LOCUS       sequence-462053-       10132 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bxb1-GT donor plasmid with downstream tandem syntax for all-in-one 
            doxycycline-inducible mScarlet expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10132)
  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 10132)
  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-GT_AIO-TetOn_mScarlet_Down-Tandem
FEATURES             Location/Qualifiers
     source          1..10132
                     /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     186..208
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     205..221
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     205..221
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     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    complement(1802..1835)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    1881..1899
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1917..1935
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1953..1971
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1989..2007
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2025..2043
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2061..2079
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2097..2115
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     2160..2184
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      2263..2272
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2269..2964
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet
                     /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFSWDILSPQFMYGSRAFTKHPADIPDYYKQSFPEGFKWERVMNFEDGG
                     AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK
                     MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH
                     STGGMDELYK"
     primer_bind     complement(2402..2420)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2670..2689
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    2968..3556
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3021..3041)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3439..3450)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     enhancer        3613..3992
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     5164..5186
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     5248..5267
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             5299..6042
                     /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             6043..6096
                     /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             6097..6804
                     /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             6829..6849
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             6853..6873
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             6877..6897
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(6962..6979)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6968..7192
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    complement(7221..7253)
                     /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(7261..7278)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(7261..7277)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7270..7292)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      7490..7945
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7577..7596)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7787..7808
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(7962..7981)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     8081..8103
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(8141..8159)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        8227..8331
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(8332..8400,8401..9192)
                     /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(8550..8569)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      9363..9951
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     9852..9871
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     10105..10122
                     /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 gcggtctccg tcgtcaggat catccggatc
     1801 cataacttcg tatagcatac attatacgaa gttatcatga tattcggcaa gcaggcatcg
     1861 actagttaat taaggtttac tccctatcag tgatagagaa cgtatgaaga gtttactccc
     1921 tatcagtgat agagaacgta tgcagacttt actccctatc agtgatagag aacgtataag
     1981 gagtttactc cctatcagtg atagagaacg tatgaccagt ttactcccta tcagtgatag
     2041 agaacgtatc tacagtttac tccctatcag tgatagagaa cgtatatcca gtttactccc
     2101 tatcagtgat agagaacgta taagctttag gcgtgtacgg tgggcgccta taaaagcaga
     2161 gctcgtttag tgaaccgtca gatcgcctgg agcaattcca caacactttt gtcttatacc
