LOCUS       sequence-485738-       11839 bp    DNA     circular SYN 10-SEP-2025
DEFINITION  Bxb1-GA donor plasmid with downstream tandem syntax for all-in-one 
            doxycycline-inducible expression of ISL1 and LHX3.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11839)
  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 11839)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Sep 10, 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_ISL1-LHX3_Down-Tandem
FEATURES             Location/Qualifiers
     source          1..11839
                     /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    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      3078..3087
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3359..3412
                     /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"
     misc_feature    4628..5216
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4681..4701)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5099..5110)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     enhancer        5273..5652
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        5654..5928
                     /label=chicken beta-actin promoter
     primer_bind     6821..6843
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     6905..6924
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             6956..7699
                     /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             7700..7753
                     /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             7754..8461
                     /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             8486..8506
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             8510..8530
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             8534..8554
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(8619..8636)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    8625..8849
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(8938..8954)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8968..8984)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8977..8999)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      9197..9652
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9284..9303)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     9494..9515
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(9669..9688)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     9788..9810
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(9848..9866)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        9934..10038
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(10039..10107,10108..10899)
                     /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(10257..10276)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      11070..11658
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     11559..11578
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     11812..11829
                     /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 ggagacatgg gagatccacc aaaaaaaaaa cgtctgattt ccctatgtgt
     2341 tggttgcggc aatcagattc acgatcagta tattctgagg gtttctccgg atttggaatg
     2401 gcatgcggca tgtttgaaat gtgcggagtg taatcagtat ttggacgaga gctgtacatg
     2461 ctttgttagg gatgggaaaa cctactgtaa aagagattat atcaggttgt acgggatcaa
     2521 atgcgccaag tgcagcatcg gcttcagcaa gaacgacttc gtgatgcgtg cccgctccaa
     2581 ggtgtatcac atcgagtgtt tccgctgtgt ggcctgcagc cgccagctca tccctgggga
     2641 cgaatttgcg cttcgggagg acggtctctt ctgccgagca gaccacgatg tggtggagag
     2701 ggccagtcta ggcgctggcg acccgctcag tcccctgcat ccagcgcggc cactgcaaat
     2761 ggcagcggag cccatctccg ccaggcagcc ggccctgcgg ccccacgtcc acaagcagcc
     2821 ggagaagacc acccgcgtgc ggactgtgct gaacgagaag cagctgcaca ccttgcggac
     2881 ctgctacgcc gcaaacccgc ggccagatgc gctcatgaag gagcaactgg tagagatgac
     2941 gggcctcagt ccccgtgtga tccgggtctg gtttcaaaac aagcggtgca aggacaagaa
     3001 gcgaagcatc atgatgaagc aactccagca gcagcagccc aatgacaaaa ctaatatcca
     3061 ggggatgaca ggaactccca tggtggctgc cagtccagag agacacgacg gtggcttaca
     3121 ggctaaccca gtggaagtac aaagttacca gccaccttgg aaagtactga gcgacttcgc
     3181 cttgcagagt gacatagatc agcctgcttt tcagcaactg gtcaattttt cagaaggagg
     3241 accgggctct aattccactg gcagtgaagt agcatcaatg tcctctcaac ttccagatac
     3301 acctaacagc atggtagcca gtcctattga ggcattgatg cacagcagag gctctggcga
     3361 gggcagaggc agcctgctga catgtggcga cgtggaagag aaccctggcc ccggtaccat
     3421 ggaagccaga ggcgaactgg gccctgccag agaatctgcc ggcggagatc tgctgctggc
     3481 cctgctggct agaagggccg acctgagaag agaaatcccc ctgtgcgccg gctgcgacca
     3541 gcacatcctg gaccggttta tcctgaaggc cctggacaga cactggcaca gcaagtgcct
     3601 gaagtgcagc gactgccaca cccccctggc cgagagatgc tttagcagag gcgagagcgt
     3661 gtactgcaag gacgacttct tcaagagatt cggcaccaag tgcgccgcct gccagctggg
