LOCUS       sequence-366939-        8772 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for inducible mammalian cell expression of the sg61 helixCAM
            and constitutive expression of an identifying fluorescent protein 
            (eGFP). Flanked with PiggyBac sites for easy line construction.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8772)
  AUTHORS   Chao G, Wannier TM, Gutierrez C, Borders NC, Appleton E, Chadha A, 
            Lebar T, Church GM
  TITLE     helixCAM: A platform for programmable cellular assembly in bacteria 
            and human cells.
  JOURNAL   Cell. 2022 Aug 23. pii: S0092-8674(22)01061-3. doi: 
            10.1016/j.cell.2022.08.012.
  PUBMED    36055250
REFERENCE   2  (bases 1 to 8772)
  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: "Cell. 2022 
            Aug 23. pii: S0092-8674(22)01061-3. doi: 
            10.1016/j.cell.2022.08.012."
COMMENT     Sequence Label: PB_TRE_sg61-PDGFR_EF1a_eGFP
FEATURES             Location/Qualifiers
     source          1..8772
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(57..161)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(187..642)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(324..345)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     536..555
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     769..791
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     783..800
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     784..800
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   930..992
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     promoter        1441..1819
                     /label=TRE3G promoter
                     /note="3rd-generation Tet-responsive promoter that can be 
                     activated by binding of Tet-On(R) 3G"
     protein_bind    1449..1467
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1485..1503
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1521..1539
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1557..1575
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1593..1611
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1629..1647
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1665..1683
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     promoter        1871..1909
                     /label=minimal CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     protein_bind    1954..1978
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     sig_peptide     1978..2037
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     CDS             2047..2064
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2191..2220
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2224..2370
                     /codon_start=1
                     /product="transmembrane domain from platelet derived growth
                     factor receptor beta"
                     /label=PDGFR-beta TM domain
                     /translation="AVGQDTQEVIVVPHSLPFKVVVISAILALVVLTIISLIILIMLWQ
                     KKPR"
     polyA_signal    2387..2594
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     promoter        2671..2882
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     LTR             2895..3163
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human 
                     T-cell leukemia virus (HTLV) type 1"
     CDS             3197..3940
                     /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             3941..3994
                     /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             join(3995..3997,3998..4000,4001..4711)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(3995..3997,3998..4000,4001..4711)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4040..4061)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4301..4320)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4648..4669
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             4712..4765
                     /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             4766..5365
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(4766..4785)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5262..5282
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5398..5519
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5435..5454)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5489..5508
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     repeat_region   6282..6316
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(6807..6823)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6807..6823)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6820..6842)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6831..6847
                     /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(6855..6861,6862..6879,6880..6885))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6900..6921
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(7038..7055)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7209..7797)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7289..7308)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7968..8759,8760..8772,1..56))
                     /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     8591..8610
