LOCUS       sequence-490944-       10303 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express A2AR mutants in Pichia Pastoris.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10303)
  AUTHORS   Wang X, Neale C, Kim SK, Goddard WA, Ye L
  TITLE     Intermediate-state-trapped mutants pinpoint G protein-coupled 
            receptor conformational allostery.
  JOURNAL   Nat Commun. 2023 Mar 10;14(1):1325. doi: 10.1038/s41467-023-36971-6.
  PUBMED    36899002
REFERENCE   2  (bases 1 to 10303)
  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: "Nat 
            Commun."; date: "2023-03-10"; volume: "14(1):1325. doi"; pages: " 
            10.1038/s41467-023-36971-6"
COMMENT     Sequence Label: pPIC9K_A2AR_316_V229C_R291AR293A
FEATURES             Location/Qualifiers
     source          1..10303
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        2..937
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 promoter
                     /note="inducible promoter, regulated by methanol"
     CDS             join(949..1005,1006..1203)
                     /codon_start=1
                     /gene="MF-alpha-1"
                     /product="N-terminal secretion signal from S. cerevisiae 
                     alpha-factor"
                     /label=alpha-factor secretion signal
                     /note="Cleavage by the Kex2 protease occurs after the 
                     dibasic KR sequence."
                     /translation="MRFPSIFTAVLFAASSALAAPVNTTTEDETAQIPAEAVIGYSDLE
                     GDFDVAVLPFSNSTNNGLLFINTTIASIAAKEEGVSLEKR"
     CDS             1204..1227
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1267..1287
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             2236..2265
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     terminator      2356..2602
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 terminator
                     /note="transcription terminator for AOX1"
     CDS             complement(3015..5549)
                     /codon_start=1
                     /gene="Pichia pastoris HIS4"
                     /product="multifunctional enzyme, required for histidine 
                     biosynthesis"
                     /label=PpHIS4
                     /note="auxotrophic marker for Pichia pastoris"
                     /translation="MTFPLLPAYASVAEFDNSLSLVGKAVFPYAADQLHNLIKFTQSTE
                     LQVNVQVESSVTEDQFEELIDNLLKLYNNGINEVILDLDLAERVVQRMIPGARVIYRTL
                     VDKVASLPANASIAVPFSSPLGDLKSFTNGGSRTVYAFSETAKLVDVTSTVASGIIPII
                     DARQLTTEYELSEDVKKFPVSEILLASLTTDRPDGLFTTLVADSSNYSLGLVYSSKKSI
                     PEAIRTQTGVYQSRRHGLWYKGATSGATQKLLGIELDCDGDCLKFVVEQTGVGFCHLER
                     TSCFGQSKGLRAMEATLWDRKSNAPEGSYTKRLFDDEVLLNAKIREEADELAEAKSKED
                     IAWECADLFYFALVRCAKYGVTLDEVERNLDMKSLKVTRRKGDAKPGYTKEQPKEESKP
                     KEVPSEGRIELCKIDVSKASSQEIEDALRRPIQKTEQIMELVKPIVDNVRQNGDKALLE
                     LTAKFDGVALKTPVLEAPFPEELMQLPDNVKRAIDLSIDNVRKFHEAQLTETLQVETCP
                     GVVCSRFARPIEKVGLYIPGGTAILPSTSLMLGVPAKVAGCKEIVFASPPKKDGTLTPE
                     VIYVAHKVGAKCIVLAGGAQAVAAMAYGTETVPKCDKIFGPGNQFVTAAKMMVQNDTSA
                     LCSIDMPAGPSEVLVIADKYADPDFVASDLLSQAEHGIDSQVILLAVDMTDKELARIED
                     AVHNQAVQLPRVEIVRKCIAHSTTLSVATYEQALEMSNQYAPEHLILQIENASSYVDQV
                     QHAGSVFVGAYSPESCGDYSSGTNHTLPTYGYARQYSGVNTATFQKFITSQDVTPEGLK
                     HIGQAVMDLAAVEGLDAHRNAVKVRMEKLGLI"
