LOCUS       Exported                8623 bp ds-DNA     circular SYN 20-FEB-2018
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8623)
  AUTHORS   UMC-Utrecht
  TITLE     Direct Submission
  JOURNAL   Exported Mar 8, 2018 from SnapGene Viewer 4.1.7
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8623
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV forward primer
     protein_bind    820..859
                     /label=TetR binding sites
     protein_bind    820..838
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    841..859
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     misc_feature    967..996
                     /label=MCS
     V_region        1027..1062
                     /label=VSV
     CDS             1063..3819
                     /codon_start=1
                     /label=INCENP RNAi
                     /translation="GTTAPGPIHLLELCDQKLMEFLCNMDNKDLVWLEEIQEEAERMFT
                     REFSKEPELMPKTPSQKNRRKKRRISYVQDENRDPIRRRLSRRKSRSSQQSTRRLRSKD
                     SVEKLATVVGENGSVLRRVTRAAAAAAAATMALAAPSSPTPESPTMLTKKPEDNHTQCQ
                     LVPVVEIGISERQNAEQHVTQLMSTEPLPRTLSPTPASATAPTSQGIPTSDEESTPKKS
                     KARILESITVSSLMATPQDPKGQGVGTGRSASKLRIAQVSPGPRDSPAFPDSPWRERVL
                     APILPDNFSTPTGSRTDSQSVRHSPIAPSSPSPRVLAQKYSLVAKQESVVRRASRRLAK
                     KTAEEPAASGRIICHSYLERLLNVEVPQKVGSEQKEPPEEAEPVAAAEPEVPENNGNNS
                     WPHNDTEIANSTPNPKPAASSPETPSAGQQEAKTDQADGPREPPQSARRKRSYKQAVSE
                     LDEEQHLEDEELQPPRSKTPSSPCPASKVVRPLRTFLHTVQRNQMLTTPTSAPRSVMKS
                     FIKRNTPLRMDPKCSFVEKERQRLENLRRKEEAEQLRRQKVEEDKRRRLEEVKLKREER
                     LRKVLQARERVEQMKEEKKKQIEQKFAQIDEKTEKAKEERLAEEKAKKKAAAKKMEEVE
                     ARRKQEEDARRLRWLQQEEEERRHQELLQKKKEEEQERLRKAAEAKRLAEQREQERREQ
                     ERREQERREQERREQERREQERQLAEQERRREQERLQAERELQEREKALRLQKEQLQRE
                     LEEKKKKEEQQRLAERQLQEEQEKKAKEAVGASKALNVTVDVQSPACTSYQMTPQGHRA
                     PPKINPDNYGMDLNSDDSTDDEAHPRKPIPTWARGTPLSQAIIHQYYHPPNLLELFGTI
                     LPLDLEDIFKKSKPRYHKRTSSAVWNSPPLQGARVPSSLAYSLKKHKL"
     CDS             3871..4590
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     misc_feature    4604..4622
                     /label=MCS
     primer_bind     4634..4651
                     /label=BGH reverse primer
     polyA_site      4640..4864
                     /label=BGH pA
     rep_origin      4910..5338
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        5352..5681
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5532..5667
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        5729..5776
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             5795..6169
                     /codon_start=1
                     /label=Zeo(R)
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    6299..6420
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6469..6485)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    6493..6509
                     /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(6517..6547)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6562..6583
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(6871..7456)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(7627..8487)
                     /codon_start=1
                     /label=Amp(R)
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(8488..8592)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggaacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc
      841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga
      901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga
      961 ctctagcgtt taaacttaag cttggtaccg agctcggatc tcgagctcaa gcttggtacc
     1021 gccaccatgt acaccgacat cgagatgaac cggttgggca aggggacgac ggccccaggg
     1081 cccattcacc tgctggagct ctgtgaccag aagctcatgg agtttctctg caacatggat
     1141 aataaggact tggtgtggct tgaggagatc caagaggagg ccgagcgcat gttcaccaga
     1201 gaattcagca aagagccaga gctgatgccc aaaacacctt ctcagaagaa ccgacggaag
     1261 aagagacgga tttcttatgt tcaggatgaa aacagagatc ccatcaggag aaggttatcc
     1321 cgcagaaagt ctcggagcag ccagcagagc acccgacgcc tccgcagcaa ggacagtgta
     1381 gagaagctgg ctacagtggt cggggagaac ggctccgttc tgcggcgtgt gacccgtgct
