LOCUS       sequence-476313-        9930 bp    DNA     circular SYN 18-JUN-2025
DEFINITION  Bxb1-GT donor plasmid with divergent syntax for all-in-one 
            doxycycline-inducible expression (empty).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9930)
  AUTHORS   Blanch-Asensio A, Ploessl DS, Wang NB, Mummery CL, Galloway KE, 
            Davis RP
  TITLE     STRAIGHT-IN Dual: a platform for dual, single-copy integrations of 
            DNA payloads and gene circuits into human induced pluripotent stem 
            cells.
  JOURNAL   bioRxiv [Preprint]. 2024 Oct 17:2024.10.17.616637. doi: 
            10.1101/2024.10.17.616637.
  PUBMED    39464154
REFERENCE   2  (bases 1 to 9930)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jun 18, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv 
            [Preprint]. 2024 Oct 17:2024.10.17.616637. doi: 
            10.1101/2024.10.17.616637."
COMMENT     Sequence Label: Bxb1-GT_AIO-TetOn_Divergent Donor
FEATURES             Location/Qualifiers
     source          1..9930
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    877..898
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(913..918,919..936,937..943)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    951..967
                     /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     975..991
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        1023..1233
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        1045..1241
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      1092..1227
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        1328..2506
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          1559..2497
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     2454..2474
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      2514..2523
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    2526..2563
                     /label=Bxb1 attB
                     /bound_moiety="Mycobacteriophage Bxb1 integrase"
                     /note="Minimal attB site for the Mycopbacteriophage Bxb1 
                     integrase (Voutev and Mann, 2017)"
     protein_bind    complement(2572..2605)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    2694..3282
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             2800..2811
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     3209..3229
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3348..3671)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLNR
                     LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTICS
                     DAA"
     primer_bind     3581..3597
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    3598..3654
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(3667..3683)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    3691..3707
                     /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(3715..3721,3722..3739,3740..3745))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3760..3781
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     regulatory      3785..3794
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      3789..3798
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(3877..3901)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     protein_bind    3946..3964
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3982..4000
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4018..4036
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4054..4072
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4090..4108
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4126..4144
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4162..4180
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     enhancer        4259..4562
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4564..4839
                     /label=chicken beta-actin promoter
     primer_bind     5732..5754
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     5816..5835
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             5867..6610
                     /codon_start=1
                     /product="modified rtTA protein that binds tightly to 
                     promoters containing the tet operator in the presence of 
                     doxycycline"
                     /label=Tet-On(R) 3G
