagggcccgt ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc
     6661 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
     6721 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct
     6781 attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg
     6841 catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctct
     6901 agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
     6961 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct
     7021 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
     7081 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt
     7141 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
     7201 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat
     7261 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt
     7321 taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg tgtggaaagt
     7381 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     7441 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     7501 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt
     7561 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg
     7621 cctctgcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt
     7681 gcaaaaagct cccgggagct tgtatatcca ttttcggatc tgatcagcac gtgttgacaa
     7741 ttaatcatcg gcatagtata tcggcatagt ataatacgac aaggtgagga actaaaccat
     7801 ggccaagttg accagtgccg ttccggtgct caccgcgcgc gacgtcgccg gagcggtcga
     7861 gttctggacc gaccggctcg ggttctcccg ggacttcgtg gaggacgact tcgccggtgt
     7921 ggtccgggac gacgtgaccc tgttcatcag cgcggtccag gaccaggtgg tgccggacaa
     7981 caccctggcc tgggtgtggg tgcgcggcct ggacgagctg tacgccgagt ggtcggaggt
     8041 cgtgtccacg aacttccggg acgcctccgg gccggccatg accgagatcg gcgagcagcc
     8101 gtgggggcgg gagttcgccc tgcgcgaccc ggccggcaac tgcgtgcact tcgtggccga
     8161 ggagcaggac tgacacgtgc tacgagattt cgattccacc gccgccttct atgaaaggtt
     8221 gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat
     8281 gctggagttc ttcgcccacc ccaacttgtt tattgcagct tataatggtt acaaataaag
     8341 caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt
     8401 gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta gctagagctt
     8461 ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca
     8521 caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact
     8581 cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct
     8641 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc
     8701 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca
     8761 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
     8821 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     8881 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     8941 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     9001 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     9061 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     9121 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     9181 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     9241 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     9301 cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     9361 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     9421 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     9481 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     9541 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     9601 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     9661 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     9721 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
     9781 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     9841 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     9901 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
     9961 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
    10021 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
    10081 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
    10141 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
    10201 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
    10261 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
    10321 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
    10381 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
    10441 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
    10501 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
    10561 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
    10621 acctgacgtc gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc
    10681 tgctctgatg ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct
    10741 gagtagtgcg cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg
    10801 aagaatctgc ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg
    10861 cgttgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat
    10921 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg
    10981 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata
    11041 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta
    11101 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc
    11161 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac
    11221 gtattagtca tcgctattac catggt
//
