LOCUS       sequence-462043-       10009 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bxb1-GA donor plasmid with downstream tandem 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 10009)
  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 10009)
  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: "bioRxiv 
            [Preprint]. 2024 Oct 17:2024.10.17.616637. doi: 
            10.1101/2024.10.17.616637."
COMMENT     Sequence Label: Bxb1-GA_AIO-TetOn_Down-Tandem Donor
FEATURES             Location/Qualifiers
     source          1..10009
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    107..128
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(143..148,149..166,167..173)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    181..197
                     /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     205..221
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        253..463
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        275..471
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      322..457
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        558..1736
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          789..1727
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1684..1704
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      1744..1753
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    1756..1793
                     /label=Bxb1 attB
                     /bound_moiety="Mycobacteriophage Bxb1 integrase"
                     /note="Minimal attB site for the Mycopbacteriophage Bxb1 
                     integrase (Voutev and Mann, 2017)"
     protein_bind    1808..1855
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     protein_bind    1900..1918
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1936..1954
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1972..1990
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2008..2026
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2044..2062
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2080..2098
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2116..2134
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     2179..2203
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      2282..2291
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      2286..2295
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    2299..2320
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2335..2340,2341..2358,2359..2365)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2373..2389
                     /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     2397..2413
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             2409..2732
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLNR
                     LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTICS
                     DAA"
     misc_feature    2426..2482
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(2483..2499)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    2798..3386
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2851..2871)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3269..3280)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     enhancer        3443..3822
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     4991..5013
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     5075..5094
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             5126..5869
                     /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             5870..5923
                     /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             5924..6631
                     /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             6656..6676
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             6680..6700
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             6704..6724
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(6789..6806)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6795..7019
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    7067..7100
                     /label=FRT3
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision with other FRT3 sites
                     (Bosch et al., 2015)"
