LOCUS       sequence-338951-       10865 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Optimized EKAREV FRET biosensor sensor for ERK.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10865)
  AUTHORS   Ponsioen B, Post JB, Buissant des Amorie JR, Laskaris D, van Ineveld
            RL, Kersten S, Bertotti A, Sassi F, Sipieter F, Cappe B, Mertens S, 
            Verlaan-Klink I, Boj SF, Vries RGJ, Rehmann H, Vandenabeele P, 
            Riquet FB, Trusolino L, Bos JL, Snippert HJG
  TITLE     Quantifying single-cell ERK dynamics in colorectal cancer organoids 
            reveals EGFR as an amplifier of oncogenic MAPK pathway signalling.
  JOURNAL   Nat Cell Biol. 2021 Apr;23(4):377-390. doi: 
            10.1038/s41556-021-00654-5. Epub 2021 Apr 1.
  PUBMED    33795873
REFERENCE   2  (bases 1 to 10865)
  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"; doi: 
            "10.1038/s41556-021-00654-5"; journalName: "Nat Cell Biol"; date: 
            "2021-04"; volume: "23"; issue: "4"; pages: "377-390"
COMMENT     Sequence Label: pLV x EF1a_EKAREN5-ires-puro
FEATURES             Location/Qualifiers
     source          1..10865
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1..21
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     misc_feature    229..781
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(373..390)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     600..619
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             808..1407
                     /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(808..827)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1304..1324
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             2299..2532
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    2604..2725
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2641..2660)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2695..2714
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      2765..2900
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2920..2939)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2921..2939)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2949..2966)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2949..2965)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2958..2980)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3107..3562
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3194..3213)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3404..3425
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3588..3692
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3693..3761,3762..4553)
                     /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(3911..3930)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4724..5312
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5213..5232
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5466..5483
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5600..5621
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5636..5641,5642..5659,5660..5666)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5674..5690
                     /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     5679..5701
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5698..5714
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5698..5714
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     5733..5753
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        5735..5753
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        5781..6007
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             6008..6188
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    6235..6360
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    6853..7086
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             7271..7315
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    7577..7694
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        7711..8873
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          7942..8864
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     8821..8841
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             join(8964..8966,8967..8969,8970..9677)
                     /codon_start=1
                     /product="yellow fluorescent protein (YFP) with fast and 
                     efficient maturation (Nagai et al., 2002)"
                     /label=Venus
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLLCTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALFKDPNEKRDHMVLL
                     EFLTAAGITEGMNELY"
     primer_bind     complement(9270..9289)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
ORIGIN
        1 cctaagaaga agcgtaaggt gtaagtcgac ggtcgagtcg agacgctagc ctagtgccat
       61 ttgttcagtg gttcgtaggg ctttccccca ctgtttggct ttcagttata tggatgatgt
      121 ggtattgggg gccaagtctg tacagcatct tgagtccctt tttaccgctg ttaccaattt
      181 tcttttgtct ttgggtaaac ccgggaattc cgcccctctc cccccctaac gttactggcc
      241 gaagccgctt ggaataaggc cggtgtgcgt ttgtctatat gttattttcc accatattgc
      301 cgtcttttgg caatgtgagg gcccggaaac ctggccctgt cttcttgacg agcattccta
      361 ggggtctttc ccctctcgcc aaaggaatgc aaggtctgtt gaatgtcgtg aaggaagcag
      421 ttcctctgga agcttcttga agacaaacaa cgtctgtagc gaccctttgc aggcagcgga
      481 accccccacc tggcgacagg tgcctctgcg gccaaaagcc acgtgtataa gatacacctg
      541 caaaggcggc acaaccccag tgccacgttg tgagttggat agttgtggaa agagtcaaat
      601 ggctctcctc aagcgtattc aacaaggggc tgaaggatgc ccagaaggta ccccattgta
      661 tgggatctga tctggggcct cggtgcacat gctttacatg tgtttagtcg aggttaaaaa
      721 aacgtctagg ccccccgaac cacggggacg tggttttcct ttgaaaaaca cgatgataat
      781 atggatatgg ccacaacggt cgccaccatg accgagtaca agcccacggt gcgcctcgcc
      841 acccgcgacg acgtccccag ggccgtacgc accctcgccg ccgcgttcgc cgactacccc
      901 gccacgcgcc acaccgtcga tccggaccgc cacatcgagc gggtcaccga gctgcaagaa
      961 ctcttcctca cgcgcgtcgg gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc
     1021 gcggtggcgg tctggaccac gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc
     1081 ggcccgcgca tggccgagtt gagcggttcc cggctggccg cgcagcaaca gatggaaggc
     1141 ctcctggcgc cgcaccggcc caaggagccc gcgtggttcc tggccaccgt cggcgtctcg
     1201 cccgaccacc agggcaaggg tctgggcagc gccgtcgtgc tccccggagt ggaggcggcc
     1261 gagcgcgccg gggtgcccgc cttcctggag acctccgcgc cccgcaacct ccccttctac
     1321 gagcggctcg gcttcaccgt caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg
     1381 tgcatgaccc gcaagcccgg tgcctgacgc ccgccccacg acccgcagcg cccgaccgaa
     1441 aggagcgcac gaccccatgc atgtattcaa tctaagcagg ctttcacttt ctcgccaact
     1501 tacaaggcct ttctgtgtaa acaatacctg aacctttacc ccgttgcccg gcaacggcca
     1561 ggtctgtgcc aagtgtttgc tgacgcaacc cccactggct ggggcttggt catgggccat
     1621 cagcgcatgc gtggaacctt ttcggctcct ctgccgatcc atactgcgga actcctagcc
     1681 gcttgttttg ctcgcagcag gtctggagca aacattatcg ggactgataa ctctgttgtc
     1741 ctatcccgca aatatacatc gtttccatgg ctgctaggct gtgctgccaa ctggatcctg
     1801 cgcgggacgt cctttgttta cgtcccgtcg gcgctgaatc ctgcggacga cccttctcgg
     1861 ggtcgcttgg gactctctcg tccccttctc cgtctgccgt tccgaccgac cacggggcgc
     1921 acctctcttt acgcggactc cccgtctgtg ccttctcatc tgccggaccg tgtgcacttc
     1981 gcttcacctc tgcacgtcgc atggagacca ccgtgaacgc ccaccaaata ttgcccaagg
     2041 tcttacataa gaggactctt ggactctcag caatgtcaac gaccgacctt gaggcatact
     2101 tcaaagactg tttgtttaaa gactgggagg agttggggga ggagattagg ttaaaggtct
     2161 ttgtactagg aggctgtagg cataaattgg tctgcgcacc agcaccatgg cgcaatcact
     2221 agagcggggt acctttaaga ccaatgactt acaaggcagc tgtagatctt agccactttt
     2281 taaaagaaaa ggggggactg gaagggctaa ttcactccca acgaagacaa gatctgcttt
     2341 ttgcttgtac tgggtctctc tggttagacc agatctgagc ctgggagctc tctggctaac
     2401 tagggaaccc actgcttaag cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg
     2461 cccgtctgtt gtgtgactct ggtaactaga gatccctcag acccttttag tcagtgtgga
     2521 aaatctctag cagtagtagt tcatgtcatc ttattattca gtatttataa cttgcaaaga
     2581 aatgaatatc agagagtgag aggaacttgt ttattgcagc ttataatggt tacaaataaa
     2641 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
     2701 tgtccaaact catcaatgta tcttatcatg tctggctcta gctatcccgc ccctaactcc
     2761 gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat
     2821 tttttttatt tatgcagagg ccgaggccgc ctcggcctct gagctattcc agaagtagtg
     2881 aggaggcttt tttggaggcc tagggacgta cccaattcgc cctatagtga gtcgtattac
     2941 gcgcgctcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg cgttacccaa
     3001 cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga agaggcccgc
     3061 accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatgggacgc gccctgtagc
     3121 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     3181 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     3241 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
     3301 ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag
     3361 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
     3421 actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg
