LOCUS       sequence-442483-       11967 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Inducible expression of SARS-CoV-2 Spike protein with C-terminal 3x 
            FLAG tag under TRE3G promoter and cloned into a 2nd generation 
            lentiviral transfer plasmid (modified pLVX with Hygro resistance)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11967)
  AUTHORS   Gunnels TF, Stranford DM, Mitrut RE, Kamat NP, Leonard JN
  TITLE     Elucidating Design Principles for Engineering Cell-Derived Vesicles 
            to Inhibit SARS-CoV-2 Infection.
  JOURNAL   Small. 2022 May;18(19):e2200125. doi: 10.1002/smll.202200125. Epub 
            2022 Apr 7.
  PUBMED    35388947
REFERENCE   2  (bases 1 to 11967)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Small."; 
            date: "2022-05"; volume: "18(19):e2200125. doi"; pages: " 
            10.1002/smll.202200125. Epub 2022 Apr 7"
COMMENT     Sequence Label: pTFG005_Spike_Inducible_3xFlag
FEATURES             Location/Qualifiers
     source          1..11967
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             307..487
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    534..659
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1156..1389
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1574..1618
                     /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    1881..1996
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1 (lacking the first T)"
     promoter        2059..2408
                     /label=TRE3GV promoter
                     /note="3rd-generation Tet-responsive promoter that can be 
                     activated by binding of Tet-On(R) 3G, optimized for 
                     retroviral and lentiviral vectors"
     protein_bind    2067..2085
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2103..2121
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2139..2157
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2175..2193
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2211..2229
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2247..2265
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2283..2301
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             6361..6426
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     promoter        6452..6951
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(6487..6510)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     6884..6903
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             6980..8005
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             6980..8005
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     misc_feature    8029..8617
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8082..8102)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8500..8511)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             9106..9363
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(9491..9507)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(9491..9507)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(9504..9526)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    9515..9531
                     /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(9539..9545,9546..9563,9564..9569))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    9584..9605
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(9722..9739)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(9893..10481)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(9973..9992)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(10652..11443,11444..11512))
                     /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     11275..11294
