LOCUS       sequence-369490-       10656 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Expresses flag-tagged EGFP in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10656)
  AUTHORS   Stransky LA, Vigeant SM, Huang B, West D, Denize T, Walton E, 
            Signoretti S, Kaelin WG Jr
  TITLE     Sensitivity of VHL mutant kidney cancers to HIF2 inhibitors does not
            require an intact p53 pathway.
  JOURNAL   Proc Natl Acad Sci U S A. 2022 Apr 5;119(14):e2120403119. doi: 
            10.1073/pnas.2120403119. Epub 2022 Mar 31.
  PUBMED    35357972
REFERENCE   2  (bases 1 to 10656)
  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: "Proc Natl 
            Acad Sci U S A."; date: "2022-04-5"; volume: "119(14):e2120403119. 
            doi"; pages: " 10.1073/pnas.2120403119. Epub 2022 Mar 31"
COMMENT     Sequence Label: pLenti_EF1a_Flag_EGFP
FEATURES             Location/Qualifiers
     source          1..10656
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    14..38
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             44..67
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             join(83..85,86..799)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(83..85,86..796)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(125..146)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(386..405)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     733..754
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(801..825)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             845..868
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        894..1393
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(929..952)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     1326..1345
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             1411..2436
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             1411..2436
                     /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"
     protein_bind    complement(2467..2500)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    2556..3144
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2609..2629)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3027..3038)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3147..3163)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3148..3164)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             3353..3586
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    3658..3779
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3695..3714)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3749..3768
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      3819..3954
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        complement(3975..3993)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     rep_origin      complement(4016..4374)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4056..4077)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4268..4287
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4401..4505
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4507..4864
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4507..4698
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4530..4550)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4715..4850
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4899..5693
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             4899..5693
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4953..4972)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5563..5582
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5881..5900)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5925..5972
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6301..6889
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6790..6809
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        7064..7367
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7369..7567
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7522..7542
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             7585..7765
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    7812..7937
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    8434..8667
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             8852..8896
                     /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    9194..9311
