LOCUS       sequence-44815-e        7644 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7644)
  AUTHORS   Gregory PA, Bert AG, Paterson EL, Barry SC, Tsykin A, Farshid G, 
            Vadas MA, Khew-Goodall Y, Goodall GJ
  TITLE     The miR-200 family and miR-205 regulate epithelial to mesenchymal 
            transition by targeting ZEB1 and SIP1.
  JOURNAL   Nat Cell Biol. 2008 May;10(5):593-601. Epub 2008 Mar 30.
  PUBMED    18376396
REFERENCE   2  (bases 1 to 7644)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat Cell 
            Biol."; date: "2008-05"; volume: "10(5)"; pages: "593-601. Epub 2008
            Mar 30"
COMMENT     Sequence Label: pCI-neo-RL-ZEB2 200bmut
FEATURES             Location/Qualifiers
     source          1..7644
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        138..517
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        518..729
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     680..700
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     726..750
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          890..1022
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1005..1024
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        1067..1085
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1089..1098
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1095..2030
                     /codon_start=1
                     /product="Renilla luciferase"
                     /label=hRluc
                     /note="human codon-optimized"
                     /translation="MASKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     polyA_signal    3348..3469
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3359..3378)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3413..3432
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      3655..4110
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3742..3761)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3952..3973
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4227..4584
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4227..4418
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4250..4270)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4435..4570
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4635..5429
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4689..4708)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5299..5318
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5493..5541
                     /note="synthetic polyadenylation signal"
     primer_bind     complement(5582..5601)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5701..5723
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5761..5779)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5847..5951
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5952..6020,6021..6812)
                     /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     complement(6170..6189)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6983..7571
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7472..7491
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tcaatattgg ccattagcca tattattcat tggttatata gcataaatca atattggcta
       61 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
      121 aatatgaccg ccatgttggc attgattatt gactagttat taatagtaat caattacggg
      181 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      241 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      301 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      361 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
      421 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
      481 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
      541 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      601 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaacaactg
      661 cgatcgcccg ccccgttgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata
      721 agcagagctc gtttagtgaa ccgtcagatc actagaagct ttattgcggt agtttatcac
      781 agttaaattg ctaacgcagt cagtgcttct gacacaacag tctcgaactt aagctgcagt
      841 gactctctta aggtagcctt gcagaagttg gtcgtgaggc actgggcagg taagtatcaa
      901 ggttacaaga caggtttaag gagaccaata gaaactgggc ttgtcgagac agagaagact
      961 cttgcgtttc tgataggcac ctattggtct tactgacatc cactttgcct ttctctccac
     1021 aggtgtccac tcccagttca attacagctc ttaaggctag agtacttaat acgactcact
     1081 ataggctagc caccatggct tccaaggtgt acgaccccga gcaacgcaaa cgcatgatca
     1141 ctgggcctca gtggtgggct cgctgcaagc aaatgaacgt gctggactcc ttcatcaact
     1201 actatgattc cgagaagcac gccgagaacg ccgtgatttt tctgcatggt aacgctgcct
     1261 ccagctacct gtggaggcac gtcgtgcctc acatcgagcc cgtggctaga tgcatcatcc
     1321 ctgatctgat cggaatgggt aagtccggca agagcgggaa tggctcatat cgcctcctgg
     1381 atcactacaa gtacctcacc gcttggttcg agctgctgaa ccttccaaag aaaatcatct
     1441 ttgtgggcca cgactggggg gcttgtctgg cctttcacta ctcctacgag caccaagaca
     1501 agatcaaggc catcgtccat gctgagagtg tcgtggacgt gatcgagtcc tgggacgagt
     1561 ggcctgacat cgaggaggat atcgccctga tcaagagcga agagggcgag aaaatggtgc
     1621 ttgagaataa cttcttcgtc gagaccatgc tcccaagcaa gatcatgcgg aaactggagc
     1681 ctgaggagtt cgctgcctac ctggagccat tcaaggagaa gggcgaggtt agacggccta
     1741 ccctctcctg gcctcgcgag atccctctcg ttaagggagg caagcccgac gtcgtccaga
     1801 ttgtccgcaa ctacaacgcc taccttcggg ccagcgacga tctgcctaag atgttcatcg
     1861 agtccgaccc tgggttcttt tccaacgcta ttgtcgaggg agctaagaag ttccctaaca
     1921 ccgagttcgt gaaggtgaag ggcctccact tcagccagga ggacgctcca gatgaaatgg
     1981 gtaagtacat caagagcttc gtggagcgcg tgctgaagaa cgagcagtaa ttctagtgga
     2041 gttggagctg ggtattgtta aaaactgcat tatgcaaaaa ttttgtacag tgttaaggcc
     2101 taaaaactgt gtggttcaga gactaattcc tgtgtttaat agcatttata ctttaagcac
     2161 aactagaaaa ttgtaagaat tgcactctac ttatgtatca ctacaaactt taaaaaacta
     2221 tgtctaattt atattaatac attttaaaaa ggtgcccgca ctaccataca tcaggacttt
     2281 tattattatt attgttattc ctttttaatt taatgtgctc gcactacaat gcatcaggac
     2341 tatgattcct ctgtactttc ctttcgctat tcatcaattt cccatttttt ttttcagctt