     primer_bind     complement(7108..7124)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7138..7154)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      7367..7822
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7454..7473)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7664..7685
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7958..7980
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(8018..8036)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        8104..8208
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(8209..8277,8278..9069)
                     /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(8427..8446)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      9240..9828
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     9729..9748
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     9982..9999
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
       61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
      121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
      181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagctta
      241 attccctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata
      301 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
      361 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc ggcctctgag
      421 ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctttgc
      481 aaagatggat aaagttttaa acagagagga atctttgcag ctaatggacc ttctaggtct
      541 tgaaaggagt gggaattggc tccggtgccc gtcagtgggc agagcgcaca tcgcccacag
      601 tccccgagaa gttgggggga ggggtcggca attgaaccgg tgcctagaga aggtggcgcg
      661 gggtaaactg ggaaagtgat gtcgtgtact ggctccgcct ttttcccgag ggtgggggag
      721 aaccgtatat aagtgcagta gtcgccgtga acgttctttt tcgcaacggg tttgccgcca
      781 gaacacaggt aagtgccgtg tgtggttccc gcgggcctgg cctctttacg ggttatggcc
      841 cttgcgtgcc ttgaattact tccacctggc tgcagtacgt gattcttgat cccgagcttc
      901 gggttggaag tgggtgggag agttcgaggc cttgcgctta aggagcccct tcgcctcgtg
      961 cttgagttga ggcctggcct gggcgctggg gccgccgcgt gcgaatctgg tggcaccttc
     1021 gcgcctgtct cgctgctttc gataagtctc tagccattta aaatttttga tgacctgctg
     1081 cgacgctttt tttctggcaa gatagtcttg taaatgcggg ccaagatctg cacactggta
     1141 tttcggtttt tggggccgcg ggcggcgacg gggcccgtgc gtcccagcgc acatgttcgg
     1201 cgaggcgggg cctgcgagcg cggccaccga gaatcggacg ggggtagtct caagctggcc
     1261 ggcctgctct ggtgcctggc ctcgcgccgc cgtgtatcgc cccgccctgg gcggcaaggc
     1321 tggcccggtc ggcaccagtt gcgtgagcgg aaagatggcc gcttcccggc cctgctgcag
     1381 ggagctcaaa atggaggacg cggcgctcgg gagagcgggc gggtgagtca cccacacaaa
     1441 ggaaaagggc ctttccgtcc tcagccgtcg cttcatgtga ctccacggag taccgggcgc
     1501 cgtccaggca cctcgattag ttctcgagct tttggagtac gtcgtcttta ggttgggggg
     1561 aggggtttta tgcgatggag tttccccaca ctgagtgggt ggagactgaa gttaggccag
     1621 cttggcactt gatgtaattc tccttggaat ttgccctttt tgagtttgga tcttggttca
     1681 ttctcaagcc tcagacagtg gttcaaagtt tttttcttcc atttcaggtg tcgtgaggaa
     1741 ttcgccacca tggccggctt gtcgacgacg gcggactccg tcgtcaggat catccggatc
     1801 cggatccgaa gttcctattc cgaagttcct attctctaga aagtatagga acttcatgat
     1861 attcggcaag caggcatcga ctagttaatt aaggtttact ccctatcagt gatagagaac
     1921 gtatgaagag tttactccct atcagtgata gagaacgtat gcagacttta ctccctatca
     1981 gtgatagaga acgtataagg agtttactcc ctatcagtga tagagaacgt atgaccagtt
     2041 tactccctat cagtgataga gaacgtatct acagtttact ccctatcagt gatagagaac
     2101 gtatatccag tttactccct atcagtgata gagaacgtat aagctttagg cgtgtacggt
     2161 gggcgcctat aaaagcagag ctcgtttagt gaaccgtcag atcgcctgga gcaattccac
     2221 aacacttttg tcttatacca actttccgta ccacttccta ccctcgtaaa cttaagaatt
     2281 cgccaccatg gcggccgcta atgtgagtta gctcactcat taggcacccc aggctttaca
     2341 ctttatgctt ccggctcgta tgttgtgtgg aattgtgagc ggataacaat ttcacacagg
     2401 aaacagctat gaccatgatt acgccaagct tgcatgcctg caggtcgact ctagaggatc
     2461 cccgggtacc gagctcgaat tcactggccg tcgttttaca acgtcgtgac tgggaaaacc
     2521 ctggcgttac ccaacttaat cgccttgcag cacatccccc tttcgccagc tggcgtaata
     2581 gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat ggcgaatggc
     2641 gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atatggtgca
     2701 ctctcagtac aatctgctct gatgccgcat agcccagccc gcctaatgag cgggcttttt
     2761 tttgaacaaa ttaagcctat aacgcggaag cggccgcaat caacctctgg attacaaaat
