LOCUS       sequence-473700-       14140 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  doxycycline inducible expression of E2F2 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 14140)
  AUTHORS   Clijsters L, Hoencamp C, Calis JJA, Marzio A, Handgraaf SM, Cuitino 
            MC, Rosenberg BR, Leone G, Pagano M
  TITLE     Cyclin F Controls Cell-Cycle Transcriptional Outputs by Directing 
            the Degradation of the Three Activator E2Fs.
  JOURNAL   Mol Cell. 2019 Jun 20;74(6):1264-1277.e7. doi: 
            10.1016/j.molcel.2019.04.010. Epub 2019 May 23.
  PUBMED    31130363
REFERENCE   2  (bases 1 to 14140)
  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: 
            "10.1016/j.molcel.2019.04.010"; journalName: "Mol Cell"; date: 
            "2019-06-20- 20"; volume: "74"; issue: "6"; pages: "1264-1277.e7"
COMMENT     Sequence Label: pTRIPZ-2xFLAG-2xSTREP-E2F2
FEATURES             Location/Qualifiers
     source          1..14140
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     286..306
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    405..993
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(458..478)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(876..887)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             1200..1433
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(1534..1551)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1540..1764
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1810..2238
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1897..1916)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2107..2128
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2247..2267)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2252..2581
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2432..2567
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2494..2513
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     polyA_signal    3789..3910
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3959..3975)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3959..3975)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3972..3994)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3983..3999
                     /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(4007..4013,4014..4031,4032..4037))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4052..4073
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4190..4207)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4361..4949)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4441..4460)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5120..5911,5912..5980))
                     /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     5743..5762
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5981..6085)
                     /gene="bla"
                     /label=AmpR promoter
     polyA_signal    6133..6267
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     LTR             6435..7068
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    7115..7240
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    7737..7970
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             8155..8199
                     /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    8497..8614
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     CDS             complement(8679..9053)
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     regulatory      9050..9059
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(9063..9081)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(9073..9120)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     protein_bind    9156..9174
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    9191..9209
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    9227..9245
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    9263..9281
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    9298..9316
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    9334..9352
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     promoter        9364..9402
                     /label=minimal CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     9399..9423
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             9433..9456
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             9472..9495
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             9511..9534
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             9571..9594
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             9631..9651
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     ncRNA           11030..11044
                     /label=miR-30a loop
                     /note="loop from the the 71-nt precursor of the human 
                     miR-30a microRNA (Zeng et al., 2002)"
                     /ncRNA_class="miRNA"
     ncRNA           11072..11198
                     /label=3' miR-30a
                     /note="sequence downstream of the 71-nt precursor of the 
                     human miR-30a microRNA (Zeng et al., 2002)"
                     /ncRNA_class="miRNA"
     promoter        complement(11293..11311)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        11324..12535
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     12372..12391
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     protein_bind    complement(12548..12581)
                     /label=loxP511
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATAC) (Shaw et al., 2021). loxP511 sites are 
                     compatible with each other, but incompatible with wild-type
                     loxP sites."
     CDS             12588..13295
                     /codon_start=1
                     /product="reverse tetracycline transactivator 3"
                     /label=rtTA3
                     /note="activates expression from Tet-responsive promoters 
                     in the presence of doxycycline"
                     /translation="MSRLDKSKVINGALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHFCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA
                     DALDDFDLDMLPG"
     protein_bind    complement(13296..13329)
                     /label=loxP511
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATAC) (Shaw et al., 2021). loxP511 sites are 
                     compatible with each other, but incompatible with wild-type
                     loxP sites."