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(11513..11617)
                     /gene="bla"
                     /label=AmpR promoter
     polyA_signal    11665..11799
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(11702..11721)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     11756..11775
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
ORIGIN
        1 ggaagggcta attcactccc aaagaagaca agatatcctt gatctgtgga tctaccacac
       61 acaaggctac ttccctgatt agcagaacta cacaccaggg ccaggggtca gatatccact
      121 gacctttgga tggtgctaca agctagtacc agttgagcca gataaggtag aagaggccaa
      181 taaaggagag aacaccagct tgttacaccc tgtgagcctg catgggatgg atgacccgga
      241 gagagaggac tggggagtgg cgagccctca gatcctgcat ataagcagct gctttttgcc
      301 tgtactgggt ctctctggtt agaccagatc tgagcctggg agctctctgg ctaactaggg
      361 aacccactgc ttaagcctca ataaagcttg ccttgagtgc ttcaagtagt gtgtgcccgt
      421 ctgttgtgtg actctggtaa ctagagatcc ctcagaccct tttagtcagt gtggaaaatc
      481 tctagcagtg gcgcccgaac agggacttga aagcgaaagg gaaaccagag gagctctctc
      541 gacgcaggac tcggcttgct gaagcgcgca cggcaagagg cgaggggcgg cgactggtga
      601 gtacgccaaa aattttgact agcggaggct agaaggagag agatgggtgc gagagcgtca
      661 gtattaagcg ggggagaatt agatcgcgat gggaaaaaat tcggttaagg ccagggggaa
      721 agaaaaaata taaattaaaa catatagtat gggcaagcag ggagctagaa cgattcgcag
      781 ttaatcctgg cctgttagaa acatcagaag gctgtagaca aatactggga cagctacaac
      841 catcccttca gacaggatca gaagaactta gatcattata taatacagta gcaaccctct
      901 attgtgtgca tcaaaggata gagataaaag acaccaagga agctttagac aagatagagg
      961 aagagcaaaa caaaagtaag accaccgcac agcaagcggc cggccgctga tcttcagacc
     1021 tggaggagga gatatgaggg acaattggag aagtgaatta tataaatata aagtagtaaa
     1081 aattgaacca ttaggagtag cacccaccaa ggcaaagaga agagtggtgc agagagaaaa
     1141 aagagcagtg ggaataggag ctttgttcct tgggttcttg ggagcagcag gaagcactat
     1201 gggcgcagcg tcaatgacgc tgacggtaca ggccagacaa ttattgtctg gtatagtgca
     1261 gcagcagaac aatttgctga gggctattga ggcgcaacag catctgttgc aactcacagt
     1321 ctggggcatc aagcagctcc aggcaagaat cctggctgtg gaaagatacc taaaggatca
     1381 acagctcctg gggatttggg gttgctctgg aaaactcatt tgcaccactg ctgtgccttg
     1441 gaatgctagt tggagtaata aatctctgga acagatttgg aatcacacga cctggatgga
     1501 gtgggacaga gaaattaaca attacacaag cttaatacac tccttaattg aagaatcgca
     1561 aaaccagcaa gaaaagaatg aacaagaatt attggaatta gataaatggg caagtttgtg
     1621 gaattggttt aacataacaa attggctgtg gtatataaaa ttattcataa tgatagtagg
     1681 aggcttggta ggtttaagaa tagtttttgc tgtactttct atagtgaata gagttaggca
     1741 gggatattca ccattatcgt ttcagaccca cctcccaacc ccgaggggac ccgacaggcc
     1801 cgaaggaata gaagaagaag gtggagagag agacagagac agatccattc gattagtgaa
     1861 cggatctcga cggtatcgcc tttaaaagaa aaggggggat tggggggtac agtgcagggg
     1921 aaagaatagt agacataata gcaacagaca tacaaactaa agaattacaa aaacaaatta
     1981 caaaaattca aaattttcgg gtttattaca gggacagcag agatccagtt tatcgatgag
     2041 gccctttcgt cttcactcga gtttactccc tatcagtgat agagaacgta tgaagagttt
     2101 actccctatc agtgatagag aacgtatgca gactttactc cctatcagtg atagagaacg
     2161 tataaggagt ttactcccta tcagtgatag agaacgtatg accagtttac tccctatcag
     2221 tgatagagaa cgtatctaca gtttactccc tatcagtgat agagaacgta tatccagttt
     2281 actccctatc agtgatagag aacgtatgtc gaggtaggcg tgtacggtgg gcgcctataa
     2341 aagcagagct cgtttagtga accgtcagat cgcctggagc aattccacaa cacttttgtc
     2401 ttatacttgg atcctgtaca cgtttagtga accgtcagat cgcctggaga cgccatccac
     2461 gctgttttga cctccataga agacaccgac tctactagag gatctgccac cagctagcgc
     2521 ggccgcgcca ccatgtttgt cttcctggtc ctgctgcctc tggtctcgtc tcagtgcgtg
     2581 aacctgacta ctagaaccca gctgcctcct gcctatacta actccttcac ccgcggcgtg
     2641 tactacccag acaaggtgtt ccgcagctcc gtgctgcact ccacccagga tctgttcctg