     3481 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac
     3541 aaaatattaa cgcttacaat ttaggtggca cttttcgggg aaatgtgcgc ggaaccccta
     3601 tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat
     3661 aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc
     3721 ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga
     3781 aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca
     3841 acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt
     3901 ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg
     3961 gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc
     4021 atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata
     4081 acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt
     4141 tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag
     4201 ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca
     4261 aactattaac tggcgaacta cttactctag cttcccggca acaattaata gactggatgg
     4321 aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg
     4381 ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag
     4441 atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg
     4501 aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag
     4561 accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga
     4621 tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt
     4681 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc
     4741 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc
     4801 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac
     4861 caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac
     4921 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt
     4981 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct
     5041 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat
     5101 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt
     5161 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg
     5221 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt
     5281 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt
     5341 tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg
     5401 tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg
     5461 agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa ccgcctctcc
     5521 ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac tggaaagcgg
     5581 gcagtgagcg caacgcaatt aatgtgagtt agctcactca ttaggcaccc caggctttac
     5641 actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa tttcacacag
     5701 gaaacagcta tgaccatgat tacgccaagc gcgcaattaa ccctcactaa agggaacaaa
     5761 agctggagct gcaagcttaa tgtagtctta tgcaatactc ttgtagtctt gcaacatggt
     5821 aacgatgagt tagcaacatg ccttacaagg agagaaaaag caccgtgcat gccgattggt
     5881 ggaagtaagg tggtacgatc gtgccttatt aggaaggcaa cagacgggtc tgacatggat
     5941 tggacgaacc actgaattgc cgcattgcag agatattgta tttaagtgcc tagctcgata
     6001 cataaacggg tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg
     6061 gaacccactg cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg
     6121 tctgttgtgt gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat
     6181 ctctagcagt ggcgcccgaa cagggacttg aaagcgaaag ggaaaccaga ggagctctct
     6241 cgacgcagga ctcggcttgc tgaagcgcgc acggcaagag gcgaggggcg gcgactggtg
     6301 agtacgccaa aaattttgac tagcggaggc tagaaggaga gagatgggtg cgagagcgtc
     6361 agtattaagc gggggagaat tagatcgcga tgggaaaaaa ttcggttaag gccaggggga
     6421 aagaaaaaat ataaattaaa acatatagta tgggcaagca gggagctaga acgattcgca
     6481 gttaatcctg gcctgttaga aacatcagaa ggctgtagac aaatactggg acagctacaa
     6541 ccatcccttc agacaggatc agaagaactt agatcattat ataatacagt agcaaccctc
     6601 tattgtgtgc atcaaaggat agagataaaa gacaccaagg aagctttaga caagatagag
     6661 gaagagcaaa acaaaagtaa gaccaccgca cagcaagcgg ccgctgatct tcagacctgg
     6721 aggaggagat atgagggaca attggagaag tgaattatat aaatataaag tagtaaaaat
     6781 tgaaccatta ggagtagcac ccaccaaggc aaagagaaga gtggtgcaga gagaaaaaag
     6841 agcagtggga ataggagctt tgttccttgg gttcttggga gcagcaggaa gcactatggg
     6901 cgcagcgtca atgacgctga cggtacaggc cagacaatta ttgtctggta tagtgcagca
     6961 gcagaacaat ttgctgaggg ctattgaggc gcaacagcat ctgttgcaac tcacagtctg
     7021 gggcatcaag cagctccagg caagaatcct ggctgtggaa agatacctaa aggatcaaca
     7081 gctcctgggg atttggggtt gctctggaaa actcatttgc accactgctg tgccttggaa
     7141 tgctagttgg agtaataaat ctctggaaca gatttggaat cacacgacct ggatggagtg
     7201 ggacagagaa attaacaatt acacaagctt aatacactcc ttaattgaag aatcgcaaaa