     misc_feature    13352..13924
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(13519..13536)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     13746..13765
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(13927..14140,1..389)
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MATEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIE
                     RVTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGSRMAELSGSRL
                     AAQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLET
                     SAPRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
ORIGIN
        1 ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc gggggcggtg ttcgccgaga
       61 tcggctcgcg catggccgag ttgagcggtt cccggctggc cgcgcagcaa cagatggaag
      121 gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt cctggccacc gtcggcgtct
      181 cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt gctccccgga gtggaggcgg
      241 ccgagcgcgc tggggtgccc gccttcctgg agacctccgc gccccgcaac ctccccttct
      301 acgagcggct cggcttcacc gtcaccgccg acgtcgaggt gcccgaagga ccgcgcacct
      361 ggtgcatgac ccgcaagccc ggtgcctgag ttcgcgtctg gaacaatcaa cctctggatt
      421 acaaaatttg tgaaagattg actggtattc ttaactatgt tgctcctttt acgctatgtg
      481 gatacgctgc tttaatgcct ttgtatcatg ctattgcttc ccgtatggct ttcattttct
      541 cctccttgta taaatcctgg ttgctgtctc tttatgagga gttgtggccc gttgtcaggc
      601 aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc cactggttgg ggcattgcca
      661 ccacctgtca gctcctttcc gggactttcg ctttccccct ccctattgcc acggcggaac
      721 tcatcgccgc ctgccttgcc cgctgctgga caggggctcg gctgttgggc actgacaatt
      781 ccgtggtgtt gtcggggaag ctgacgtcct ttccatggct gctcgcctgt gttgccacct
      841 ggattctgcg cgggacgtcc ttctgctacg tcccttcggc cctcaatcca gcggaccttc
      901 cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga
      961 cgagtcggat ctccctttgg gccgcctccc cgcctggaat taattctgca gtcgagacct
     1021 agaaaaacat ggagcaatca caagtagcaa tacagcagct accaatgctg attgtgcctg
     1081 gctagaagca caagaggagg aggaggtggg ttttccagtc acacctcagg tacctttaag
     1141 accaatgact tacaaggcag ctgtagatct tagccacttt ttaaaagaaa agaggggact
     1201 ggaagggcta attcactccc aacgaagaca agatctgctt tttgcttgta ctgggtctct
     1261 ctggttagac cagatctgag cctgggagct ctctggctaa ctagggaacc cactgcttaa
     1321 gcctcaataa agcttgcctt gagtgcttca agtagtgtgt gcccgtctgt tgtgtgactc
     1381 tggtaactag agatccctca gaccctttta gtcagtgtgg aaaatctcta gcagtagtag
     1441 ttcatgtcat cttattattc agtatttata acttgcaaag aaatgaatat cagagagtga
     1501 gaggccttga cattgtttaa acccgctgat cagcctcgac tgtgccttct agttgccagc
     1561 catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg
     1621 tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc
     1681 tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg
     1741 ctggggatgc ggtgggctct atggcttctg aggcggaaag aaccagctgg ggctctaggg
     1801 ggtatcccca cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca
     1861 gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct
     1921 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt
     1981 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac
     2041 gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct
     2101 ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt
     2161 ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac
     2221 aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc
     2281 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg
     2341 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     2401 agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc
     2461 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     2521 tgcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa
     2581 aaagctcccg ggagcttgta tatccatttt cggatctgat cagcacgtga tgaaaaagcc
     2641 tgaactcacc gcgacgtctg tcgagaagtt tctgatcgaa aagttcgaca gcgtctccga
     2701 cctgatgcag ctctcggagg gcgaagaatc tcgtgctttc agcttcgatg taggagggcg
     2761 tggatatgtc ctgcgggtaa atagctgcgc cgatggtttc tacaaagatc gttatgttta
     2821 tcggcacttt gcatcggccg cgctcccgat tccggaagtg cttgacattg gggaattaat
     2881 tcagcgagag cctgacctat tgcatctccc gccgtgcaca gggtgtcacg ttgcaagacc
     2941 tgcctgaaac cgaactgccc gctgttctgc agccggtcgc ggaggccatg gatgcgatcg
     3001 ctgcggccga tcttagccag acgagcgggt tcggcccatt cggaccgcaa ggaatcggtc
     3061 aatacactac atggcgtgat ttcatatgcg cgattgctga tccccatgtg tatcactggc
     3121 aaactgtgat ggacgacacc gtcagtgcgt ccgtcgcgca ggctctcgat gagctgatgc
     3181 tttgggccga ggactgcccc gaagtccggc acctcgtgca cgcggatttc ggctccaaca
     3241 atgtcctgac ggacaatggc cgcataacag cggtcattga ctggagcgag gcgatgttcg
     3301 gggattccca atacgaggtc gccaacatct tcttctggag gccgtggttg gcttgtatgg
     3361 agcagcagac gcgctacttc gagcggaggc atccggagct tgcaggatcg ccgcggctcc
     3421 gggcgtatat gctccgcatt ggtcttgacc aactctatca gagcttggtt gacggcaatt
     3481 tcgatgatgc agcttgggcg cagggtcgat gcgacgcaat cgtccgatcc ggagccggga
     3541 ctgtcgggcg tacacaaatc gcccgcagaa gcgcggccgt ctggaccgat ggctgtgtag
     3601 aagtactcgc cgatagtgga aaccgacgcc ccagcactcg tccgagggca aaggaatagc
     3661 acgtgctacg agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg
     3721 ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg
     3781 cccaccccaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     3841 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     3901 atgtatctta tcatgtctgt ataccgtcga cctctagcta gagcttggcg taatcatggt
     3961 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg
     4021 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt
     4081 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg
     4141 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg
     4201 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     4261 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     4321 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     4381 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     4441 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     4501 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     4561 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     4621 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     4681 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     4741 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     4801 gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     4861 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     4921 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     4981 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     5041 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     5101 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
     5161 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     5221 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
     5281 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     5341 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     5401 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