     2701 cccttcttca gcaacgtgac ctggttccac gccatccacg tgagcggcac caatggcacc
     2761 aagcggttcg acaatcccgt gctgccattc aacgatggcg tgtacttcgc ctccaccgag
     2821 aagagcaaca tcatccgcgg ctggatcttc ggcaccaccc tggactccaa gacccagagc
     2881 ctgctgatcg tgaacaatgc caccaacgtg gtcatcaagg tgtgcgagtt ccagttctgc
     2941 aatgatccat tcctgggcgt gtactaccac aagaacaata agtcctggat ggagagcgag
     3001 ttccgcgtgt acagctccgc caacaattgc accttcgagt acgtgtccca gcccttcctg
     3061 atggacctgg agggcaagca gggcaatttc aagaacctgc gcgagttcgt gttcaagaat
     3121 atcgatggct acttcaagat ctactccaag cacaccccca tcaacctggt gcgcgacctg
     3181 ccacagggct tcagcgccct ggagccactg gtggatctgc caatcggcat caacatcacc
     3241 aggttccaga ccctgctggc cctgcaccgc agctacctga ccccaggcga cagctccagc
     3301 ggatggaccg ctggagctgc tgcctactac gtgggctacc tgcagccccg caccttcctg
     3361 ctgaagtaca acgagaatgg caccatcacc gacgccgtgg attgcgccct ggatccactg
     3421 tccgagacaa agtgcaccct gaagagcttc accgtggaga agggcatcta ccagacctcc
     3481 aatttccgcg tgcagccaac cgagagcatc gtgcgcttcc ccaatatcac caacctgtgc
     3541 ccattcggcg aggtgttcaa cgctaccagg ttcgccagcg tgtacgcttg gaatcgcaag
     3601 cgcatctcca actgcgtggc cgactacagc gtgctgtaca actccgccag cttctccacc
     3661 ttcaagtgct acggcgtgtc ccccaccaag ctgaatgatc tgtgcttcac caacgtgtac
     3721 gccgatagct tcgtgatcag gggcgacgag gtgcgccaga tcgctccagg acagaccggc
     3781 aagatcgctg actacaatta caagctgccc gacgatttca ccggctgcgt gatcgcctgg
     3841 aactccaaca atctggatag caaagtgggc ggcaactaca attacctgta ccgcctgttc
     3901 cgcaagtcca atctgaagcc attcgagcgc gacatctcca ccgagatcta ccaggctgga
     3961 agcaccccat gcaatggagt ggagggcttc aactgctact tccccctgca gagctacggc
     4021 ttccagccaa ccaacggagt gggataccag ccatacaggg tggtggtgct gtccttcgag
     4081 ctgctgcacg ctccagctac cgtgtgcgga ccaaagaaga gcaccaatct ggtgaagaac
     4141 aagtgcgtga acttcaattt caacggcctg accggaaccg gcgtgctgac cgagtccaac
     4201 aagaagttcc tgccattcca gcagttcgga agggacatcg ctgataccac cgacgccgtg
     4261 cgcgacccac agaccctgga gatcctggat atcaccccat gctccttcgg cggcgtgagc
     4321 gtgatcaccc caggaaccaa taccagcaac caggtggccg tgctgtacca ggacgtgaat
     4381 tgcaccgagg tgccagtggc tatccacgct gatcagctga ccccaacctg gcgcgtgtac
     4441 agcaccggat ccaacgtgtt ccagacccgc gccggatgcc tgatcggagc tgagcacgtg
     4501 aacaattcct acgagtgcga catcccaatc ggagctggaa tctgcgccag ctaccagacc
     4561 cagaccaact ccccaaggag ggctcgcagc gtggccagcc agtccatcat cgcctacacc
     4621 atgtccctgg gcgccgagaa tagcgtggcc tacagcaaca attccatcgc catcccaacc
     4681 aacttcacca tctccgtgac caccgagatc ctgcccgtgt ccatgaccaa gaccagcgtg
     4741 gactgcacca tgtacatctg cggcgattcc accgagtgca gcaacctgct gctgcagtac
     4801 ggcagcttct gcacccagct gaatcgcgcc ctgaccggaa tcgctgtgga gcaggataag
     4861 aacacccagg aggtgttcgc ccaggtgaag cagatctaca agaccccccc aatcaaggac
     4921 ttcggcggct tcaatttcag ccagatcctg cccgatccaa gcaagccctc caagcgcagc
     4981 ttcatcgagg acctgctgtt caacaaggtg accctggccg atgccggctt catcaagcag
     5041 tacggcgatt gcctgggcga catcgctgcc cgcgacctga tctgcgccca gaagttcaat
     5101 ggcctgaccg tgctgccacc actgctgacc gatgagatga tcgctcagta cacctccgcc
     5161 ctgctggccg gaaccatcac cagcggatgg accttcggcg ctggagccgc cctgcagatc
     5221 cccttcgcca tgcagatggc ctaccgcttc aacggcatcg gcgtgaccca gaatgtgctg
     5281 tacgagaacc agaagctgat cgccaatcag ttcaactccg ccatcggcaa gatccaggac
     5341 tccctgtcca gcaccgccag cgccctgggc aagctgcagg atgtggtgaa tcagaacgcc
     5401 caggccctga ataccctggt gaagcagctg tccagcaact tcggcgccat ctccagcgtg
     5461 ctgaatgata tcctgagccg cctggacaag gtggaggctg aggtgcagat cgataggctg
     5521 atcaccggcc gcctgcagtc cctgcagacc tacgtgaccc agcagctgat cagggctgct
     5581 gagatcaggg ccagcgccaa tctggctgct accaagatgt ccgagtgcgt gctgggacag
     5641 agcaagaggg tggacttctg cggcaagggc taccacctga tgtccttccc acagagcgcc