     2221 aactttccgt accacttcct accctcgtaa acttaagaat tcgccaccat ggtgagcaag
     2281 ggcgaggcag tgatcaagga gttcatgcgg ttcaaggtgc acatggaggg ctccatgaac
     2341 ggccacgagt tcgagatcga gggcgagggc gagggccgcc cctacgaggg cacccagacc
     2401 gccaagctga aggtgaccaa gggtggcccc ctgcccttct cctgggacat cctgtcccct
     2461 cagttcatgt acggctccag ggccttcacc aagcaccccg ccgacatccc cgactactat
     2521 aagcagtcct tccccgaggg cttcaagtgg gagcgcgtga tgaacttcga ggacggcggc
     2581 gccgtgaccg tgacccagga cacctccctg gaggacggca ccctgatcta caaggtgaag
     2641 ctccgcggca ccaacttccc tcctgacggc cccgtaatgc agaagaagac aatgggctgg
     2701 gaagcgtcca ccgagcggtt gtaccccgag gacggcgtgc tgaagggcga cattaagatg
     2761 gccctgcgcc tgaaggacgg cggccgctac ctggcggact tcaagaccac ctacaaggcc
     2821 aagaagcccg tgcagatgcc cggcgcctac aacgtcgacc gcaagttgga catcacctcc
     2881 cacaacgagg actacaccgt ggtggaacag tacgaacgct ccgagggccg ccactccacc
     2941 ggcggcatgg acgagctgta caagtaaaat caacctctgg attacaaaat ttgtgaaaga
     3001 ttgactggta ttcttaacta tgttgctcct tttacgctat gtggatacgc tgctttaatg
     3061 cctttgtatc atgctattgc ttcccgtatg gctttcattt tctcctcctt gtataaatcc
     3121 tggttgctgt ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc
     3181 actgtgtttg ctgacgcaac ccccactggt tggggcattg ccaccacctg tcagctcctt
     3241 tccgggactt tcgctttccc cctccctatt gccacggcgg aactcatcgc cgcctgcctt
     3301 gcccgctgct ggacaggggc tcggctgttg ggcactgaca attccgtggt gttgtcgggg
     3361 aagctgacgt cctttccatg gctgctcgcc tgtgttgcca cctggattct gcgcgggacg
     3421 tccttctgct acgtcccttc ggccctcaat ccagcggacc ttccttcccg cggcctgctg
     3481 ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg gatctccctt
     3541 tgggccgcct ccccgccctt tgggccgcct ccccgcgcta gtgctgggga tgcggtgggc
     3601 tctatgggtc tcgacattga ttattgacta gttattaata gtaatcaatt acggggtcat
     3661 tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg
     3721 gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa
     3781 cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact
     3841 tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta
     3901 aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt
     3961 acatctacgt attagtcatc gctattacca tggtcgaggt gagccccacg ttctgcttca
     4021 ctctccccat ctcccccccc tccccacccc caattttgta tttatttatt ttttaattat
     4081 tttgtgcagc gatgggggcg gggggggggg gggcgcgcgc caggcggggc gggggggggg
     4141 gggggggggg gggggggggg gggggggggg ggggggggcg cccaggcggg ggggggggcc
     4201 ccgaaggttt ctttttttgc cggggggggg gggggggggc cccctaaaaa aaggagggcg
     4261 ggcggcgggg ggggggggcg gggccgcgct ctccccccgt gccccgctcc gccgccgcct
     4321 cgcgccgccc gccccggctc tgactgaccg cgttactccc acaggtgagc gggcgggacg
     4381 gcccttctcc tccgggctgt aattagcgct tggtttaatg acggcttgtt tcttttctgt
     4441 ggctgcgtga aagccttgag gggctccggg agggcccttt gtgcgggggg agcggctcgg
     4501 ggggtgcgtg cgtgtgtgtg tgcgtgggga gcgccgcgtg cggctccgcg ctgcccggcg
     4561 gctgtgagcg ctgcgggcgc ggcgcggggc tttgtgcgct ccgcagtgtg cgcgagggga
     4621 gcgcggccgg gggcggtgcc ccgcggtgcg gggggggctg cgaggggaac aaaggctgcg
     4681 tgcggggtgt gtgcgtgggg gggtgagcag ggggtgtggg cgcgtcggtc gggctgcaac
     4741 cccccctgca cccccctccc cgagttgctg agcgcggccc ggcttcgggt gcggggctcc
     4801 gtacggggcg tggcgcgggg ctcgccgtgc cgggcggggg gtggcggcag gtgggggtgc
     4861 cgggcggccc ccggagcgcc ggcggctgtc gaggcgcggc gagccgcagc cattgccttt
     4921 tatggtaatc gtgcgagagg gcgcagggac ttcctttgtc ccaaatctgt gcggagccga
     4981 aatctgggag gcgccgccgc accccctcta gcgggcgcgg ggcgaagcgg tgcggcgccg
     5041 gcaggaagga aatgggcggg gagggccttc gtgcgtcgcc gcgccgccgt ccccttctcc
     5101 ctctccagcc tcggggctgt ccgcgggggg acggctgcct tcggggggga cggggcaggg
     5161 cggggttcgg cttctggcgt gtgaccggcg gctctagagc ctctgctaac catgttcatg
     5221 ccttcttctt tttcctacag ctcctgggca acgtgctggt tattgtgctg tctcatcatt
     5281 ttggcaaaga atgtcgacat gagccggctg gacaagagca aagtgatcaa cagcgccctg
     5341 gaactgctga acggcgtggg catcgagggc ctgaccaccc ggaagctggc ccagaaactg
     5401 ggcgtggaac agcccaccct gtactggcac gtgaagaaca agcgggccct gctggacgcc
     5461 ctgcccatcg agatgctgga ccggcaccac acccacagct gccctctgga aggcgagagc
     5521 tggcaggact tcctgcggaa caacgccaag agctacagat gcgccctgct gagccaccgg
     5581 gacggcgcca aagtgcacct gggcaccaga cccaccgaga agcagtacga gacactggaa
     5641 aaccagctgg ccttcctgtg ccagcagggc ttcagcctgg aaaacgccct gtacgccctg
     5701 agcgccgtgg gccacttcac cctgggctgc gtgctggaag aacaggaaca ccaggtcgcc
     5761 aaagaggaaa gagagacacc caccaccgac agcatgcccc ccctgctgaa gcaggccatc
     5821 gagctgttcg acagacaggg cgccgagccc gccttcctgt tcggcctgga actgatcatc
     5881 tgcggcctcg agaagcagct gaagtgcgag agcggcggac ccaccgacgc cctggacgac
     5941 ttcgacctgg acatgctgcc cgccgatgcc ctggatgatt ttgatctgga tatgctccct