     5461 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     5521 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     5581 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     5641 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     5701 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     5761 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     5821 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     5881 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     5941 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
     6001 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     6061 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtcgacg
     6121 gatcgggaga tcaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc
     6181 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc
     6241 atcaatgtat cttatcatgt ctggatcaac tggataactc aagctaacca aaatcatccc
     6301 aaacttccca ccccataccc tattaccact gccaattacc tgtggtttca tttactctaa
     6361 acctgtgatt cctctgaatt attttcattt taaagaaatt gtatttgtta aatatgtact
     6421 acaaacttag tagttggaag ggctaattca ctcccaaaga agacaagata tccttgatct
     6481 gtggatctac cacacacaag gctacttccc tgattagcag aactacacac cagggccagg
     6541 ggtcagatat ccactgacct ttggatggtg ctacaagcta gtaccagttg agccagataa
     6601 ggtagaagag gccaataaag gagagaacac cagcttgtta caccctgtga gcctgcatgg
     6661 gatggatgac ccggagagag aagtgttaga gtggaggttt gacagccgcc tagcatttca
     6721 tcacgtggcc cgagagctgc atccggagta cttcaagaac tgctgatatc gagcttgcta
     6781 caagggactt tccgctgggg actttccagg gaggcgtggc ctgggcggga ctggggagtg
     6841 gcgagccctc agatcctgca tataagcagc tgctttttgc ctgtactggg tctctctggt
     6901 tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg cttaagcctc
     6961 aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt gactctggta
     7021 actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagt ggcgcccgaa
     7081 cagggacttg aaagcgaaag ggaaaccaga ggagctctct cgacgcagga ctcggcttgc
     7141 tgaagcgcgc acggcaagag gcgaggggcg gcgactggtg agtacgccaa aaattttgac
     7201 tagcggaggc tagaaggaga gagatgggtg cgagagcgtc agtattaagc gggggagaat
     7261 tagatcgcga tgggaaaaaa ttcggttaag gccaggggga aagaaaaaat ataaattaaa
     7321 acatatagta tgggcaagca gggagctaga acgattcgca gttaatcctg gcctgttaga
     7381 aacatcagaa ggctgtagac aaatactggg acagctacaa ccatcccttc agacaggatc
     7441 agaagaactt agatcattat ataatacagt agcaaccctc tattgtgtgc atcaaaggat
     7501 agagataaaa gacaccaagg aagctttaga caagatagag gaagagcaaa acaaaagtaa
     7561 gaccaccgca cagcaagcgg ccggccgctg atcttcagac ctggaggagg agatatgagg
     7621 gacaattgga gaagtgaatt atataaatat aaagtagtaa aaattgaacc attaggagta
     7681 gcacccacca aggcaaagag aagagtggtg cagagagaaa aaagagcagt gggaatagga
     7741 gctttgttcc ttgggttctt gggagcagca ggaagcacta tgggcgcagc gtcaatgacg
     7801 ctgacggtac aggccagaca attattgtct ggtatagtgc agcagcagaa caatttgctg
     7861 agggctattg aggcgcaaca gcatctgttg caactcacag tctggggcat caagcagctc
     7921 caggcaagaa tcctggctgt ggaaagatac ctaaaggatc aacagctcct ggggatttgg
     7981 ggttgctctg gaaaactcat ttgcaccact gctgtgcctt ggaatgctag ttggagtaat
     8041 aaatctctgg aacagatttg gaatcacacg acctggatgg agtgggacag agaaattaac
     8101 aattacacaa gcttaataca ctccttaatt gaagaatcgc aaaaccagca agaaaagaat
     8161 gaacaagaat tattggaatt agataaatgg gcaagtttgt ggaattggtt taacataaca
     8221 aattggctgt ggtatataaa attattcata atgatagtag gaggcttggt aggtttaaga
     8281 atagtttttg ctgtactttc tatagtgaat agagttaggc agggatattc accattatcg
     8341 tttcagaccc acctcccaac cccgagggga cccgacaggc ccgaaggaat agaagaagaa
     8401 ggtggagaga gagacagaga cagatccatt cgattagtga acggatcggc actgcgtgcg