     CDS             complement(5962..6777)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(5962..6777)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     6685..6704
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     6769..6788
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(6942..6961)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     misc_feature    7156..7904
                     /label=AOX1 3' fragment
                     /note="region downstream of Pichia pastoris AOX1 gene"
     primer_bind     complement(7940..7962)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    8048..8188
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     8103..8122
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(8203..8220)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(8374..8962)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(8454..8473)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(9133..9924,9925..9993))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     9756..9775
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(9994..10098)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     10166..10184
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 agatctaaca tccaaagacg aaaggttgaa tgaaaccttt ttgccatccg acatccacag
       61 gtccattctc acacataagt gccaaacgca acaggagggg atacactagc agcagaccgt
      121 tgcaaacgca ggacctccac tcctcttctc ctcaacaccc acttttgcca tcgaaaaacc
      181 agcccagtta ttgggcttga ttggagctcg ctcattccaa ttccttctat taggctacta
      241 acaccatgac tttattagcc tgtctatcct ggcccccctg gcgaggttca tgtttgttta
      301 tttccgaatg caacaagctc cgcattacac ccgaacatca ctccagatga gggctttctg
      361 agtgtggggt caaatagttt catgttcccc aaatggccca aaactgacag tttaaacgct
      421 gtcttggaac ctaatatgac aaaagcgtga tctcatccaa gatgaactaa gtttggttcg
      481 ttgaaatgct aacggccagt tggtcaaaaa gaaacttcca aaagtcggca taccgtttgt
      541 cttgtttggt attgattgac gaatgctcaa aaataatctc attaatgctt agcgcagtct
      601 ctctatcgct tctgaacccc ggtgcacctg tgccgaaacg caaatgggga aacacccgct
      661 ttttggatga ttatgcattg tctccacatt gtatgcttcc aagattctgg tgggaatact
      721 gctgatagcc taacgttcat gatcaaaatt taactgttct aacccctact tgacagcaat
      781 atataaacag aaggaagctg ccctgtctta aacctttttt tttatcatca ttattagctt
      841 actttcataa ttgcgactgg ttccaattga caagcttttg attttaacga cttttaacga
      901 caacttgaga agatcaaaaa acaactaatt attcgaagga tccaaacgat gagatttcct
      961 tcaattttta ctgcagtttt attcgcagca tcctccgcat tagctgctcc agtcaacact
     1021 acaacagaag atgaaacggc acaaattccg gctgaagctg tcatcggtta ctcagattta
     1081 gaaggggatt tcgatgttgc tgttttgcca ttttccaaca gcacaaataa cgggttattg
     1141 tttataaata ctactattgc cagcattgct gctaaagaag aaggggtatc tctcgaaaaa
     1201 agagactaca aggacgatga cgataagtct aacaacaaca acaacaacaa caacaacaac
     1261 cttggcgaaa acttgtattt ccagggcgct cccatcatgg gctcctcggt gtacatcacg
     1321 gtggagctgg ccattgctgt gctggccatc ctgggcaatg tgctggtgtg ctgggccgtg
     1381 tggctcaaca gcaacctgca gaacgtcacc aactactttg tggtgtcact ggcggcggcc
     1441 gacatcgcag tgggtgtgct cgccatcccc tttgccatca ccatcagcac cgggttctgc
     1501 gctgcctgcc acggctgcct cttcattgcc tgcttcgtcc tggtcctcac gcagagctcc
     1561 atcttcagtc tcctggccat cgccattgac cgctacattg ccatccgcat cccgctccgg
     1621 tacaatggct tggtgaccgg cacgagggct aagggcatca ttgccatctg ctgggtgctg
     1681 tcgtttgcca tcggcctgac tcccatgcta ggttggaaca actgcggtca gccaaaggag