     6001 gctgacgctc tcgatgactt cgatctcgat atgctgccag gcgagggcag gggaagtctt
     6061 ctaacatgcg gggacgtgga ggaaaatccc ggccccgtgt ctaagggcga agagctgatt
     6121 aaggagaaca tgcacatgaa gctgtacatg gagggcaccg tggacaacca tcacttcaag
     6181 tgcacatccg agggcgaagg caagccctac gagggcaccc agaccatgag aatcaaggtg
     6241 gtcgagggcg gccctctccc cttcgccttc gacatcctgg ctactagctt cctctacggc
     6301 agcaagacct tcatcaacca cacccagggc atccccgact tcttcaagca gtccttccct
     6361 gagggcttca catgggagag agtcaccaca tacgaagacg ggggcgtgct gaccgctacc
     6421 caggacacca gcctccagga cggctgcctc atctacaacg tcaagatcag aggggtgaac
     6481 ttcacatcca acggccctgt gatgcagaag aaaacactcg gctgggaggc cttcaccgag
     6541 acgctgtacc ccgctgacgg cggcctggaa ggcagaaacg acatggccct gaagctcgtg
     6601 ggcgggagcc atctgatcgc aaacgccaag accacatata gatccaagaa acccgctaag
     6661 aacctcaaga tgcctggcgt ctactatgtg gactacagac tggaaagaat caaggaggcc
     6721 aacaacgaga cctacgtcga gcagcacgag gtggcagtgg ccagatactg cgacctccct
     6781 agcaaactgg ggcacaagct taattccgga ctcagatctc gagctgatcc aaaaaagaag
     6841 agaaaggtag atccaaaaaa gaagagaaag gtagatccaa aaaagaagag aaaggtagga
     6901 tccaccggat ctagataact gatcataatc agccatacca catttgaaac ccgctgatca
     6961 gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc
     7021 ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg
     7081 cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg
     7141 gaggattggg aagacaatag caggcatgct ggggatgcgg tgggctctat ggctagccgc
     7201 tgcaagaact cttcctcacg ataacttcgt ataggagact ttatacgaag ttaagcgctc
     7261 actggccgtc gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg
     7321 ccttgcagca catccccctt tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg
     7381 cccttcccaa cagttgcgca gcctgaatgg cgaatggcgc ctgatgcggt attttctcct
     7441 tacgcatctg tgcggtattt cacaccgcat acgtcaaagc aaccatagta cgcgccctgt
     7501 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     7561 agcgccttag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     7621 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     7681 cacctcgacc ccaaaaaact tgatttgggt gatggttcac gtagtgggcc atcgccctga
     7741 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     7801 caaactggaa caacactcaa ctctatctcg ggctattctt ttgatttata agggattttg
     7861 ccgatttcgg tctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt
     7921 aacaaaatat taacgtttac aattttatgg tgcactctca gtacaatctg ctctgatgcc
     7981 gcatagttaa gccagccccg acacccgcca acacccgctg acgcgccctg acgggcttgt
     8041 ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag
     8101 aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg gcctcgtgat acgcctattt
     8161 ttataggtta atgtcatgat aataatggtt tcttagacgt caggtggcac ttttcgggga
     8221 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
     8281 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt
     8341 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct
     8401 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt
     8461 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt
     8521 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac
     8581 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac
     8641 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct
     8701 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg
     8761 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg
     8821 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca
     8881 atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa
     8941 caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt
     9001 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc
     9061 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg
     9121 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt
     9181 aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt
     9241 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc
     9301 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     9361 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     9421 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     9481 ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac
     9541 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     9601 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     9661 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     9721 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     9781 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
     9841 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
     9901 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
     9961 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct
    10021 gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct
    10081 cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga ag
//