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        9383..10561
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          9614..10552
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     10509..10529
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     promoter        10578..10596
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 gatctggcca atcacaagtt tgtacaaaaa agcaggctaa atggactaca aggacgacga
       61 tgacaagggt ggcggaggga gcgtgagcaa gggcgaggag ctgttcaccg gggtggtgcc
      121 catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg
      181 cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct
      241 gcccgtgccc tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg
      301 ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt
      361 ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa
      421 gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga
      481 cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg tctatatcat
      541 ggccgacaag cagaagaacg gcatcaaggt gaacttcaag atccgccaca acatcgagga
      601 cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt
      661 gctgctgccc gacaaccact acctgagcac ccagtccgcc ctgagcaaag accccaacga
      721 gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat
      781 ggacgagctg tacaagtgaa acccagcttt cttgtacaaa gtggtgggag gaggtggatc
      841 tggcgactac aaggacgacg atgacaagtg actcgaggtt aacgaattct accgggtagg
      901 ggaggcgctt ttcccaaggc agtctggagc atgcgcttta gcagccccgc tgggcacttg
      961 gcgctacaca agtggcctct ggcctcgcac acattccaca tccaccggta ggcgccaacc
     1021 ggctccgttc tttggtggcc ccttcgcgcc accttctact cctcccctag tcaggaagtt
     1081 cccccccgcc ccgcagctcg cgtcgtgcag gacgtgacaa atggaagtag cacgtctcac
     1141 tagtctcgtg cagatggaca gcaccgctga gcaatggaag cgggtaggcc tttggggcag
     1201 cggccaatag cagctttgct ccttcgcttt ctgggctcag aggctgggaa ggggtgggtc
     1261 cgggggcggg ctcaggggcg ggctcagggg cggggcgggc gcccgaaggt cctccggagg
     1321 cccggcattc tgcacgcttc aaaagcgcac gtctgccgcg ctgttctcct cttcctcatc
     1381 tccgggcctt tcgacctgca tcccgccacc atgaaaaagc ctgaactcac cgcgacgtct
     1441 gtcgagaagt ttctgatcga aaagttcgac agcgtctccg acctgatgca gctctcggag
     1501 ggcgaagaat ctcgtgcttt cagcttcgat gtaggagggc gtggatatgt cctgcgggta
     1561 aatagctgcg ccgatggttt ctacaaagat cgttatgttt atcggcactt tgcatcggcc
     1621 gcgctcccga ttccggaagt gcttgacatt ggggaattca gcgagagcct gacctattgc
     1681 atctcccgcc gtgcacaggg tgtcacgttg caagacctgc ctgaaaccga actgcccgct
     1741 gttctgcagc cggtcgcgga ggccatggat gcgatcgctg cggccgatct tagccagacg
     1801 agcgggttcg gcccattcgg accgcaagga atcggtcaat acactacatg gcgtgatttc
     1861 atatgcgcga ttgctgatcc ccatgtgtat cactggcaaa ctgtgatgga cgacaccgtc
     1921 agtgcgtccg tcgcgcaggc tctcgatgag ctgatgcttt gggccgagga ctgccccgaa
     1981 gtccggcacc tcgtgcacgc ggatttcggc tccaacaatg tcctgacgga caatggccgc
     2041 ataacagcgg tcattgactg gagcgaggcg atgttcgggg attcccaata cgaggtcgcc
     2101 aacatcttct tctggaggcc gtggttggct tgtatggagc agcagacgcg ctacttcgag
     2161 cggaggcatc cggagcttgc aggatcgccg cggctccggg cgtatatgct ccgcattggt
     2221 cttgaccaac tctatcagag cttggttgac ggcaatttcg atgatgcagc ttgggcgcag
     2281 ggtcgatgcg acgcaatcgt ccgatccgga gccgggactg tcgggcgtac acaaatcgcc
     2341 cgcagaagcg cggccgtctg gaccgatggc tgtgtagaag tactcgccga tagtggaaac
     2401 cgacgcccca gcactcgtcc gagggcaaag gaatagtgta caagtaggaa ttcgtcgagg
     2461 gacctaataa cttcgtatag catacattat acgaagttat acatgtttaa gggttccggt
     2521 tccactaggt acaattcgat atcaagctta tcgataatca acctctggat tacaaaattt
     2581 gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt ggatacgctg
     2641 ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc tcctccttgt
     2701 ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg caacgtggcg
     2761 tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc accacctgtc
     2821 agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa ctcatcgccg
     2881 cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat tccgtggtgt
     2941 tgtcggggaa atcatcgtcc tttccttggc tgctcgcctg tgttgccacc tggattctgc
     3001 gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg
     3061 gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag acgagtcgga
     3121 tctccctttg ggccgcctcc ccgcatcgat accgtcgacc tcgatcgaga cctagaaaaa
     3181 catggagcaa tcacaagtag caatacagca gctaccaatg ctgattgtgc ctggctagaa
     3241 gcacaagagg aggaggaggt gggttttcca gtcacacctc aggtaccttt aagaccaatg