     2821 ttgtgaaaga ttgactggta ttcttaacta tgttgctcct tttacgctat gtggatacgc
     2881 tgctttaatg cctttgtatc atgctattgc ttcccgtatg gctttcattt tctcctcctt
     2941 gtataaatcc tggttgctgt ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg
     3001 cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt tggggcattg ccaccacctg
     3061 tcagctcctt tccgggactt tcgctttccc cctccctatt gccacggcgg aactcatcgc
     3121 cgcctgcctt gcccgctgct ggacaggggc tcggctgttg ggcactgaca attccgtggt
     3181 gttgtcgggg aagctgacgt cctttccatg gctgctcgcc tgtgttgcca cctggattct
     3241 gcgcgggacg tccttctgct acgtcccttc ggccctcaat ccagcggacc ttccttcccg
     3301 cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg
     3361 gatctccctt tgggccgcct ccccgccctt tgggccgcct ccccgcgcta gtgctgggga
     3421 tgcggtgggc tctatgggtc tcgacattga ttattgacta gttattaata gtaatcaatt
     3481 acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat
     3541 ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt
     3601 cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa
     3661 actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc
     3721 aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct
     3781 acttggcagt acatctacgt attagtcatc gctattacca tggtcgaggt gagccccacg
     3841 ttctgcttca ctctccccat ctcccccccc tccccacccc caattttgta tttatttatt
     3901 ttttaattat tttgtgcagc gatgggggcg gggggggggg gcgcgcgcca ggcggggcgg
     3961 gggggggggg gggggggggg gggggggggg gggggggggg gggggggcgc cgggccgggg
     4021 gggggggggg gggggttttt ctttttgggg gggggggggg cgggggggcc cctaaaaaaa
     4081 agggcggcgc cgcggggggg gggcggcgcg cgcggccctc tccccgtgcc ccgctccgcc
     4141 gccgcctcgc gccgcccgcc ccggctctga ctgaccgcgt tactcccaca ggtgagcggg
     4201 cgggacggcc cttctcctcc gggctgtaat tagcgcttgg tttaatgacg gcttgtttct
     4261 tttctgtggc tgcgtgaaag ccttgagggg ctccgggagg gccctttgtg cggggggagc
     4321 ggctcggggg gtgcgtgcgt gtgtgtgtgc gtggggagcg ccgcgtgcgg ctccgcgctg
     4381 cccggcggct gtgagcgctg cgggcgcggc gcggggcttt gtgcgctccg cagtgtgcgc
     4441 gaggggagcg cggccggggg cggtgccccg cggtgcgggg ggggctgcga ggggaacaaa
     4501 ggctgcgtgc ggggtgtgtg cgtggggggg tgagcagggg gtgtgggcgc gtcggtcggg
     4561 ctgcaacccc ccctgcaccc ccctccccga gttgctgagc gcggcccggc ttcgggtgcg
     4621 gggctccgta cggggcgtgg cgcggggctc gccgtgccgg gcggggggtg gcggcaggtg
     4681 ggggtgccgg gcggcccccg gagcgccggc ggctgtcgag gcgcggcgag ccgcagccat
     4741 tgccttttat ggtaatcgtg cgagagggcg cagggacttc ctttgtccca aatctgtgcg
     4801 gagccgaaat ctgggaggcg ccgccgcacc ccctctagcg ggcgcggggc gaagcggtgc
     4861 ggcgccggca ggaaggaaat gggcggggag ggccttcgtg cgtcgccgcg ccgccgtccc
     4921 cttctccctc tccagcctcg gggctgtccg cggggggacg gctgccttcg ggggggacgg
     4981 ggcagggcgg ggttcggctt ctggcgtgtg accggcggct ctagagcctc tgctaaccat
     5041 gttcatgcct tcttcttttt cctacagctc ctgggcaacg tgctggttat tgtgctgtct
     5101 catcattttg gcaaagaatg tcgacatgag ccggctggac aagagcaaag tgatcaacag
     5161 cgccctggaa ctgctgaacg gcgtgggcat cgagggcctg accacccgga agctggccca
     5221 gaaactgggc gtggaacagc ccaccctgta ctggcacgtg aagaacaagc gggccctgct
     5281 ggacgccctg cccatcgaga tgctggaccg gcaccacacc cacagctgcc ctctggaagg
     5341 cgagagctgg caggacttcc tgcggaacaa cgccaagagc tacagatgcg ccctgctgag
     5401 ccaccgggac ggcgccaaag tgcacctggg caccagaccc accgagaagc agtacgagac
     5461 actggaaaac cagctggcct tcctgtgcca gcagggcttc agcctggaaa acgccctgta
     5521 cgccctgagc gccgtgggcc acttcaccct gggctgcgtg ctggaagaac aggaacacca
     5581 ggtcgccaaa gaggaaagag agacacccac caccgacagc atgccccccc tgctgaagca
     5641 ggccatcgag ctgttcgaca gacagggcgc cgagcccgcc ttcctgttcg gcctggaact
     5701 gatcatctgc ggcctcgaga agcagctgaa gtgcgagagc ggcggaccca ccgacgccct
     5761 ggacgacttc gacctggaca tgctgcccgc cgatgccctg gatgattttg atctggatat
     5821 gctccctgct gacgctctcg atgacttcga tctcgatatg ctgccaggcg agggcagggg
     5881 aagtcttcta acatgcgggg acgtggagga aaatcccggc cccgtgtcta agggcgaaga
     5941 gctgattaag gagaacatgc acatgaagct gtacatggag ggcaccgtgg acaaccatca
     6001 cttcaagtgc acatccgagg gcgaaggcaa gccctacgag ggcacccaga ccatgagaat
     6061 caaggtggtc gagggcggcc ctctcccctt cgccttcgac atcctggcta ctagcttcct
     6121 ctacggcagc aagaccttca tcaaccacac ccagggcatc cccgacttct tcaagcagtc
     6181 cttccctgag ggcttcacat gggagagagt caccacatac gaagacgggg gcgtgctgac