     2401 aagtaaccac acaattttag gcctcaattt tttttttttt ctgtgaagga acttgaagtg
     2461 atgcatgtgt gaatttaaga taccgaagtc ttaaagtgac ctggacgtga aggaaaaagt
     2521 aagatgagaa ataaagaaag cctttgtaag gtggttttaa aagccttata tgcaaacctt
     2581 ttaatctgtg tttctgcaag tgccatcctt gtacagtgtt aagagggtaa catgggttac
     2641 ctttgcacca gcttcagtgt taagctcacc ctgttctttg aagcacccat gtcagcagta
     2701 gaagaatagg cagcagttcc ttagtttaca tatgtttgtg caattatttt ctgtactttt
     2761 ttgttcatta attttgtcag cagtacacca aactgttttt gcaacaaaaa aatttttttt
     2821 gcattcattt aattttaggt caaataacat tttatttatg tggctcattt tatatttcct
     2881 aattttattt atttcatact gtagtgtaca gcagtatagt tcttcaatat atagatatat
     2941 tttagtaaaa aaggaacatg acgttgatca tttgggcaaa ttttacgtaa agagaagagc
     3001 atttattgtg ttttggaaca ttaattgtga gatgggattt ttcaatttta ttattttatt
     3061 tttgtttttt tccaattact ggaaattcca aatttgggaa cttttgatac gatcttgtga
     3121 aaacactgta ttttcgactg aaaattccac tttcttcatc ttgtttttta gctaaaaaga
     3181 gggactgtta aatacaatgt atgataccat gacaaaaatc tttcctgaat tgtctttgta
     3241 aaaggactat tgaattttca atttgtaatt tcttttgaaa atgaccatgc tcgaaactag
     3301 agcggccgct tccctttagt gagggttaat gcttcgagca gacatgataa gatacattga
     3361 tgagtttgga caaaccacaa ctagaatgca gtgaaaaaaa tgctttattt gtgaaatttg
     3421 tgatgctatt gctttatttg taaccattat aagctgcaat aaacaagtta acaacaacaa
     3481 ttgcattcat tttatgtttc aggttcaggg ggagatgtgg gaggtttttt aaagcaagta
     3541 aaacctctac aaatgtggta aaatccgata aggatcgatc cgggctggcg taatagcgaa
     3601 gaggcccgca ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga atggacgcgc
     3661 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac
     3721 ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg
     3781 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt
     3841 tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc
     3901 cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct
     3961 tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga
     4021 ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga
     4081 attttaacaa aatattaacg cttacaattt cctgatgcgg tattttctcc ttacgcatct
     4141 gtgcggtatt tcacaccgca tacgcggatc tgcgcagcac catggcctga aataacctct
     4201 gaaagaggaa cttggttagg taccttctga ggcggaaaga accagctgtg gaatgtgtgt
     4261 cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
     4321 ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     4381 caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg
     4441 cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt
     4501 tatgcagagg ccgaggccgc ctcggcctct gagctattcc agaagtagtg aggaggcttt
     4561 tttggaggcc taggcttttg caaaaagctt gattcttctg acacaacagt ctcgaactta
     4621 aggctagagc caccatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg
     4681 tggagaggct attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg
     4741 tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg
     4801 ccctgaatga actgcaggac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc
     4861 cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg
     4921 aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca
     4981 tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc
     5041 aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg
     5101 atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg
     5161 cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata
     5221 tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg
     5281 accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat
     5341 gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct
     5401 tctatcgcct tcttgacgag ttcttctgag cgggactctg gggttcgaaa tgaccgacca
     5461 agcgacgccc aacctgccat cacgatggcc gcaataaaat atctttattt tcattacatc
     5521 tgtgtgttgg ttttttgtgt gaatcgatag cgataaggat ccgcgtatgg tgcactctca
     5581 gtacaatctg ctctgatgcc gcatagttaa gccagccccg acacccgcca acacccgctg
     5641 acgcgccctg acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct
     5701 ccgggagctg catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg
     5761 gcctcgtgat acgcctattt ttataggtta atgtcatgat aataatggtt tcttagacgt
     5821 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     5881 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     5941 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
     6001 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
     6061 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
     6121 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
     6181 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc
     6241 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
     6301 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
     6361 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
     6421 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
     6481 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
     6541 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
     6601 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
     6661 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
     6721 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
     6781 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     6841 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     6901 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     6961 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     7021 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     7081 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt
     7141 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     7201 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     7261 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     7321 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     7381 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     7441 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     7501 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     7561 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     7621 ttgctcacat ggctcgacag atct
//