     1741 ggcaagaacc actcccaggg ctgcggggag ggccaagtgg cctgtctctt tgaggatgtg
     1801 gtccccatga actacatggt gtacttcaac ttctttgcct gtgtgctggt gcccctgctg
     1861 ctcatgctgg gtgtctattt gcggatcttc ctggcggcgc gacgacagct gaagcagatg
     1921 gagagccagc ctctgccggg ggagcgggca cggtccacac tgcagaagga gtgccatgct
     1981 gccaagtcac tggccatcat tgtggggctc tttgccctct gctggctgcc cctacacatc
     2041 atcaactgct tcactttctt ctgccccgac tgcagccacg cccctctctg gctcatgtac
     2101 ctggccatcg tcctctccca caccaattcg gttgtgaatc ccttcattta cgcctacgct
     2161 atcgccgagt tccgccagac cttccgcaag atcattcgca gccacgtcct gaggcagcaa
     2221 gaacctttca aggcacatca ccatcaccat caccatcacc atcactgagc ggccgcgaat
     2281 taattcgcct tagacatgac tgttcctcag ttcaagttgg gcacttacga gaagaccggt
     2341 cttgctagat tctaatcaag aggatgtcag aatgccattt gcctgagaga tgcaggcttc
     2401 atttttgata cttttttatt tgtaacctat atagtatagg attttttttg tcattttgtt
     2461 tcttctcgta cgagcttgct cctgatcagc ctatctcgca gctgatgaat atcttgtggt
     2521 aggggtttgg gaaaatcatt cgagtttgat gtttttcttg gtatttccca ctcctcttca
     2581 gagtacagaa gattaagtga gacgttcgtt tgtgcaagct tatcgataag ctttaatgcg
     2641 gtagtttatc acagttaaat tgctaacgca gtcaggcacc gtgtatgaaa tctaacaatg
     2701 cgctcatcgt catcctcggc accgtcaccc tggatgctgt aggcataggc ttggttatgc
     2761 cggtactgcc gggcctcttg cgggatatcg tccattccga cagcatcgcc agtcactatg
     2821 gcgtgctgct agcgctatat gcgttgatgc aatttctatg cgcacccgtt ctcggagcac
     2881 tgtccgaccg ctttggccgc cgcccagtcc tgctcgcttc gctacttgga gccactatcg
     2941 actacgcgat catggcgacc acacccgtcc tgtggatcta tcgaatctaa atgtaagtta
     3001 aaatctctaa ataattaaat aagtcccagt ttctccatac gaaccttaac agcattgcgg
     3061 tgagcatcta gaccttcaac agcagccaga tccatcactg cttggccaat atgtttcagt
     3121 ccctcaggag ttacgtcttg tgaagtgatg aacttctgga aggttgcagt gttaactccg
     3181 ctgtattgac gggcatatcc gtacgttggc aaagtgtggt tggtaccgga ggagtaatct
     3241 ccacaactct ctggagagta ggcaccaaca aacacagatc cagcgtgttg tacttgatca
     3301 acataagaag aagcattctc gatttgcagg atcaagtgtt caggagcgta ctgattggac
     3361 atttccaaag cctgctcgta ggttgcaacc gatagggttg tagagtgtgc aatacacttg
     3421 cgtacaattt caacccttgg caactgcaca gcttggttgt gaacagcatc ttcaattctg
     3481 gcaagctcct tgtctgtcat atcgacagcc aacagaatca cctgggaatc aataccatgt
     3541 tcagcttgag acagaaggtc tgaggcaacg aaatctggat cagcgtattt atcagcaata
     3601 actagaactt cagaaggccc agcaggcatg tcaatactac acagggctga tgtgtcattt
     3661 tgaaccatca tcttggcagc agtaacgaac tggtttcctg gaccaaatat tttgtcacac
     3721 ttaggaacag tttctgttcc gtaagccata gcagctactg cctgggcgcc tcctgctagc
     3781 acgatacact tagcaccaac cttgtgggca acgtagatga cttctggggt aagggtacca
     3841 tccttcttag gtggagatgc aaaaacaatt tctttgcaac cagcaacttt ggcaggaaca
     3901 cccagcatca gggaagtgga aggcagaatt gcggttccac caggaatata gaggccaact
     3961 ttctcaatag gtcttgcaaa acgagagcag actacaccag ggcaagtctc aacttgcaac
     4021 gtctccgtta gttgagcttc atggaatttc ctgacgttat ctatagagag atcaatggct
     4081 ctcttaacgt tatctggcaa ttgcataagt tcctctggga aaggagcttc taacacaggt
     4141 gtcttcaaag cgactccatc aaacttggca gttagttcta aaagggcttt gtcaccattt
     4201 tgacgaacat tgtcgacaat tggtttgact aattccataa tctgttccgt tttctggata
     4261 ggacgacgaa gggcatcttc aatttcttgt gaggaggcct tagaaacgtc aattttgcac
     4321 aattcaatac gaccttcaga agggacttct ttaggtttgg attcttcttt aggttgttcc
     4381 ttggtgtatc ctggcttggc atctcctttc cttctagtga cctttaggga cttcatatcc
     4441 aggtttctct ccacctcgtc caacgtcaca ccgtacttgg cacatctaac taatgcaaaa