     6241 cgctacccag gacaccagcc tccaggacgg ctgcctcatc tacaacgtca agatcagagg
     6301 ggtgaacttc acatccaacg gccctgtgat gcagaagaaa acactcggct gggaggcctt
     6361 caccgagacg ctgtaccccg ctgacggcgg cctggaaggc agaaacgaca tggccctgaa
     6421 gctcgtgggc gggagccatc tgatcgcaaa cgccaagacc acatatagat ccaagaaacc
     6481 cgctaagaac ctcaagatgc ctggcgtcta ctatgtggac tacagactgg aaagaatcaa
     6541 ggaggccaac aacgagacct acgtcgagca gcacgaggtg gcagtggcca gatactgcga
     6601 cctccctagc aaactggggc acaagcttaa ttccggactc agatctcgag ctgatccaaa
     6661 aaagaagaga aaggtagatc caaaaaagaa gagaaaggta gatccaaaaa agaagagaaa
     6721 ggtaggatcc accggatcta gataactgat cataatcagc cataccacat ttgaaacccg
     6781 ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt
     6841 gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat
     6901 tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag
     6961 caagggggag gattgggaag acaatagcag gcatgctggg gatgcggtgg gctctatggc
     7021 tagccgctgc aagaactctt cctcacggat ccgaagttcc tattccgaag ttcctattct
     7081 tcaaatagta taggaacttc agcgctcact ggccgtcgtt ttacaaaggg cgaattcact
     7141 ggccgtcgtt ttacaacgtc gtgactggga aaaccctggc gttacccaac ttaatcgcct
     7201 tgcagcacat ccccctttcg ccagctggcg taatagcgaa gaggcccgca ccgatcgccc
     7261 ttcccaacag ttgcgcagcc tgaatggcga atggcgcctg atgcggtatt ttctccttac
     7321 gcatctgtgc ggtatttcac accgcatacg tcaaagcaac catagtacgc gccctgtagc
     7381 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     7441 gccttagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     7501 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
     7561 ctcgacccca aaaaacttga tttgggtgat ggttcacgta gtgggccatc gccctgatag
     7621 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
     7681 actggaacaa cactcaactc tatctcgggc tattcttttg atttataagg gattttgccg
     7741 atttcggtct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac
     7801 aaaatattaa cgtttacaat tttatggtgc actctcagta caatctgctc tgatgccgca
     7861 tagttaagcc agccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg
     7921 ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg
     7981 ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg cctattttta
     8041 taggttaatg tcatgataat aatggtttct tagacgtcag gtggcacttt tcggggaaat
     8101 gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
     8161 agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa
     8221 catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac
     8281 ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac
     8341 atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt
     8401 ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc
     8461 gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca
     8521 ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc
     8581 ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag
     8641 gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa
     8701 ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg
     8761 gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa
     8821 ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg
     8881 gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt
     8941 gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt
     9001 caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag
     9061 cattggtaac tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat
     9121 ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct
     9181 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     9241 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     9301 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     9361 agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc
     9421 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     9481 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     9541 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     9601 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
     9661 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     9721 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     9781 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     9841 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg
     9901 ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc
     9961 cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaag
//