     3721 aattcctcct acacaggtcg tgcggagagc ccaggacttc gtgtaccatc tgcactgctt
     3781 cgcctgcgtc gtgtgcaaga gacagctggc caccggcgac gagttctacc tgatggaaga
     3841 tagccggctc gtgtgtaaag ccgactacga gacagccaag cagagagagg ccgaggccac
     3901 cgccaaaaga cccagaacca ccatcaccgc caagcagctg gaaaccctga agtccgccta
     3961 caacaccagc cccaagcccg ccagacatgt gcgggaacag ctgagcagcg agacaggcct
     4021 ggacatgaga gtggtgcaag tgtggttcca gaacagacgg gccaaagaga agcggctgaa
     4081 gaaggacgcc ggcaggcaga gatggggcca gtacttccgg aacatgaagc ggtccagagg
     4141 cggcagcaag agcgacaagg acagcgtgca ggaaggccag gacagcgacg ccgaggtgtc
     4201 cttcccagat gagcctagcc tggccgaaat gggcccagcc aatggcctgt atggcagcct
     4261 gggagagcca acccaggctc tgggcagacc ttctggcgcc ctgggcaact tctctctgga
     4321 acacggcgga ctggccggac ccgagcagta tagagaactg aggcccggca gcccttacgg
     4381 cgtgccacct tctccagctg cccctcagtc tctgccagga cctcagcctc tgctgtccag
     4441 cctggtgtac cccgatacaa gcctgggcct ggtgccatct ggcgctccag gcggacctcc
     4501 tccaatgaga gtgctggccg gcaatggccc tagcagcgat ctgagcacag gcagctctgg
     4561 cggctacccc gattttcctg ccagccctgc ctcctggctg gacgaagtgg atcacgccca
     4621 gttctgaaat caacctctgg attacaaaat ttgtgaaaga ttgactggta ttcttaacta
     4681 tgttgctcct tttacgctat gtggatacgc tgctttaatg cctttgtatc atgctattgc
     4741 ttcccgtatg gctttcattt tctcctcctt gtataaatcc tggttgctgt ctctttatga
     4801 ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac
     4861 ccccactggt tggggcattg ccaccacctg tcagctcctt tccgggactt tcgctttccc
     4921 cctccctatt gccacggcgg aactcatcgc cgcctgcctt gcccgctgct ggacaggggc
     4981 tcggctgttg ggcactgaca attccgtggt gttgtcgggg aagctgacgt cctttccatg
     5041 gctgctcgcc tgtgttgcca cctggattct gcgcgggacg tccttctgct acgtcccttc
     5101 ggccctcaat ccagcggacc ttccttcccg cggcctgctg ccggctctgc ggcctcttcc
     5161 gcgtcttcgc cttcgccctc agacgagtcg gatctccctt tgggccgcct ccccgccctt
     5221 tgggccgcct ccccgcgcta gtgctgggga tgcggtgggc tctatgggtc tcgacattga
     5281 ttattgacta gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg
     5341 gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc
     5401 cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat
     5461 tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat
     5521 catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat
     5581 gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc
     5641 gctattacca tggtcgaggt gagccccacg ttctgcttca ctctccccat ctcccccccc
     5701 tccccacccc caattttgta tttatttatt ttttaattat tttgtgcagc gatgggggcg
     5761 gggggggggg ggcgcgcgcc aggcggggcg gggcggggcg aggggcgggg cggggcgagg
     5821 cggagaggtg cggcggcagc caatcagagc ggcgcgctcc gaaagtttcc ttttatggcg
     5881 aggcggcggc ggcggcggcc ctataaaaag cgaagcgcgc ggcgggcggg agtcgctgcg
     5941 cgctgccttc gccccgtgcc ccgctccgcc gccgcctcgc gccgcccgcc ccggctctga
     6001 ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc gggctgtaat
     6061 tagcgcttgg tttaatgacg gcttgtttct tttctgtggc tgcgtgaaag ccttgagggg
     6121 ctccgggagg gccctttgtg cggggggagc ggctcggggg gtgcgtgcgt gtgtgtgtgc
     6181 gtggggagcg ccgcgtgcgg ctccgcgctg cccggcggct gtgagcgctg cgggcgcggc
     6241 gcggggcttt gtgcgctccg cagtgtgcgc gaggggagcg cggccggggg cggtgccccg
     6301 cggtgcgggg ggggctgcga ggggaacaaa ggctgcgtgc ggggtgtgtg cgtggggggg
     6361 tgagcagggg gtgtgggcgc gtcggtcggg ctgcaacccc ccctgcaccc ccctccccga
     6421 gttgctgagc gcggcccggc ttcgggtgcg gggctccgta cggggcgtgg cgcggggctc
     6481 gccgtgccgg gcggggggtg gcggcaggtg ggggtgccgg gcggcccccg gagcgccggc
     6541 ggctgtcgag gcgcggcgag ccgcagccat tgccttttat ggtaatcgtg cgagagggcg
     6601 cagggacttc ctttgtccca aatctgtgcg gagccgaaat ctgggaggcg ccgccgcacc
     6661 ccctctagcg ggcgcggggc gaagcggtgc ggcgccggca ggaaggaaat gggcggggag
     6721 ggccttcgtg cgtcgccgcg ccgccgtccc cttctccctc tccagcctcg gggctgtccg
     6781 cggggggacg gctgccttcg ggggggacgg ggcagggcgg ggttcggctt ctggcgtgtg
     6841 accggcggct ctagagcctc tgctaaccat gttcatgcct tcttcttttt cctacagctc
     6901 ctgggcaacg tgctggttat tgtgctgtct catcattttg gcaaagaatg tcgacatgag
     6961 ccggctggac aagagcaaag tgatcaacag cgccctggaa ctgctgaacg gcgtgggcat
     7021 cgagggcctg accacccgga agctggccca gaaactgggc gtggaacagc ccaccctgta
     7081 ctggcacgtg aagaacaagc gggccctgct ggacgccctg cccatcgaga tgctggaccg
     7141 gcaccacacc cacagctgcc ctctggaagg cgagagctgg caggacttcc tgcggaacaa
     7201 cgccaagagc tacagatgcg ccctgctgag ccaccgggac ggcgccaaag tgcacctggg
     7261 caccagaccc accgagaagc agtacgagac actggaaaac cagctggcct tcctgtgcca
     7321 gcagggcttc agcctggaaa acgccctgta cgccctgagc gccgtgggcc acttcaccct
     7381 gggctgcgtg ctggaagaac aggaacacca ggtcgccaaa gaggaaagag agacacccac
     7441 caccgacagc atgccccccc tgctgaagca ggccatcgag ctgttcgaca gacagggcgc
     7501 cgagcccgcc ttcctgttcg gcctggaact gatcatctgc ggcctcgaga agcagctgaa
     7561 gtgcgagagc ggcggaccca ccgacgccct ggacgacttc gacctggaca tgctgcccgc
     7621 cgatgccctg gatgattttg atctggatat gctccctgct gacgctctcg atgacttcga