     8461 ccaattctgc agacaaatgg cagtattcat ccacaatttt aaaagaaaag gggggattgg
     8521 ggggtacagt gcaggggaaa gaatagtaga cataatagca acagacatac aaactaaaga
     8581 attacaaaaa caaattacaa aaattcaaaa ttttcgggtt tattacaggg acagcagaga
     8641 tccagtttgg ttagtaccgg gcccgctcta gtccggaatc agtcctgctc ctcggccacg
     8701 aagtgcacgc agttgccggc cgggtcgcgc agggcgaact cccgccccca cggctgctcg
     8761 ccgatctcgg tcatggccgg cccggaggcg tcccggaagt tcgtggacac gacctccgac
     8821 cactcggcgt acagctcgtc caggccgcgc acccacaccc aggccagggt gttgtccggc
     8881 accacctggt cctggaccgc gctgatgaac agggtcacgt cgtcccggac cacaccggcg
     8941 aagtcgtcct ccacgaagtc ccgggagaac ccgagccggt cggtccagaa ctcgaccgct
     9001 ccggcgacgt cgcgcgcggt gagcaccgga acggcactgg tcaacttggc catggtggcc
     9061 ctcctatagt gagtcgtatt atactatgcc gatatactat gccgatgatt aattgtcaac
     9121 acgtgctgca ggtccgaggt tctagacgag tttactccct atcagtgata gagaacgatg
     9181 tcgagtttac tccctatcag tgatagagaa cgtatgtcga gtttactccc tatcagtgat
     9241 agagaacgta tgtcgagttt actccctatc agtgatagag aacgtatgtc gagtttatcc
     9301 ctatcagtga tagagaacgt atgtcgagtt tactccctat cagtgataga gaacgtatgt
     9361 cgaggtaggc gtgtacggtg ggaggcctat ataagcagag ctcgtttagt gaaccgtcag
     9421 atcgcaacca tggactacaa ggacgacgat gacaaggtac caggatctat ggattataaa
     9481 gatgatgatg ataaagggtc ggccgccagc tggagccacc ctcagttcga gaagggagga
     9541 ggaagcggcg gaggcagcgg aggaggaagc tggagccacc cgcagttcga gaaaggagct
     9601 ggatccagtg tggtggaatt gggagcatca ccaaaaaaga agagaaaggt acccgtggct
     9661 caattcctgc aagggccccg ggccttggct tcggccgctg ggcagacccc gaaggtggtg
     9721 cccgcgatga gccccacaga gctgtggcca tccggcctca gcagccccca gctctgccca
     9781 gctactgcta cctactacac accgctgtac ccgcagacgg cgcctcccgc agcggcgcca
     9841 ggcacctgcc tcgacgccac tccccacgga cccgagggcc aagttgtgcg atgcctgccg
     9901 gcaggccggc tgccggccaa aaggaagctg gatctggagg ggattgggag gcccgtcgtc
     9961 cctgagttcc caacccccaa ggggaagtgc atcagagtgg atggcctccc cagccccaaa
    10021 acccccaaat cccccgggga gaagactcgg tatgacactt cgctggggct gctcaccaag
    10081 aagttcattt acctcctgag cgagtcagag gatggggtcc tggacctgaa ctgggccgct
    10141 gaggtgctgg acgtgcagaa gcggcgcatc tatgacatca ccaacgtgct ggaaggcatc
    10201 cagctcatcc gcaagaaggc caagaacaac atccagtggg taggcagggg aatgtttgaa
    10261 gaccccacca gacctgggaa gcagcaacag ctggggcagg agctgaagga gctgatgaac
    10321 acggagcagg ccttggacca gctcatccag agctgctctc tgagcttcaa gcacctgact
    10381 gaggacaagg ccaacaagag gctggcctat gtgacttacc aggatatccg tgctgttggc
    10441 aactttaagg agcagacagt gattgccgtc aaggcccctc cgcagacgag actggaagtg
    10501 cccgacagga ctgaggacaa cctgcagata tatctcaaga gcacccaagg gcccatcgaa
    10561 gtctacctgt gcccagagga ggtgcaggag ccggacagtc cttccgagga gcctctcccc
    10621 tctacctcca ccctctgccc cagccctgac tctgcccagc ccagcagcag caccgaccct
    10681 agcatcatgg agcccacagc atcctcagtg ccagcaccag cgccaacccc ccagcaggcc
    10741 ccaccgcctc catccctggt ccccttggag gctactgaca gcctgctgga gctgccgcac
    10801 ccactcctgc agcagactga ggaccagttc ctgtccccga ccctggcgtg cagctcccct
    10861 ctgatcagct tctccccatc cttggaccag gacgactacc tgtggggctt ggaggcgggt
    10921 gagggcatca gcgatctctt cgactcctac gaccttgggg acctgttgat taattgagaa
    10981 ggtatattgc tgttgacagt gagcgatctc gcttgggcga gagtaagtag tgaagccaca
    11041 gatgtactta ctctcgccca agcgagagtg cctactgcct cggaattcaa ggggctactt
    11101 taggagcaat tatcttgttt actaaaactg aataccttgc tatctctttg atacattttt
    11161 acaaagctga attaaaatgg tataaattaa atcacttttt tcaattggaa gactaatgcg
    11221 gccggccatt actccgtctc gtgtcttgtt gcatatgtct gctggtttgt ttgatgttgt
    11281 ttgcgggcgg gccctatagt gagtcgtatt acctaggacg cgtggcctcc gcgccgggtt