     4501 taaaataagt cagcacattc ccaggctata tcttccttgg atttagcttc tgcaagttca
     4561 tcagcttcct ccctaatttt agcgttcaac aaaacttcgt cgtcaaataa ccgtttggta
     4621 taagaacctt ctggagcatt gctcttacga tcccacaagg tggcttccat ggctctaaga
     4681 ccctttgatt ggccaaaaca ggaagtgcgt tccaagtgac agaaaccaac acctgtttgt
     4741 tcaaccacaa atttcaagca gtctccatca caatccaatt cgatacccag caacttttga
     4801 gttgctccag atgtagcacc tttataccac aaaccgtgac gacgagattg gtagactcca
     4861 gtttgtgtcc ttatagcctc cggaatagac tttttggacg agtacaccag gcccaacgag
     4921 taattagaag agtcagccac caaagtagtg aatagaccat cggggcggtc agtagtcaaa
     4981 gacgccaaca aaatttcact gacagggaac tttttgacat cttcagaaag ttcgtattca
     5041 gtagtcaatt gccgagcatc aataatgggg attataccag aagcaacagt ggaagtcaca
     5101 tctaccaact ttgcggtctc agaaaaagca taaacagttc tactaccgcc attagtgaaa
     5161 cttttcaaat cgcccagtgg agaagaaaaa ggcacagcga tactagcatt agcgggcaag
     5221 gatgcaactt tatcaaccag ggtcctatag ataaccctag cgcctgggat catcctttgg
     5281 acaactcttt ctgccaaatc taggtccaaa atcacttcat tgataccatt attgtacaac
     5341 ttgagcaagt tgtcgatcag ctcctcaaat tggtcctctg taacggatga ctcaacttgc
     5401 acattaactt gaagctcagt cgattgagtg aacttgatca ggttgtgcag ctggtcagca
     5461 gcatagggaa acacggcttt tcctaccaaa ctcaaggaat tatcaaactc tgcaacactt
     5521 gcgtatgcag gtagcaaggg aaatgtcata cttgaagtcg gacagtgagt gtagtcttga
     5581 gaaattctga agccgtattt ttattatcag tgagtcagtc atcaggagat cctctacgcc
     5641 ggacgcatcg tggccgacct gcaggggggg gggggcgctg aggtctgcct cgtgaagaag
     5701 gtgttgctga ctcataccag gcctgaatcg ccccatcatc cagccagaaa gtgagggagc
     5761 cacggttgat gagagctttg ttgtaggtgg accagttggt gattttgaac ttttgctttg
     5821 ccacggaacg gtctgcgttg tcgggaagat gcgtgatctg atccttcaac tcagcaaaag
     5881 ttcgatttat tcaacaaagc cgccgtcccg tcaagtcagc gtaatgctct gccagtgtta
     5941 caaccaatta accaattctg attagaaaaa ctcatcgagc atcaaatgaa actgcaattt
     6001 attcatatca ggattatcaa taccatattt ttgaaaaagc cgtttctgta atgaaggaga
     6061 aaactcaccg aggcagttcc ataggatggc aagatcctgg tatcggtctg cgattccgac
     6121 tcgtccaaca tcaatacaac ctattaattt cccctcgtca aaaataaggt tatcaagtga
     6181 gaaatcacca tgagtgacga ctgaatccgg tgagaatggc aaaagcttat gcatttcttt
     6241 ccagacttgt tcaacaggcc agccattacg ctcgtcatca aaatcactcg catcaaccaa
     6301 accgttattc attcgtgatt gcgcctgagc gagacgaaat acgcgatcgc tgttaaaagg
     6361 acaattacaa acaggaatcg aatgcaaccg gcgcaggaac actgccagcg catcaacaat
     6421 attttcacct gaatcaggat attcttctaa tacctggaat gctgttttcc cggggatcgc
     6481 agtggtgagt aaccatgcat catcaggagt acggataaaa tgcttgatgg tcggaagagg
     6541 cataaattcc gtcagccagt ttagtctgac catctcatct gtaacatcat tggcaacgct
     6601 acctttgcca tgtttcagaa acaactctgg cgcatcgggc ttcccataca atcgatagat
     6661 tgtcgcacct gattgcccga cattatcgcg agcccattta tacccatata aatcagcatc
     6721 catgttggaa tttaatcgcg gcctcgagca agacgtttcc cgttgaatat ggctcataac
     6781 accccttgta ttactgttta tgtaagcaga cagttttatt gttcatgatg atatattttt
     6841 atcttgtgca atgtaacatc agagattttg agacacaacg tggctttccc ccccccccct
     6901 gcaggtcggc atcaccggcg ccacaggtgc ggttgctggc gcctatatcg ccgacatcac
     6961 cgatggggaa gatcgggctc gccacttcgg gctcatgagc gcttgtttcg gcgtgggtat
     7021 ggtggcaggc cccgtggccg ggggactgtt gggcgccatc tccttgcatg caccattcct
     7081 tgcggcggcg gtgctcaacg gcctcaacct actactgggc tgcttcctaa tgcaggagtc
     7141 gcataaggga gagcgtcgag tatctatgat tggaagtatg ggaatggtga tacccgcatt
     7201 cttcagtgtc ttgaggtctc ctatcagatt atgcccaact aaagcaaccg gaggaggaga