                     /translation="MSRLDKSKVINSALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHSCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLKQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA
                     DALDDFDLDMLPADALDDFDLDMLPG"
     CDS             6611..6664
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             6665..7372
                     /codon_start=1
                     /product="enhanced monomeric blue fluorescent protein 
                     (Subach et al., 2011)"
                     /label=mTagBFP2
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMR
                     IKVVEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGV
                     LTATQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDM
                     ALKLVGGSHLIANAKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVA
                     RYCDLPSKLGHKLN"
     CDS             7397..7417
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7421..7441
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7445..7465
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(7530..7547)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    7536..7760
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    complement(7789..7821)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     primer_bind     complement(7829..7845)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      8058..8513
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(8145..8164)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     8355..8376
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     8649..8671
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(8709..8727)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        8795..8899
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(8900..8968,8969..9760)
                     /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(9118..9137)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
      781 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
      841 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
      901 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
      961 ataacaattt cacacaggaa acagctatga ccatgattac gccaagctta attccctccc
     1021 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc
     1081 taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct
     1141 gactaatttt ttttatttat gcagaggccg aggccgcctc ggcctctgag ctattccaga
     1201 agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctttgc aaagatggat
     1261 aaagttttaa acagagagga atctttgcag ctaatggacc ttctaggtct tgaaaggagt
     1321 gggaattggc tccggtgccc gtcagtgggc agagcgcaca tcgcccacag tccccgagaa
     1381 gttgggggga ggggtcggca attgaaccgg tgcctagaga aggtggcgcg gggtaaactg
     1441 ggaaagtgat gtcgtgtact ggctccgcct ttttcccgag ggtgggggag aaccgtatat
     1501 aagtgcagta gtcgccgtga acgttctttt tcgcaacggg tttgccgcca gaacacaggt
     1561 aagtgccgtg tgtggttccc gcgggcctgg cctctttacg ggttatggcc cttgcgtgcc
     1621 ttgaattact tccacctggc tgcagtacgt gattcttgat cccgagcttc gggttggaag
     1681 tgggtgggag agttcgaggc cttgcgctta aggagcccct tcgcctcgtg cttgagttga
     1741 ggcctggcct gggcgctggg gccgccgcgt gcgaatctgg tggcaccttc gcgcctgtct
     1801 cgctgctttc gataagtctc tagccattta aaatttttga tgacctgctg cgacgctttt
     1861 tttctggcaa gatagtcttg taaatgcggg ccaagatctg cacactggta tttcggtttt
     1921 tggggccgcg ggcggcgacg gggcccgtgc gtcccagcgc acatgttcgg cgaggcgggg
     1981 cctgcgagcg cggccaccga gaatcggacg ggggtagtct caagctggcc ggcctgctct
     2041 ggtgcctggc ctcgcgccgc cgtgtatcgc cccgccctgg gcggcaaggc tggcccggtc
     2101 ggcaccagtt gcgtgagcgg aaagatggcc gcttcccggc cctgctgcag ggagctcaaa
     2161 atggaggacg cggcgctcgg gagagcgggc gggtgagtca cccacacaaa ggaaaagggc
     2221 ctttccgtcc tcagccgtcg cttcatgtga ctccacggag taccgggcgc cgtccaggca
     2281 cctcgattag ttctcgagct tttggagtac gtcgtcttta ggttgggggg aggggtttta
     2341 tgcgatggag tttccccaca ctgagtgggt ggagactgaa gttaggccag cttggcactt
     2401 gatgtaattc tccttggaat ttgccctttt tgagtttgga tcttggttca ttctcaagcc
     2461 tcagacagtg gttcaaagtt tttttcttcc atttcaggtg tcgtgaggaa ttcgccacca
     2521 tggccggctt gtcgacgacg gcggtctccg tcgtcaggat catccggatc cataacttcg
     2581 tatagcatac attatacgaa gttatcatga tattcggcaa gcaggcatcg aaatgtcgag
     2641 acccatagag cccaccgcat ccccagcact agcgcgggga ggcggcccaa agggcgggga