    11341 ttggcgcctc ccgcgggcgc ccccctcctc acggcgagcg ctgccacgtc agacgaaggg
    11401 cgcagcgagc gtcctgatcc ttccgcccgg acgctcagga cagcggcccg ctgctcataa
    11461 gactcggcct tagaacccca gtatcagcag aaggacattt taggacggga cttgggtgac
    11521 tctagggcac tggttttctt tccagagagc ggaacaggcg aggaaaagta gtcccttctc
    11581 ggcgattctg cggagggatc tccgtggggc ggtgaacgcc gatgattata taaggacgcg
    11641 ccgggtgtgg cacagctagt tccgtcgcag ccgggatttg ggtcgcggtt cttgtttgtg
    11701 gatcgctgtg atcgtcactt ggtgagtagc gggctgctgg gctggccggg gctttcgtgg
    11761 ccgccgggcc gctcggtggg acggaagcgt gtggagagac cgccaagggc tgtagtctgg
    11821 gtccgcgagc aaggttgccc tgaactgggg gttgggggga gcgcagcaaa atggcggctg
    11881 ttcccgagtc ttgaatggaa gacgcttgtg aggcgggctg tgaggtcgtt gaaacaaggt
    11941 ggggggcatg gtgggcggca agaacccaag gtcttgaggc cttcgctaat gcgggaaagc
    12001 tcttattcgg gtgagatggg ctggggcacc atctggggac cctgacgtga agtttgtcac
    12061 tgactggaga actcggtttg tcgtctgttg cgggggcggc agttatggcg gtgccgttgg
    12121 gcagtgcacc cgtacctttg ggagcgcgcg ccctcgtcgt gtcgtgacgt cacccgttct
    12181 gttggcttat aatgcagggt ggggccacct gccggtaggt gtgcggtagg cttttctccg
    12241 tcgcaggacg cagggttcgg gcctagggta ggctctcctg aatcgacagg cgccggacct
    12301 ctggtgaggg gagggataag tgaggcgtca gtttctttgg tcggttttat gtacctatct
    12361 tcttaagtag ctgaagctcc ggttttgaac tatgcgctcg gggttggcga gtgtgttttg
    12421 tgaagttttt taggcacctt ttgaaatgta atcatttggg tcaatatgta attttcagtg
    12481 ttagactagt aaattgtccg ctaaattctg gccgtttttg gcttttttgt tagacgctag
    12541 cggatccata acttcgtata gtatacatta tacgaagtta tgccaccatg tctaggctgg
    12601 acaagagcaa agtcataaac ggagctctgg aattactcaa tggtgtcggt atcgaaggcc
    12661 tgacgacaag gaaactcgct caaaagctgg gagttgagca gcctaccctg tactggcacg
    12721 tgaagaacaa gcgggccctg ctcgatgccc tgccaatcga gatgctggac aggcatcata
    12781 cccacttctg ccccctggaa ggcgagtcat ggcaagactt tctgcggaac aacgccaagt
    12841 cataccgctg tgctctcctc tcacatcgcg acggggctaa agtgcatctc ggcacccgcc
    12901 caacagagaa acagtacgaa accctggaaa atcagctcgc gttcctgtgt cagcaaggct
    12961 tctccctgga gaacgcactg tacgctctgt ccgccgtggg ccactttaca ctgggctgcg
    13021 tattggagga acaggagcat caagtagcaa aagaggaaag agagacacct accaccgatt
    13081 ctatgccccc acttctgaga caagcaattg agctgttcga ccggcaggga gccgaacctg
    13141 ccttcctttt cggcctggaa ctaatcatat gtggcctgga gaaacagcta aagtgcgaaa
    13201 gcggcgggcc gaccgacgcc cttgacgatt ttgacttaga catgctccca gccgatgccc
    13261 ttgacgattt tgaccttgac atgctccccg ggtaaataac ttcgtatagt atacattata
    13321 cgaagttatg gatccgcggc cgcaaattcc gcccctctcc ctcccccccc cctaacgtta
    13381 ctggccgaag ccgcttggaa taaggccggt gtgcgtttgt ctatatgtta ttttccacca
    13441 tattgccgtc ttttggcaat gtgagggccc ggaaacctgg ccctgtcttc ttgacgagca
    13501 ttcctagggg tctttcccct ctcgccaaag gaatgcaagg tctgttgaat gtcgtgaagg
    13561 aagcagttcc tctggaagct tcttgaagac aaacaacgtc tgtagcgacc ctttgcaggc
    13621 agcggaaccc cccacctggc gacaggtgcc tctgcggcca aaagccacgt gtataagata
    13681 cacctgcaaa ggcggcacaa ccccagtgcc acgttgtgag ttggatagtt gtggaaagag
    13741 tcaaatggct ctcctcaagc gtattcaaca aggggctgaa ggatgcccag aaggtacccc
    13801 attgtatggg atctgatctg gggcctcggt gcacatgctt tacatgtgtt tagtcgaggt
    13861 taaaaaaacg tctaggcccc ccgaaccacg gggacgtggt tttcctttga aaaacacgat
    13921 aataccatgg ccaccgagta caagcccacg gtgcgcctcg ccacccgcga cgacgtcccc
    13981 cgggccgtac gcaccctcgc cgccgcgttc gccgactacc ccgccacgcg ccacaccgtc
    14041 gacccggacc gccacatcga gcgggtcacc gagctgcaag aactcttcct cacgcgcgtc
    14101 gggctcgaca tcggcaaggt gtgggtcgcg gacgacggcg
//