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
       61 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
      121 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
      181 ccacctaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca
      241 gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga
      301 ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg
      361 actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat
      421 caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag
      481 ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga
      541 agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa
      601 ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtcccattcg ccattcaggc
      661 tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga
      721 aagggggatg tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac
      781 gttgtaaaac gacggccagt gagcgcgcct cgttcattca cgtttttgaa cccgtggagg
      841 acgggcagac tcgcggtgca aatgtgtttt acagcgtgat ggagcagatg aagatgctcg
      901 acacgctgca gaacacgcag ctagattaac cctagaaaga taatcatatt gtgacgtacg
      961 ttaaagataa tcatgtgtaa aattgacgca tgtgttttat cggtctgtat atcgaggttt
     1021 atttattaat ttgaatagat attaagtttt attatattta cacttacata ctaataataa
     1081 attcaacaaa caatttattt atgtttattt atttattaaa aaaaacaaaa actcaaaatt
     1141 tcttctataa agtaacaaaa cttttatgag ggacagcccc cccccaaagc ccccagggat
     1201 gtaattacgt ccctcccccg ctagggggca gcagcgagcc gcccggggct ccgctccggt
     1261 ccggcgctcc ccccgcatcc ccgagccggc agcgtgcggg gacagcccgg gcacggggaa
     1321 ggtggcacgg gatcgctttc ctctgaacgc ttctcgctgc tctttgagcc tgcagacacc
     1381 tggggggata cggggaaaag gcctccacgg ccactagtct ttcgtcttca agaattcctc
     1441 gagtttactc cctatcagtg atagagaacg tatgaagagt ttactcccta tcagtgatag
     1501 agaacgtatg cagactttac tccctatcag tgatagagaa cgtataagga gtttactccc
     1561 tatcagtgat agagaacgta tgaccagttt actccctatc agtgatagag aacgtatcta
     1621 cagtttactc cctatcagtg atagagaacg tatatccagt ttactcccta tcagtgatag
     1681 agaacgtata agctttaggc gtgtacggtg ggcgcctata aaagcagagc tcgtttagtg
     1741 aaccgtcaga tcgcctggag caattccaca acacttttgt cttataccaa ctttccgtac
     1801 cacttcctac cctcgtaaag gtctagagct agcgaattcg aagccaccaa ctggcaggtg
     1861 cgtatgtcga ggtaggcgtg tacggtggga ggcctatata agcagagctc gtttagtgaa
     1921 ccgtcagatc gcctggagaa ttggctagca tcaacaagtt tgtacaaaaa agttggcatg
     1981 gagacagaca cactcctgct atgggtactg ctgctctggg ttccaggttc cactggtgac
     2041 ggaagccatc accaccatca tcacggtgca tcccccgagg ataaaatcca ggctctgaag
     2101 cagaaaaatg cgcaactaaa gcaagaaaat gcggcgcttg agcaaaagaa tgctcaacta
     2161 aaacaggaga tacaagcgtt agaacaagac gaacaaaaac tcatctcaga agaggatctg
     2221 aatgctgtgg gccaggacac gcaggaggtc atcgtggtgc cacactcctt gccctttaag
     2281 gtggtggtga tctcagccat cctggccctg gtggtgctca ccatcatctc ccttatcatc
     2341 ctcatcatgc tttggcagaa gaagccacgt taacggtacc cctcgactgt gcctactagt
     2401 tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact
     2461 cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat
     2521 tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc
     2581 aggcatgctg gggatgcggt gggctcttcg aatttaaatc ggatccagct gaatctaagt
     2641 ggatctgcga tcgctccggt gcccgtcagt gggcagagcg cacatcgccc acagtccccg
     2701 agaagttggg gggaggggtc ggcaattgaa cgggtgccta gagaaggtgg cgcggggtaa
     2761 actgggaaag tgatgtcgtg tactggctcc gcctttttcc cgagggtggg ggagaaccgt
     2821 atataagtgc agtagtcgcc gtgaacgttc tttttcgcaa cgggtttgcc gccagaacac
     2881 agctgaagct tcgaggggct cgcatctctc cttcacgcgc ccgccgccct acctgaggcc
     2941 gccatccacg ccggttgagt cgcgttctgc cgcctcccgc ctgtggtgcc tcctgaactg
     3001 cgtccgccgt ctaggtaagt ttaaagctca ggtcgagacc gggcctttgt ccggcgctcc
     3061 cttggagcct acctagactc agccggctct ccacgctttg cctgaccctg cttgctcaac
     3121 tctacgtctt tgtttcgttt tctgttctgc gccgttacag atctaagctg tgaccggcgc
     3181 ctactctaga gccgccatgt ctagactgga caagagcaaa gtcataaact ctgctctgga
     3241 attactcaat ggagtcggta tcgaaggcct gacgacaagg aaactcgctc aaaagctggg
     3301 agttgagcag cctaccctgt actggcacgt gaagaacaag cgggccctgc tcgatgccct
     3361 gccaatcgag atgctggaca ggcatcatac ccactcctgc cccctggaag gcgagtcatg
     3421 gcaagacttt ctgcggaaca acgccaagtc ataccgctgt gctctcctct cacatcgcga
     3481 cggggctaaa gtgcatctcg gcacccgccc aacagagaaa cagtacgaaa ccctggaaaa
     3541 tcagctcgcg ttcctgtgtc agcaaggctt ctccctggag aacgcactgt acgctctgtc
     3601 cgccgtgggc cactttacac tgggctgcgt attggaggaa caggagcatc aagtagcaaa
     3661 agaggaaaga gagacaccta ccaccgattc tatgccccca cttctgaaac aagcaattga
     3721 gctgttcgac cggcagggag ccgaacctgc cttccttttc ggcctggaac taatcatatg
     3781 tggcctggag aaacagctaa agtgcgaaag cggcgggccg accgacgccc ttgacgattt
     3841 tgacttagac atgctcccag ccgatgccct tgacgacttt gaccttgata tgctgcctgc
     3901 tgacgctctt gacgattttg accttgacat gctccccggg gagggcagag gaagtcttct
     3961 aacatgcggt gacgtggagg agaatcccgg ccctatggtg agcaagggcg aggagctgtt
     4021 caccggggtg gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc acaagttcag
     4081 cgtgtccggc gagggcgagg gcgatgccac ctacggcaag ctgaccctga agttcatctg
     4141 caccaccggc aagctgcccg tgccctggcc caccctcgtg accaccctga cctacggcgt
     4201 gcagtgcttc agccgctacc ccgaccacat gaagcagcac gacttcttca agtccgccat