     7261 ccagcaagaa aagaatgaac aagaattatt ggaattagat aaatgggcaa gtttgtggaa
     7321 ttggtttaac ataacaaatt ggctgtggta tataaaatta ttcataatga tagtaggagg
     7381 cttggtaggt ttaagaatag tttttgctgt actttctata gtgaatagag ttaggcaggg
     7441 atattcacca ttatcgtttc agacccacct cccaaccccg aggggacccg acaggcccga
     7501 aggaatagaa gaagaaggtg gagagagaga cagagacaga tccattcgat tagtgaacgg
     7561 atctcgacgg tatcggtttt aaaagaaaag gggggattgg ggggtacagt gcaggggaaa
     7621 gaatagtaga cataatagca acagacatac aaactaaaga attacaaaaa caaattacaa
     7681 aaattcaaaa ttttatcgat aagcttgggt ggctccggtg cccgtcagtg ggcagagcgc
     7741 acatcgccca cagtccccga gaagttgggg ggaggggtcg gcaattgaac cggtgcctag
     7801 agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt actggctccg cctttttccc
     7861 gagggtgggg gagaaccgta tataagtgca gtagtcgccg tgaacgttct ttttcgcaac
     7921 gggtttgccg ccagaacaca ggtaagtgcc gtgtgtggtt cccgcgggcc tggcctcttt
     7981 acgggttatg gcccttgcgt gccttgaatt acttccacct ggctccagta cgtgattctt
     8041 gatcccgagc tggagccagg ggcgggcctt gcgctttagg agccccttcg cctcgtgctt
     8101 gagttgaggc ctggcctggg cgctggggcc gccgcgtgcg aatctggtgg caccttcgcg
     8161 cctgtctcgc tgctttcgat aagtctctag ccatttaaaa tttttgatga cctgctgcga
     8221 cgcttttttt ctggcaagat agtcttgtaa atgcgggcca ggatctgcac actggtattt
     8281 cggtttttgg ggccgcgggc ggcgacgggg cccgtgcgtc ccagcgcaca tgttcggcga
     8341 ggcggggcct gcgagcgcgg ccaccgagaa tcggacgggg gtagtctcaa gctggccggc
     8401 ctgctctggt gcctggcctc gcgccgccgt gtatcgcccc gccctgggcg gcaaggctgg
     8461 cccggtcggc accagttgcg tgagcggaaa gatggccgct tcccggccct gctccagggg
     8521 gctcaaaatg gaggacgcgg cgctcgggag agcgggcggg tgagtcaccc acacaaagga
     8581 aaggggcctt tccgtcctca gccgtcgctt catgtgactc cacggagtac cgggcgccgt
     8641 ccaggcacct cgattagttc tggagctttt ggagtacgtc gtctttaggt tggggggagg
     8701 ggttttatgc gatggagttt ccccacactg agtgggtgga gactgaagtt aggccagctt
     8761 ggcacttgat gtaattctcc ttggaatttg ccctttttga gtttggatct tggttcattc
     8821 tcaagcctca gacagtggtt caaagttttt ttcttccatt tcaggtgtcg tgaggatccc
     8881 cgggcgcgcc acgcgtaccg atagcttcga attctgcagt cgacgatatc actagtctcg
     8941 agcattttgg caaagaattc ggcatggtga gcaagggcga ggagctgttc accggggtgg
     9001 tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg
     9061 agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gcttctatgc accaccggca
     9121 agctgcccgt gccctggccc accctcgtga ccaccctggg ctacggcctg cagtgcttcg
     9181 cccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct
     9241 acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg
     9301 tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg
     9361 aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac aacgtctata
     9421 tcaccgccga caagcagaag aacggcatca aggccaactt caagatccgc cacaacatcg
     9481 aggacggcgg cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc
     9541 ccgtgctgct gcccgacaac cactacctga gctaccagtc cgccctgttc aaagacccca
     9601 acgagaagcg cgatcacatg gtcctgctgg agttcctgac cgccgccggg atcactgagg
     9661 gcatgaacga gctgtacctc gagatggcgg acgaggagaa gctgccgccc ggctgggaga
     9721 agcgcatgag ccgcagctca ggccgagtgt actacttcaa ccacatcact aacgccagcc
     9781 agtgggagcg gcccagcggc aacagcagca gtggtggcaa aaacgggcag ggggagcctg
     9841 ccaggggtac cagtgctggt ggtagtgctg gtggtagtgc tggtggttcc ggcagtgctg
     9901 gtggtagtgc tggtggtagt accagtgctg gtggtagtgc tggtggtagt gctggtggta
     9961 gtgctggtgg tagtgctggt ggttccggca gtgctggtgg tagtgctggt ggtagtacca
    10021 gtgctggtgg tagtgctggt ggtagtgctg gtggtagtgc tggtggtagt gctggtggtt
    10081 ccggaccaga tgtccctaga actccagtgg atccagcatg gctgtcattc caatttccgg
    10141 gcggccgcat ggtgagtaaa ggggaagaac tgtttactgg cgtggtgcca attctggtcg
    10201 aactggatgg agatgtaaat gggcataaat ttagtgtgtc tggagaaggg gaaggagacg
    10261 ctacatatgg gaaactgaca ctgaaattta tttgtactac tggaaaactg cctgtgcctt
    10321 ggcctacact cgtgacaacc ctgtcttggg gagtgcagtg ttttgcccgg tatcctgatc
    10381 atatgaaaca gcatgatttt ttcaaatctg ccatgcctga gggatatgtc caggaacgga
    10441 caattttctt taaagatgat gggaattata aaacacgggc cgaagtgaaa tttgaagggg
    10501 atacactggt gaatcggatt gaactgaaag gaattgattt taaagaagat gggaatattc
    10561 tgggccataa actggaatat aattatttca gcgataatgt ctacattaca gccgataaac
    10621 agaagaatgg gattaaagcc aattttaaaa ttcggcataa tattgaagat ggaggggtgc
    10681 agctcgccga tcattatcag cagaatacac ctattgggga tgggcctgtg ctgctgcctg
    10741 ataatcatta tctgagcaca cagtctgccc tgagcaagga ccctaatgaa aagcgggacc
    10801 atatggtcct gctggaattt ctgactgccg ccggcattac actcgggatg gatgaactgt
    10861 ctaga
//
