LOCUS       sequence-47074-e        6440 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 6440)
  AUTHORS   Braun JE, Huntzinger E, Fauser M, Izaurralde E
  TITLE     GW182 proteins directly recruit cytoplasmic deadenylase complexes to
            miRNA targets.
  JOURNAL   Mol Cell. 2011 Oct 7;44(1):120-33.
  PUBMED    21981923
REFERENCE   2  (bases 1 to 6440)
  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: "Mol Cell.";
            date: "2011-10-7"; volume: "44(1)"; pages: "120-33"
COMMENT     Sequence Label: pT7-EGFP-C1-HsNot6L
FEATURES             Location/Qualifiers
     source          1..6440
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     613..632
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        613..631
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      659..668
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(665..667,668..670,671..1381)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(665..667,668..670,671..1381)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(710..731)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(971..990)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1318..1339
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    3228..3349
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3265..3284)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3319..3338
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3356..3811)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3493..3514)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3705..3724
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3838..3942
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3944..4301
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3944..4135
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3967..3987)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4152..4287
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4336..5130
                     /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             4336..5130
                     /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(4390..4409)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5000..5019
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5318..5337)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5362..5409
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5738..6326
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6227..6246
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca ctgcttactg
      601 gcttatcgaa attaatacga ctcactatag ggagacccaa gctctagcgc taccggtcgc
      661 caccatggtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct
      721 ggacggcgac gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac
      781 ctacggcaag ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc
      841 caccctcgtg accaccctga cctacggcgt gcagtgcttc agccgctacc ccgaccacat
      901 gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat
      961 cttcttcaag gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac
     1021 cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg
     1081 gcacaagctg gagtacaact acaacagcca caacgtctat atcatggccg acaagcagaa
     1141 gaacggcatc aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct
     1201 cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa
     1261 ccactacctg agcacccagt ccgccctgag caaagacccc aacgagaagc gcgatcacat
     1321 ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa
     1381 gtccggactc agatctcgag ctagactaat agggatgcca aaggaaaaat atgatcctcc
     1441 agatcctcgc agaatttata ccatcatgtc agcagaggag gtagccaatg ggaaaaaatc
     1501 tcactgggca gaattagaaa tctcgggtag agtgcggagc ctaagtacat cactttggtc
     1561 attgacacac ttgacagcgc tgcacctaaa tgacaattac cttagtcgca ttccacctga
     1621 tattgccaag cttcataatc tggtttacct ggatctgtca tccaataaac tcagaagttt
     1681 accagcagaa ctaggaaaca tggtgtctct cagggaattg cttttaaata acaatctgtt
     1741 acgggttttg ccttatgaac ttggtcggct cttccagcta caaactctag gtttgaaagg
     1801 caatccttta tcacaggata ttctcaactt ataccaggac ccagatggaa cccgaaagct
     1861 actgaacttc atgcttgaca atctcgcagt tcatccagag cagcttcctc cgaggccatg
     1921 gattacatta aaagaacgag accaaattct gccgtcagca tcattcacgg ttatgtgtta
     1981 caatgtgtta tgtgataaat acgctacccg gcagctatat ggctattgcc catcctgggc
     2041 attaaactgg gaatacagga aaaagggaat tatggaagaa attgttaact gtgacgcaga
     2101 tatcattagt cttcaggaag tggaaacaga gcaatacttc actctctttc tgccagcatt
     2161 gaaggagcgt ggatatgatg gatttttttc tccaaagtca cgtgccaaaa tcatgtctga
     2221 gcaggagaga aagcatgtag atggttgtgc aatattcttc aaaacagaaa aatttacatt
     2281 ggtgcagaag catacagtgg aatttaacca agtggcgatg gctaattcag atggatccga
     2341 agctatgctg aacagagtga tgacaaaaga taacattggt gtcgctgtgg tattagaggt
     2401 ccacaaagaa ctatttggag caggtatgaa gcctattcat gctgcagaca aacagctgct
     2461 tatagtggca aatgcccaca tgcattggga cccagagtat tctgatgtga agctcatcca
     2521 gaccatgatg tttgtctcag aggttaaaaa cattctggag aaagcctcta gtaggcctgg
     2581 aagcccaact gcagatccta attccatccc gctggtgcta tgtgcagatc ttaactcatt
     2641 gccagattca ggtgttgtgg aatacttaag caatggagga gtagctgaca accataaaga
     2701 cttcaaggaa ctaaggtaca atgagtgtct tatgaacttc agctgcaatg gaaagaatgg
     2761 aagctcagaa gggagaatca cacatggctt ccaacttaag agcgcctatg aaaataactt
     2821 gatgccttac accaattaca cctttgattt caaaggcgtg attgactaca ttttctattc
     2881 caagactcat atgaacgtgc ttggtgtcct ggggccttta gatcctcaat ggctggttga
     2941 gaacaacatc actgggtgtc cacaccctca catcccttca gaccacttct cactgttaac
     3001 acaacttgaa ctccaccctc cactcctgcc tcttgtcaat ggtgttcact tgcctaatcg
     3061 gaggtaggaa ttctgcagtc gacggtaccg cgggcccggg atccaccgga tctagataac
     3121 tgatcataat cagccatacc acatttgtag aggttttact tgctttaaaa aacctcccac
     3181 acctccccct gaacctgaaa cataaaatga atgcaattgt tgttgttaac ttgtttattg
     3241 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
     3301 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttaa cgcgtaaatt
     3361 gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag ctcatttttt
     3421 aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac cgagataggg
     3481 ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga ctccaacgtc
     3541 aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc accctaatca
     3601 agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg gagcccccga
     3661 tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa
     3721 ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc
     3781 gccgcgctta atgcgccgct acagggcgcg tcaggtggca cttttcgggg aaatgtgcgc
     3841 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     3901 taaccctgat aaatgcttca ataatattga aaaaggaaga gtcctgaggc ggaaagaacc
     3961 agctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa
     4021 gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc
     4081 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc
     4141 taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct
     4201 gactaatttt ttttatttat gcagaggccg aggccgcctc ggcctctgag ctattccaga
     4261 agtagtgagg aggctttttt ggaggcctag gcttttgcaa agatcgatca agagacagga
     4321 tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg
     4381 gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc
     4441 gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt
     4501 gccctgaatg aactgcaaga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt
     4561 ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc
     4621 gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc
     4681 atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac
     4741 caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag
     4801 gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag
     4861 gcgagcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat
     4921 atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg
     4981 gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa
     5041 tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc
     5101 ttctatcgcc ttcttgacga gttcttctga gcgggactct ggggttcgaa atgaccgacc
     5161 aagcgacgcc caacctgcca tcacgagatt tcgattccac cgccgccttc tatgaaaggt
     5221 tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca
     5281 tgctggagtt cttcgcccac cctaggggga ggctaactga aacacggaag gagacaatac
     5341 cggaaggaac ccgcgctatg acggcaataa aaagacagaa taaaacgcac ggtgttgggt
     5401 cgtttgttca taaacgcggg gttcggtccc agggctggca ctctgtcgat accccaccga
     5461 gaccccattg gggccaatac gcccgcgttt cttccttttc cccaccccac cccccaagtt
     5521 cgggtgaagg cccagggctc gcagccaacg tcggggcggc aggccctgcc atagcctcag
     5581 gttactcata tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg
     5641 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact
     5701 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     5761 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     5821 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     5881 ctgtccttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     5941 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     6001 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     6061 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     6121 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     6181 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     6241 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     6301 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     6361 ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     6421 accgtattac cgccatgcat
//