     1441 gcggctgcag ctgccgcggc taccatggca ttggctgcac cttcttcacc cacccctgag
     1501 tctcccacga tgctgactaa gaagcccgag gataaccaca cccagtgcca gctggtgcct
     1561 gtggtggaga tcggcatcag tgagcgccag aatgctgagc agcatgtcac ccagctcatg
     1621 tccaccgagc ctctgccccg cactctgtcc ccgactccag cttcagccac agctccaacc
     1681 tcccagggca tcccgacatc agatgaggaa tcaacaccta agaagtcgaa ggccaggata
     1741 ctggagtcca tcacagtgag ctccctgatg gctacacccc aggaccccaa gggtcaaggg
     1801 gtcgggacgg ggcggtctgc gtctaagctc aggattgcgc aggtctcccc tggcccacgg
     1861 gactcgccag cctttccaga ttctccatgg cgggagcggg tgctggctcc catcctgccg
     1921 gataacttct ccacgcccac gggctctcgc acggactctc aatcggtgcg gcacagcccg
     1981 atcgccccgt cttccccgag tccccgagtc ttagcccaga agtactctct ggtggccaaa
     2041 caggaaagtg ttgtccgcag ggcgagcaga aggcttgcca agaagactgc cgaagagcca
     2101 gctgcctctg gccgcatcat ctgtcacagt tacctggaga ggctcctgaa tgttgaggtg
     2161 ccccagaaag ttggttctga gcagaaggaa ccccccgagg aggctgagcc tgtggcggca
     2221 gctgagccag aggtccctga gaacaacgga aataactcgt ggccccacaa tgacacagag
     2281 atcgccaaca gcacacccaa cccgaagcct gcagccagca gcccggaaac accctctgca
     2341 gggcagcaag aggccaagac ggaccaagca gatggaccca gagagccacc gcagagtgcc
     2401 aggaggaagc gcagctacaa gcaggccgtg agtgagctgg acgaggagca gcacctggag
     2461 gatgaggagc tgcagccccc caggagcaag accccttcct caccctgccc agccagcaag
     2521 gtggtacggc ccctccggac ctttctgcac acagtgcaga ggaaccagat gctcacgacc
     2581 ccgacctcag ccccacgcag cgtcatgaag tcctttatta agcgcaacac tcccctgcgc
     2641 atggacccca agtgcagctt cgtcgagaag gagcggcagc gcctggagaa tctgcggcgg
     2701 aaggaggagg ccgagcagct gcgcaggcag aaggtggagg aggacaagcg gcggcggctg
     2761 gaggaggtga agctgaagcg tgaggaacgc ctccgcaagg tgctgcaggc ccgcgagcgg
     2821 gtggagcaga tgaaggagga gaagaaaaag caaattgagc agaagtttgc tcagatcgac
     2881 gagaagactg agaaggccaa ggaggagcgg ctggcagagg agaaggccaa gaaaaaggcg
     2941 gcggccaaga agatggagga ggtggaagca cgcaggaagc aggaagagga tgcacgtagg
     3001 ctcaggtggc tgcagcagga ggaggaagag cggcggcacc aagagctgct gcagaagaag
     3061 aaggaagagg agcaggagcg gctgcggaag gcggccgagg ctaagcggct ggcagagcag
     3121 cgggagcagg agcggcggga gcaggagcgg cgcgagcagg agcggcgcga gcaggagcgg
     3181 cgggagcagg agcggcgcga gcaggagcga cagctggcag agcaggagcg tcggcgggag
     3241 caggagcggc tccaggccga gagggagctg caggagcggg agaaggccct gcggctgcag
     3301 aaggagcagc tgcagaggga actggaggag aagaagaaga aggaagagca gcagcgtctg
     3361 gctgagcggc agctgcagga ggagcaagag aagaaagcca aggaggcagt aggggccagc
     3421 aaggccctga atgtgactgt ggacgtgcag tctccagctt gtacctcata tcaaatgact
     3481 ccgcaagggc acagggcccc tcccaagatc aacccagata actacgggat ggatctgaat
     3541 agcgacgact ccaccgatga tgaggcccat ccccggaagc ccatccccac ctgggcccga
     3601 ggcaccccgc tcagccaggc tatcattcac cagtactacc acccaccgaa ccttctggag
     3661 ctctttggaa ccattctccc actggacttg gaggatatct tcaagaagag caagccccgc
     3721 tatcacaagc gcaccagctc tgctgtctgg aactcaccgc ccctgcaggg cgccagggtc
     3781 cccagcagcc tggcctacag cctgaagaag cacaagcttc gaattctgca gtcgacggta
     3841 ccgcgggccc gggatccacc ggtcgccacc atggtgagca agggcgagga gctgttcacc
     3901 ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg
     3961 tccggcgagg gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc
     4021 accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag
     4081 tgcttcagcc gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc
     4141 gaaggctacg tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc
     4201 gccgaggtga agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac
     4261 ttcaaggagg acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac
     4321 gtctatatca tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa gatccgccac
     4381 aacatcgagg acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc
     4441 gacggccccg tgctgctgcc cgacaaccac tacctgagca cccagtccgc cctgagcaaa
     4501 gaccccaacg agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc
     4561 actctcggca tggacgagct gtacaagtaa agcggccgcg actctagagg gcccgtttaa
     4621 acccgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc
     4681 cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag
     4741 gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag
     4801 gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc ggtgggctct
     4861 atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca cgcgccctgt
     4921 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     4981 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     5041 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     5101 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
     5161 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     5221 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
     5281 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaattaa
     5341 ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa
     5401 gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc
     5461 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc
     5521 taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct
     5581 gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag ctattccaga
     5641 agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg ggagcttgta
     5701 tatccatttt cggatctgat cagcacgtgt tgacaattaa tcatcggcat agtatatcgg
     5761 catagtataa tacgacaagg tgaggaacta aaccatggcc aagttgacca gtgccgttcc
     5821 ggtgctcacc gcgcgcgacg tcgccggagc ggtcgagttc tggaccgacc ggctcgggtt
     5881 ctcccgggac ttcgtggagg acgacttcgc cggtgtggtc cgggacgacg tgaccctgtt
     5941 catcagcgcg gtccaggacc aggtggtgcc ggacaacacc ctggcctggg tgtgggtgcg
     6001 cggcctggac gagctgtacg ccgagtggtc ggaggtcgtg tccacgaact tccgggacgc
     6061 ctccgggccg gccatgaccg agatcggcga gcagccgtgg gggcgggagt tcgccctgcg
     6121 cgacccggcc ggcaactgcg tgcacttcgt ggccgaggag caggactgac acgtgctacg
     6181 agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga
     6241 cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa
     6301 cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa
     6361 taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta
     6421 tcatgtctgt ataccgtcga cctctagcta gagcttggcg taatcatggt catagctgtt
     6481 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa
     6541 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact
     6601 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
     6661 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
     6721 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     6781 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     6841 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     6901 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     6961 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     7021 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     7081 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     7141 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     7201 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     7261 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     7321 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     7381 cggcaaacaa accaccgctg gtagcggttt ttttgtttgc aagcagcaga ttacgcgcag
     7441 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     7501 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     7561 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     7621 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     7681 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     7741 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     7801 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     7861 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     7921 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     7981 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     8041 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     8101 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     8161 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     8221 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     8281 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     8341 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     8401 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     8461 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     8521 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     8581 aggggttccg cgcacatttc cccgaaaagt gccacctgac gtc
//