     5701 ccacacggag tggtgttcct gcacgtgacc tacgtgccag cccaggagaa gaacttcacc
     5761 accgctccag ctatctgcca cgatggcaag gctcacttcc cacgcgaggg cgtgttcgtg
     5821 tccaacggca cccactggtt cgtgacccag cgcaatttct acgagcccca gatcatcacc
     5881 accgacaata ccttcgtgag cggcaactgc gacgtggtca tcggaatcgt gaacaatacc
     5941 gtgtacgatc ccctgcagcc agagctggac tccttcaagg aggagctgga taagtacttc
     6001 aagaatcaca ccagccccga cgtggatctg ggcgacatct ccggcatcaa tgccagcgtg
     6061 gtgaacatcc agaaggagat cgaccgcctg aacgaggtgg ccaagaatct gaacgagtcc
     6121 ctgatcgatc tgcaggagct gggcaagtac gagcagtaca tcaagtggcc atggtacatc
     6181 tggctgggct tcatcgccgg cctgatcgcc atcgtgatgg tgaccatcat gctgtgctgc
     6241 atgacctcct gctgcagctg cctgaagggc tgctgctcct gcggcagctg ctgcaagttc
     6301 gatgaggacg atagcgagcc cgtgctgaag ggcgtcaaac tgcactatac aggcaccggt
     6361 gattacaagg accacgatgg cgactataag gatcacgaca tcgactacaa ggacgatgac
     6421 gacaagtaaa cgcgtcatat ggaattctac cgggtagggg aggcgctttt cccaaggcag
     6481 tctggagcat gcgctttagc agccccgctg ggcacttggc gctacacaag tggcctctgg
     6541 cctcgcacac attccacatc caccggtagg cgccaaccgg ctccgttctt tggtggcccc
     6601 ttcgcgccac cttctactcc tcccctagtc aggaagttcc cccccgcccc gcagctcgcg
     6661 tcgtgcagga cgtgacaaat ggaagtagca cgtctcacta gtctcgtgca gatggacagc
     6721 accgctgagc aatggaagcg ggtaggcctt tggggcagcg gccaatagca gctttgctcc
     6781 ttcgctttct gggctcagag gctgggaagg ggtgggtccg ggggcgggct caggggcggg
     6841 ctcaggggcg gggcgggcgc ccgaaggtcc tccggaggcc cggcattctg cacgcttcaa
     6901 aagcgcacgt ctgccgcgct gttctcctct tcctcatctc cgggcctttc gacctgcagc
     6961 ccaagcttac cggcgcgcca tgaaaaagcc tgaactcacc gcgacgtctg tcgagaagtt
     7021 tctgatcgaa aagttcgaca gcgtctccga cctgatgcag ctctcggagg gcgaagaatc
     7081 tcgtgctttc agcttcgatg taggagggcg tggatatgtc ctgcgggtaa atagctgcgc
     7141 cgatggtttc tacaaagatc gttatgttta tcggcacttt gcatcggccg cgctcccgat
     7201 tccggaagtg cttgacattg gggaattcag cgagagcctg acctattgca tctcccgccg
     7261 tgcacagggt gtcacgttgc aagacctgcc tgaaaccgaa ctgcccgctg ttctgcagcc
     7321 ggtcgcggag gccatggatg cgatcgctgc ggccgatctt agccagacga gcgggttcgg
     7381 cccattcgga ccgcaaggaa tcggtcaata cactacatgg cgtgatttca tatgcgcgat
     7441 tgctgatccc catgtgtatc actggcaaac tgtgatggac gacaccgtca gtgcgtccgt
     7501 cgcgcaggct ctcgatgagc tgatgctttg ggccgaggac tgccccgaag tccggcacct
     7561 cgtgcacgcg gatttcggct ccaacaatgt cctgacggac aatggccgca taacagcggt
     7621 cattgactgg agcgaggcga tgttcgggga ttcccaatac gaggtcgcca acatcttctt
     7681 ctggaggccg tggttggctt gtatggagca gcagacgcgc tacttcgagc ggaggcatcc
     7741 ggagcttgca ggatcgccgc ggctccgggc gtatatgctc cgcattggtc ttgaccaact
     7801 ctatcagagc ttggttgacg gcaatttcga tgatgcagct tgggcgcagg gtcgatgcga
     7861 cgcaatcgtc cgatccggag ccgggactgt cgggcgtaca caaatcgccc gcagaagcgc
     7921 ggccgtctgg accgatggct gtgtagaagt actcgccgat agtggaaacc gacgccccag
     7981 cactcgtccg agggcaaagg aatagtctag acggggcgcg tctggaacaa tcaacctctg
     8041 gattacaaaa tttgtgaaag attgactggt attcttaact atgttgctcc ttttacgcta
     8101 tgtggatacg ctgctttaat gcctttgtat catgctattg cttcccgtat ggctttcatt
     8161 ttctcctcct tgtataaatc ctggttgctg tctctttatg aggagttgtg gcccgttgtc
     8221 aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg ttggggcatt
     8281 gccaccacct gtcagctcct ttccgggact ttcgctttcc ccctccctat tgccacggcg
     8341 gaactcatcg ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt gggcactgac
     8401 aattccgtgg tgttgtcggg gaagctgacg tcctttccat ggctgctcgc ctgtgttgcc
     8461 acctggattc tgcgcgggac gtccttctgc tacgtccctt cggccctcaa tccagcggac
     8521 cttccttccc gcggcctgct gccggctctg cggcctcttc cgcgtcttcg ccttcgccct
     8581 cagacgagtc ggatctccct ttgggccgcc tccccgcctg gaattaattc tgcagtcgag