     3301 acttacaagg cagctgtaga tcttagccac tttttaaaag aaaagggggg actggaaggg
     3361 ctaattcact cccaacgaag acaagatctg ctttttgctt gtactgggtc tctctggtta
     3421 gaccagatct gagcctggga gctctctggc taactaggga acccactgct taagcctcaa
     3481 taaagcttgc cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac
     3541 tagagatccc tcagaccctt ttagtcagtg tggaaaatct ctagcagtag tagttcatgt
     3601 catcttatta ttcagtattt ataacttgca aagaaatgaa tatcagagag tgagaggaac
     3661 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
     3721 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat
     3781 catgtctggc tctagctatc ccgcccctaa ctccgcccat cccgccccta actccgccca
     3841 gttccgccca ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg
     3901 ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggga
     3961 cgtacccaat tcgccctata gtgagtcgta ttagcatgtg agcaaaaggc cagcataaat
     4021 caaaagaata gaccgagata gggttgagtg ttgttccagt ttggaacaag agtccactat
     4081 taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac
     4141 tacgtgaacc atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc
     4201 ggaaccctaa agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga
     4261 gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca
     4321 cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtcaggtg
     4381 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
     4441 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
     4501 agagtcctga ggcggaaaga accagctgtg gaatgtgtgt cagttagggt gtggaaagtc
     4561 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag
     4621 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
     4681 gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc
     4741 cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc
     4801 ctcggcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg
     4861 caaagatcga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc
     4921 acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga
     4981 caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt
     5041 ttgtcaagac cgacctgtcc ggtgccctga atgaactgca agacgaggca gcgcggctat
     5101 cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg
     5161 gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg
     5221 ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc
     5281 cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga
     5341 tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag
     5401 ccgaactgtt cgccaggctc aaggcgagca tgcccgacgg cgaggatctc gtcgtgaccc
     5461 atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg
     5521 actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata
     5581 ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg
     5641 ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac
     5701 tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc
     5761 caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat
     5821 gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccctaggg ggaggctaac
     5881 tgaaacacgg aaggagacaa taccggaagg aacccgcgct atgacggcaa taaaaagaca
     5941 gaataaaacg cacggtgttg ggtcgtttgt tcataaacgc ggggttcggt cccagggctg
     6001 gcactctgtc gataccccac cgagacccca ttggggccaa tacgcccgcg tttcttcctt
     6061 ttccccaccc caccccccaa gttcgggtga aggcccaggg ctcgcagcca acgtcggggc
     6121 ggcaggccct gccatagcct caggttactc atatatactt tagattgatt taaaacttca
     6181 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     6241 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     6301 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     6361 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     6421 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
     6481 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     6541 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     6601 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     6661 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     6721 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     6781 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     6841 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     6901 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     6961 gttatcccct gattctgtgg ataaccgtat taccgccatg cattagttat taatagtaat
     7021 caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg
     7081 taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt
     7141 atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac
     7201 ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtacg ccccctattg
     7261 acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc ttatgggact
     7321 ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt
     7381 ggcagtacat caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc
     7441 ccattgacgt caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc
     7501 gtaacaactc cgccccattg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata
     7561 taagcagcgc gttttgcctg tactgggtct ctctggttag accagatctg agcctgggag
     7621 ctctctggct aactagggaa cccactgctt aagcctcaat aaagcttgcc ttgagtgctt
     7681 caagtagtgt gtgcccgtct gttgtgtgac tctggtaact agagatccct cagacccttt
     7741 tagtcagtgt ggaaaatctc tagcagtggc gcccgaacag ggacttgaaa gcgaaaggga
     7801 aaccagagga gctctctcga cgcaggactc ggcttgctga agcgcgcacg gcaagaggcg
     7861 aggggcggcg actggtgagt acgccaaaaa ttttgactag cggaggctag aaggagagag
     7921 atgggtgcga gagcgtcagt attaagcggg ggagaattag atcgcgatgg gaaaaaattc
     7981 ggttaaggcc agggggaaag aaaaaatata aattaaaaca tatagtatgg gcaagcaggg
     8041 agctagaacg attcgcagtt aatcctggcc tgttagaaac atcagaaggc tgtagacaaa
     8101 tactgggaca gctacaacca tcccttcaga caggatcaga agaacttaga tcattatata
     8161 atacagtagc aaccctctat tgtgtgcatc aaaggataga gataaaagac accaaggaag
     8221 ctttagacaa gatagaggaa gagcaaaaca aaagtaagac caccgcacag caagcggccg
     8281 gccgctgatc ttcagacctg gaggaggaga tatgagggac aattggagaa gtgaattata
     8341 taaatataaa gtagtaaaaa ttgaaccatt aggagtagca cccaccaagg caaagagaag
     8401 agtggtgcag agagaaaaaa gagcagtggg aataggagct ttgttccttg ggttcttggg
     8461 agcagcagga agcactatgg gcgcagcgtc aatgacgctg acggtacagg ccagacaatt
     8521 attgtctggt atagtgcagc agcagaacaa tttgctgagg gctattgagg cgcaacagca
     8581 tctgttgcaa ctcacagtct ggggcatcaa gcagctccag gcaagaatcc tggctgtgga
     8641 aagataccta aaggatcaac agctcctggg gatttggggt tgctctggaa aactcatttg
     8701 caccactgct gtgccttgga atgctagttg gagtaataaa tctctggaac agatttggaa
     8761 tcacacgacc tggatggagt gggacagaga aattaacaat tacacaagct taatacactc
     8821 cttaattgaa gaatcgcaaa accagcaaga aaagaatgaa caagaattat tggaattaga
     8881 taaatgggca agtttgtgga attggtttaa cataacaaat tggctgtggt atataaaatt
     8941 attcataatg atagtaggag gcttggtagg tttaagaata gtttttgctg tactttctat
     9001 agtgaataga gttaggcagg gatattcacc attatcgttt cagacccacc tcccaacccc
     9061 gaggggaccc gacaggcccg aaggaataga agaagaaggt ggagagagag acagagacag
     9121 atccattcga ttagtgaacg gatcggcact gcgtgcgcca attctgcaga caaatggcag
     9181 tattcatcca caattttaaa agaaaagggg ggattggggg gtacagtgca ggggaaagaa
     9241 tagtagacat aatagcaaca gacatacaaa ctaaagaatt acaaaaacaa attacaaaaa
     9301 ttcaaaattt tcgggtttat tacagggaca gcagagatcc agtttggtta gtaccgggcc
     9361 cgctctagcc ctagatttaa atggctccgg tgcccgtcag tgggcagagc gcacatcgcc
     9421 cacagtcccc gagaagttgg ggggaggggt cggcaattga accggtgcct agagaaggtg
     9481 gcgcggggta aactgggaaa gtgatgtcgt gtactggctc cgcctttttc ccgagggtgg
     9541 gggagaaccg tatataagtg cagtagtcgc cgtgaacgtt ctttttcgca acgggtttgc
     9601 cgccagaaca caggtaagtg ccgtgtgtgg ttcccgcggg cctggcctct ttacgggtta
     9661 tggcccttgc gtgccttgaa ttacttccac ctggctgcag tacgtgattc ttgatcccga
     9721 gcttcgggtt ggaagtgggt gggagagttc gaggccttgc gcttaaggag ccccttcgcc
     9781 tcgtgcttga gttgaggcct ggcctgggcg ctggggccgc cgcgtgcgaa tctggtggca
     9841 ccttcgcgcc tgtctcgctg ctttcgataa gtctctagcc atttaaaatt tttgatgacc
     9901 tgctgcgacg ctttttttct ggcaagatag tcttgtaaat gcgggccaag atctgcacac
     9961 tggtatttcg gtttttgggg ccgcgggcgg cgacggggcc cgtgcgtccc agcgcacatg
    10021 ttcggcgagg cggggcctgc gagcgcggcc accgagaatc ggacgggggt agtctcaagc
    10081 tggccggcct gctctggtgc ctggcctcgc gccgccgtgt atcgccccgc cctgggcggc
    10141 aaggctggcc cggtcggcac cagttgcgtg agcggaaaga tggccgcttc ccggccctgc
    10201 tgcagggagc tcaaaatgga ggacgcggcg ctcgggagag cgggcgggtg agtcacccac
    10261 acaaaggaaa agggcctttc cgtcctcagc cgtcgcttca tgtgactcca cggagtaccg
    10321 ggcgccgtcc aggcacctcg attagttctc gagcttttgg agtacgtcgt ctttaggttg
    10381 gggggagggg ttttatgcga tggagtttcc ccacactgag tgggtggaga ctgaagttag
    10441 gccagcttgg cacttgatgt aattctcctt ggaatttgcc ctttttgagt ttggatcttg
    10501 gttcattctc aagcctcaga cagtggttca aagttttttt cttccatttc aggtgtcgtg
    10561 aggaattagc ttggtactaa tacgactcac tatagggaga cccaagctgg ctagccgcgg
    10621 aatctagact gccggatcga attgtcgacg aattcg
//