     2701 ggcggcccaa agggagatcc gactcgtctg agggcgaagg cgaagacgcg gaagaggccg
     2761 cagagccggc agcaggccgc gggaaggaag gtccgctgga ttgagggccg aagggacgta
     2821 gcagaaggac gtcccgcgca gaatccaggt ggcaacacag gcgagcagcc atggaaagga
     2881 cgtcagcttc cccgacaaca ccacggaatt gtcagtgccc aacagccgag cccctgtcca
     2941 gcagcgggca aggcaggcgg cgatgagttc cgccgtggca atagggaggg ggaaagcgaa
     3001 agtcccggaa aggagctgac aggtggtggc aatgccccaa ccagtggggg ttgcgtcagc
     3061 aaacacagtg cacaccacgc cacgttgcct gacaacgggc cacaactcct cataaagaga
     3121 cagcaaccag gatttataca aggaggagaa aatgaaagcc atacgggaag caatagcatg
     3181 atacaaaggc attaaagcag cgtatccaca tagcgtaaaa ggagcaacat agttaagaat
     3241 accagtcaat ctttcacaaa ttttgtaatc cagaggttga ttgcggccgc ttccgcgtta
     3301 taggcttaat ttgttcaaaa aaaagcccgc tcattaggcg ggctgggcta tgcggcatca
     3361 gagcagattg tactgagagt gcaccatatg cggtgtgaaa taccgcacag atgcgtaagg
     3421 agaaaatacc gcatcaggcg ccattcgcca ttcaggctgc gcaactgttg ggaagggcga
     3481 tcggtgcggg cctcttcgct attacgccag ctggcgaaag ggggatgtgc tgcaaggcga
     3541 ttaagttggg taacgccagg gttttcccag tcacgacgtt gtaaaacgac ggccagtgaa
     3601 ttcgagctcg gtacccgggg atcctctaga gtcgacctgc aggcatgcaa gcttggcgta
     3661 atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     3721 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     3781 agcggccgcc atggtggcga attcttaagt ttacgagggt aggaagtggt acggaaagtt
     3841 ggtataagac aaaagtgttg tggaattgct ccaggcgatc tgacggttca ctaaacgagc
     3901 tctgctttta taggcgccca ccgtacacgc ctaaagctta tacgttctct atcactgata
     3961 gggagtaaac tggatatacg ttctctatca ctgataggga gtaaactgta gatacgttct
     4021 ctatcactga tagggagtaa actggtcata cgttctctat cactgatagg gagtaaactc
     4081 cttatacgtt ctctatcact gatagggagt aaagtctgca tacgttctct atcactgata
     4141 gggagtaaac tcttcatacg ttctctatca ctgataggga gtaaacctta attaactagt
     4201 cttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg
     4261 ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga
     4321 cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat
     4381 gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa
     4441 gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca
     4501 tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc gctattacca
     4561 tggtcgaggt gagccccacg ttctgcttca ctctccccat ctcccccccc tccccacccc
     4621 caattttgta tttatttatt ttttaattat tttgtgcagc gatgggggcg gggggggggg
     4681 gggcgcgcgc caggcggggc ggggcggggc gaggggcggg gcggggcgag gcggagaggt
     4741 gcggcggcag ccaatcagag cggcgcgctc cgaaagtttc cttttatggc gaggcggcgg
     4801 cggcggcggc cctataaaaa gcgaagcgcg cggcgggcgg gagtcgctgc gcgctgcctt
     4861 cgccccgtgc cccgctccgc cgccgcctcg cgccgcccgc cccggctctg actgaccgcg
     4921 ttactcccac aggtgagcgg gcgggacggc ccttctcctc cgggctgtaa ttagcgcttg
     4981 gtttaatgac ggcttgtttc ttttctgtgg ctgcgtgaaa gccttgaggg gctccgggag
     5041 ggccctttgt gcggggggag cggctcgggg ggtgcgtgcg tgtgtgtgtg cgtggggagc
     5101 gccgcgtgcg gctccgcgct gcccggcggc tgtgagcgct gcgggcgcgg cgcggggctt
     5161 tgtgcgctcc gcagtgtgcg cgaggggagc gcggccgggg gcggtgcccc gcggtgcggg
     5221 gggggctgcg aggggaacaa aggctgcgtg cggggtgtgt gcgtgggggg gtgagcaggg
     5281 ggtgtgggcg cgtcggtcgg gctgcaaccc cccctgcacc cccctccccg agttgctgag
     5341 cgcggcccgg cttcgggtgc ggggctccgt acggggcgtg gcgcggggct cgccgtgccg
     5401 ggcggggggt ggcggcaggt gggggtgccg ggcggccccc ggagcgccgg cggctgtcga
     5461 ggcgcggcga gccgcagcca ttgcctttta tggtaatcgt gcgagagggc gcagggactt
     5521 cctttgtccc aaatctgtgc ggagccgaaa tctgggaggc gccgccgcac cccctctagc
     5581 gggcgcgggg cgaagcggtg cggcgccggc aggaaggaaa tgggcgggga gggccttcgt
     5641 gcgtcgccgc gccgccgtcc ccttctccct ctccagcctc ggggctgtcc gcggggggac
     5701 ggctgccttc gggggggacg gggcagggcg gggttcggct tctggcgtgt gaccggcggc
     5761 tctagagcct ctgctaacca tgttcatgcc ttcttctttt tcctacagct cctgggcaac
     5821 gtgctggtta ttgtgctgtc tcatcatttt ggcaaagaat gtcgacatga gccggctgga
     5881 caagagcaaa gtgatcaaca gcgccctgga actgctgaac ggcgtgggca tcgagggcct
     5941 gaccacccgg aagctggccc agaaactggg cgtggaacag cccaccctgt actggcacgt
     6001 gaagaacaag cgggccctgc tggacgccct gcccatcgag atgctggacc ggcaccacac
     6061 ccacagctgc cctctggaag gcgagagctg gcaggacttc ctgcggaaca acgccaagag
     6121 ctacagatgc gccctgctga gccaccggga cggcgccaaa gtgcacctgg gcaccagacc
     6181 caccgagaag cagtacgaga cactggaaaa ccagctggcc ttcctgtgcc agcagggctt