     4261 gcccgaaggc tacgtccagg agcgcaccat cttcttcaag gacgacggca actacaagac
     4321 ccgcgccgag gtgaagttcg agggcgacac cctggtgaac cgcatcgagc tgaagggcat
     4381 cgacttcaag gaggacggca acatcctggg gcacaagctg gagtacaact acaacagcca
     4441 caacgtctat atcatggccg acaagcagaa gaacggcatc aaggtgaact tcaagatccg
     4501 ccacaacatc gaggacggca gcgtgcagct cgccgaccac taccagcaga acacccccat
     4561 cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agcacccagt ccgccctgag
     4621 caaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg
     4681 gatcactctc ggcatggacg agctgtacaa ggagggcaga ggaagtcttc taacatgcgg
     4741 tgacgtggag gagaatcccg gccctatgac cgagtacaag cccacggtgc gcctcgccac
     4801 ccgcgacgac gtccccaggg ccgtacgcac cctcgccgcc gcgttcgccg actaccccgc
     4861 cacgcgccac accgtcgatc cggaccgcca catcgagcgg gtcaccgagc tgcaagaact
     4921 cttcctcacg cgcgtcgggc tcgacatcgg caaggtgtgg gtcgcggacg acggcgccgc
     4981 ggtggcggtc tggaccacgc cggagagcgt cgaagcgggg gcggtgttcg ccgagatcgg
     5041 cccgcgcatg gccgagttga gcggttcccg gctggccgcg cagcaacaga tggaaggcct
     5101 cctggcgccg caccggccca aggagcccgc gtggttcctg gccaccgtcg gcgtctcgcc
     5161 cgaccaccag ggcaagggtc tgggcagcgc cgtcgtgctc cccggagtgg aggcggccga
     5221 gcgcgccggg gtgcccgcct tcctggagac ctccgcgccc cgcaacctcc ccttctacga
     5281 gcggctcggc ttcaccgtca ccgccgacgt cgaggtgccc gaaggaccgc gcacctggtg
     5341 catgacccgc aagcccggtg cctgaaatca acctctggat tacaaaattt gtgggtaaac
     5401 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
     5461 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat
     5521 catgtctgga attgactcaa atgatgtcaa ttagtctatc agaagctcat ctggtctccc
     5581 ttccggggga caagacatcc ctgtttaata tttaaacagc agtgttccca aactgggttc
     5641 ttatatccct tgctctggtc aaccaggttg cagggtttcc tgtcctcaca ggaacgaagt
     5701 ccctaaagaa acagtggcag ccaggtttag ccccggaatt gactggattc cttttttagg
     5761 gcccattggt atggcttttt ccccgtatcc ccccaggtgt ctgcaggctc aaagagcagc
     5821 gagaagcgtt cagaggaaag cgatcccgtg ccaccttccc cgtgcccggg ctgtccccgc
     5881 acgctgccgg ctcggggatg cggggggagc gccggaccgg agcggagccc cgggcggctc
     5941 gctgctgccc cctagcgggg gagggacgta attacatccc tgggggcttt gggggggggc
     6001 tgtccctgat atctataaca agaaaatata tatataataa gttatcacgt aagtagaaca
     6061 tgaaataaca atataattat cgtatgagtt aaatcttaaa agtcacgtaa aagataatca
     6121 tgcgtcattt tgactcacgc ggtcgttata gttcaaaatc agtgacactt accgcattga
     6181 caagcacgcc tcacgggagc tccaagcggc gactgagatg tcctaaatgc acagcgacgg
     6241 attcgcgcta tttagaaaga gagagcaata tttcaagaat gcatgcgtca attttacgca
     6301 gactatcttt ctagggttaa tctagctgca tcaggatcat atcgtcgggt cttttttccg
     6361 gctcagtcat cgcccaagct ggcgctatct gggcatcggg gaggaagaag cccgtgcctt
     6421 ttcccgcgag gttgaagcgg catggaaaga gtttgccgag gatgactgct gctgcattga
     6481 cgttgagcga aaacgcacgt ttaccatgat gattcgggaa ggtgtggcca tgcacgcctt
     6541 taacggtgaa ctgttcgttc aggccacctg ggataccagt tcgtcgcggc ttttccggac
     6601 acagttccgg atggtcagcc cgaagcgcat cagcaacccg aacaataccg gcgacagccg
     6661 gaactgccgt gccggtgtgc agattaatga cagcggtgcg gcgctgggat attacgtcag
     6721 cgaggacggg tatcctggct ggatgccgca gaaatggaca tggatacccc gtgagttacc
     6781 cggcgggcgc gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg
     6841 ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa
     6901 tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac
     6961 ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt
     7021 gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
     7081 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     7141 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     7201 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     7261 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     7321 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     7381 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     7441 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     7501 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     7561 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     7621 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag
     7681 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     7741 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     7801 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     7861 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     7921 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     7981 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     8041 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     8101 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     8161 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     8221 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     8281 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     8341 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     8401 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     8461 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     8521 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     8581 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     8641 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     8701 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     8761 gcaaaaacag ga
//