     7681 tctcgatatg ctgccaggcg agggcagggg aagtcttcta acatgcgggg acgtggagga
     7741 aaatcccggc cccgtgtcta agggcgaaga gctgattaag gagaacatgc acatgaagct
     7801 gtacatggag ggcaccgtgg acaaccatca cttcaagtgc acatccgagg gcgaaggcaa
     7861 gccctacgag ggcacccaga ccatgagaat caaggtggtc gagggcggcc ctctcccctt
     7921 cgccttcgac atcctggcta ctagcttcct ctacggcagc aagaccttca tcaaccacac
     7981 ccagggcatc cccgacttct tcaagcagtc cttccctgag ggcttcacat gggagagagt
     8041 caccacatac gaagacgggg gcgtgctgac cgctacccag gacaccagcc tccaggacgg
     8101 ctgcctcatc tacaacgtca agatcagagg ggtgaacttc acatccaacg gccctgtgat
     8161 gcagaagaaa acactcggct gggaggcctt caccgagacg ctgtaccccg ctgacggcgg
     8221 cctggaaggc agaaacgaca tggccctgaa gctcgtgggc gggagccatc tgatcgcaaa
     8281 cgccaagacc acatatagat ccaagaaacc cgctaagaac ctcaagatgc ctggcgtcta
     8341 ctatgtggac tacagactgg aaagaatcaa ggaggccaac aacgagacct acgtcgagca
     8401 gcacgaggtg gcagtggcca gatactgcga cctccctagc aaactggggc acaagcttaa
     8461 ttccggactc agatctcgag ctgatccaaa aaagaagaga aaggtagatc caaaaaagaa
     8521 gagaaaggta gatccaaaaa agaagagaaa ggtaggatcc accggatcta gataactgat
     8581 cataatcagc cataccacat ttgaaacccg ctgatcagcc tcgactgtgc cttctagttg
     8641 ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc
     8701 cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc
     8761 tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag
     8821 gcatgctggg gatgcggtgg gctctatggc tagccgctgc aagaactctt cctcacggat
     8881 ccgaagttcc tattccgaag ttcctattct tcaaatagta taggaacttc agcgctcact
     8941 ggccgtcgtt ttacaaaggg cgaattcact ggccgtcgtt ttacaacgtc gtgactggga
     9001 aaaccctggc gttacccaac ttaatcgcct tgcagcacat ccccctttcg ccagctggcg
     9061 taatagcgaa gaggcccgca ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga
     9121 atggcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatacg
     9181 tcaaagcaac catagtacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt
     9241 acgcgcagcg tgaccgctac acttgccagc gccttagcgc ccgctccttt cgctttcttc
     9301 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct
     9361 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga tttgggtgat
     9421 ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc
     9481 acgttcttta atagtggact cttgttccaa actggaacaa cactcaactc tatctcgggc
     9541 tattcttttg atttataagg gattttgccg atttcggtct attggttaaa aaatgagctg
     9601 atttaacaaa aatttaacgc gaattttaac aaaatattaa cgtttacaat tttatggtgc
     9661 actctcagta caatctgctc tgatgccgca tagttaagcc agccccgaca cccgccaaca
     9721 cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg
     9781 accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga
     9841 cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat aatggtttct
     9901 tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc
     9961 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa
    10021 tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt
    10081 gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct
    10141 gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc
    10201 cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta
    10261 tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac
    10321 tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc
    10381 atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac
    10441 ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg
    10501 gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac
    10561 gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc
    10621 gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt
    10681 gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga
    10741 gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc
    10801 cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag
    10861 atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca
    10921 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
    10981 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
    11041 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
    11101 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
    11161 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt
    11221 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
    11281 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
    11341 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
    11401 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
    11461 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
    11521 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
    11581 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
    11641 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
    11701 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt
    11761 accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca
    11821 gtgagcgagg aagcggaag
//