     7261 tttcatggta aatttctctg acttttggtc atcagtagac tcgaactgtg agactatctc
     7321 ggttatgaca gcagaaatgt ccttcttgga gacagtaaat gaagtcccac caataaagaa
     7381 atccttgtta tcaggaacaa acttcttgtt tcgaactttt tcggtgcctt gaactataaa
     7441 atgtagagtg gatatgtcgg gtaggaatgg agcgggcaaa tgcttacctt ctggaccttc
     7501 aagaggtatg tagggtttgt agatactgat gccaacttca gtgacaacgt tgctatttcg
     7561 ttcaaaccat tccgaatcca gagaaatcaa agttgtttgt ctactattga tccaagccag
     7621 tgcggtcttg aaactgacaa tagtgtgctc gtgttttgag gtcatctttg tatgaataaa
     7681 tctagtcttt gatctaaata atcttgacga gccagacgat aataccaatc taaactcttt
     7741 aaacgttaaa ggacaagtat gtctgcctgt attaaacccc aaatcagctc gtagtctgat
     7801 cctcatcaac ttgaggggca ctatcttgtt ttagagaaat ttgcggagat gcgatatcga
     7861 gaaaaaggta cgctgatttt aaacgtgaaa tttatctcaa gatctgctgc ctcgcgcgtt
     7921 tcggtgatga cggtgaaaac ctctgacaca tgcagctccc ggagacggtc acagcttgtc
     7981 tgtaagcgga tgccgggagc agacaagccc gtcagggcgc gtcagcgggt gttggcgggt
     8041 gtcggggcgc agccatgacc cagtcacgta gcgatagcgg agtgtatact ggcttaacta
     8101 tgcggcatca gagcagattg tactgagagt gcaccatatg cggtgtgaaa taccgcacag
     8161 atgcgtaagg agaaaatacc gcatcaggcg ctcttccgct tcctcgctca ctgactcgct
     8221 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     8281 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     8341 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     8401 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     8461 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     8521 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     8581 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     8641 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     8701 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     8761 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt
     8821 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     8881 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     8941 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     9001 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     9061 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     9121 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     9181 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     9241 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     9301 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     9361 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     9421 tagtttgcgc aacgttgttg ccattgctgc aggcatcgtg gtgtcacgct cgtcgtttgg
     9481 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     9541 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     9601 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     9661 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     9721 gcgaccgagt tgctcttgcc cggcgtcaac acgggataat accgcgccac atagcagaac
     9781 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     9841 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     9901 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     9961 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
    10021 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
    10081 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat
    10141 tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc gtcttcaaga
    10201 attaattctc atgtttgaca gcttatcatc gataagctga ctcatgttgg tattgtgaaa
    10261 tagacgcaga tcgggaacac tgaaaaataa cagttattat tcg
//