     6241 cagcctggaa aacgccctgt acgccctgag cgccgtgggc cacttcaccc tgggctgcgt
     6301 gctggaagaa caggaacacc aggtcgccaa agaggaaaga gagacaccca ccaccgacag
     6361 catgcccccc ctgctgaagc aggccatcga gctgttcgac agacagggcg ccgagcccgc
     6421 cttcctgttc ggcctggaac tgatcatctg cggcctcgag aagcagctga agtgcgagag
     6481 cggcggaccc accgacgccc tggacgactt cgacctggac atgctgcccg ccgatgccct
     6541 ggatgatttt gatctggata tgctccctgc tgacgctctc gatgacttcg atctcgatat
     6601 gctgccaggc gagggcaggg gaagtcttct aacatgcggg gacgtggagg aaaatcccgg
     6661 ccccgtgtct aagggcgaag agctgattaa ggagaacatg cacatgaagc tgtacatgga
     6721 gggcaccgtg gacaaccatc acttcaagtg cacatccgag ggcgaaggca agccctacga
     6781 gggcacccag accatgagaa tcaaggtggt cgagggcggc cctctcccct tcgccttcga
     6841 catcctggct actagcttcc tctacggcag caagaccttc atcaaccaca cccagggcat
     6901 ccccgacttc ttcaagcagt ccttccctga gggcttcaca tgggagagag tcaccacata
     6961 cgaagacggg ggcgtgctga ccgctaccca ggacaccagc ctccaggacg gctgcctcat
     7021 ctacaacgtc aagatcagag gggtgaactt cacatccaac ggccctgtga tgcagaagaa
     7081 aacactcggc tgggaggcct tcaccgagac gctgtacccc gctgacggcg gcctggaagg
     7141 cagaaacgac atggccctga agctcgtggg cgggagccat ctgatcgcaa acgccaagac
     7201 cacatataga tccaagaaac ccgctaagaa cctcaagatg cctggcgtct actatgtgga
     7261 ctacagactg gaaagaatca aggaggccaa caacgagacc tacgtcgagc agcacgaggt
     7321 ggcagtggcc agatactgcg acctccctag caaactgggg cacaagctta attccggact
     7381 cagatctcga gctgatccaa aaaagaagag aaaggtagat ccaaaaaaga agagaaaggt
     7441 agatccaaaa aagaagagaa aggtaggatc caccggatct agataactga tcataatcag
     7501 ccataccaca tttgaaaccc gctgatcagc ctcgactgtg ccttctagtt gccagccatc
     7561 tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct
     7621 ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg
     7681 gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg
     7741 ggatgcggtg ggctctatgg ctagccgctg caagaactct tcctcacgat aacttcgtat
     7801 aggagacttt atacgaagtt aagcgctcac tggccgtcgt tttacaacgt cgtgactggg
     7861 aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc
     7921 gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg
     7981 aatggcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatac
     8041 gtcaaagcaa ccatagtacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt
     8101 tacgcgcagc gtgaccgcta cacttgccag cgccttagcg cccgctcctt tcgctttctt
     8161 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc
     8221 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg atttgggtga
     8281 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc
     8341 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaact ctatctcggg
     8401 ctattctttt gatttataag ggattttgcc gatttcggtc tattggttaa aaaatgagct
     8461 gatttaacaa aaatttaacg cgaattttaa caaaatatta acgtttacaa ttttatggtg
     8521 cactctcagt acaatctgct ctgatgccgc atagttaagc cagccccgac acccgccaac
     8581 acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt
     8641 gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag
     8701 acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc
     8761 ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     8821 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
     8881 atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt
     8941 tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc
     9001 tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat
     9061 ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct
     9121 atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca
     9181 ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg
     9241 catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa
     9301 cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg
     9361 ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga
     9421 cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg
     9481 cgaactactt actctagctt cccggcaaca attaatagac tggatggagg cggataaagt
     9541 tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg
     9601 agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc
     9661 ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca
     9721 gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc
     9781 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     9841 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     9901 agaccccgta gaaaagatca aaggatcttc
//