     8641 acctagaaaa acatggagca atcacaagta gcaatacagc agctaccaat gctgattgtg
     8701 cctggctaga agcacaagag gaggaggagg tgggttttcc agtcacacct caggtacctt
     8761 taagaccaat gacttacaag gcagctgtag atcttagcca ctttttaaaa gaaaagaggg
     8821 gactggaagg gctaattcac tcccaacgaa gacaagatat ccttgatctg tggatctacc
     8881 acacacaagg ctacttccct gattagcaga actacacacc agggccaggg gtcagatatc
     8941 cactgacctt tggatggtgc tacaagctag taccagttga gccagataag gtagaagagg
     9001 ccaataaagg agagaacacc agcttgttac accctgtgag cctgcatggg atggatgacc
     9061 cggagagaga agctacaagg grctttccgc tggggacttt ccagggaggc gtggcctggg
     9121 cgggactggg gagtggcgag ccctcagatc ctgcatataa gcagctgctt tttgcctgta
     9181 ctgggtctct ctggttagac cagatctgag cctgggagct ctctggctaa ctagggaacc
     9241 cactgcttaa gcctcaataa agcttgcctt gagtgcttca agtagtgtgt gcccgtctgt
     9301 tgtgtgactc tggtaactag agatccctca gaccctttta gtcagtgtgg aaaatctcta
     9361 gcagtagtag ttcatgtcat cttattattc agtatttata acttgcaaag aaatgaatat
     9421 cagagagtga gaggccttga cattgctagc gtttaccgtc gacctctagc tagagcttgg
     9481 cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca
     9541 acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca
     9601 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
     9661 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
     9721 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     9781 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     9841 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     9901 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     9961 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
    10021 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
    10081 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
    10141 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
    10201 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
    10261 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
    10321 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
    10381 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
    10441 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
    10501 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
    10561 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
    10621 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
    10681 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
    10741 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
    10801 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
    10861 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
    10921 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
    10981 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
    11041 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
    11101 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
    11161 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
    11221 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
    11281 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
    11341 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
    11401 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
    11461 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
    11521 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
    11581 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
    11641 ctgacgtcga cggatcggga gatcaacttg tttattgcag cttataatgg ttacaaataa
    11701 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
    11761 ttgtccaaac tcatcaatgt atcttatcat gtctggatca actggataac tcaagctaac
    11821 caaaatcatc ccaaacttcc caccccatac cctattacca ctgccaatta cctgtggttt
    11881 catttactct aaacctgtga ttcctctgaa ttattttcat tttaaagaaa ttgtatttgt
    11941 taaatatgta ctacaaactt agtagtt
//
