LOCUS       sequence-435688-        6740 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 6740)
  AUTHORS   Dundr M, Hoffmann-Rohrer U, Hu Q, Grummt I, Rothblum LI, Phair RD, 
            Misteli T
  TITLE     A kinetic framework for a mammalian RNA polymerase in vivo.
  JOURNAL   Science. 2002 Nov 22. 298(5598):1623-6.
  PUBMED    12446911
REFERENCE   2  (bases 1 to 6740)
  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: "Science. 
            2002 Nov 22. 298(5598):1623-6."
COMMENT     Sequence Label: GFP-RRN3/TIF1A
FEATURES             Location/Qualifiers
     source          1..6740
                     /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"
     regulatory      607..616
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(613..615,616..618,619..1329)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(613..615,616..618,619..1329)
                     /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(919..938)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1266..1287
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     regulatory      1370..1379
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    3528..3649
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3565..3584)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3619..3638
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3656..4111)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3793..3814)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4005..4024
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4138..4242
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4244..4601
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4244..4435
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4267..4287)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4452..4587
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4636..5430
                     /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             4636..5430
                     /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(4690..4709)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5300..5319
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5618..5637)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5662..5709
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6038..6626
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6527..6546
                     /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 gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
      661 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
      721 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
      781 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc
      841 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
      901 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
      961 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     1021 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat catggccgac
     1081 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     1141 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     1201 cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc
     1261 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     1321 ctgtacaagt ccggactcag atctcgagct caagcttcga attccggtac catggcggca
     1381 ccgctgcttc acacgcgttt gccgggagat gcggccgctt cgtcctctgc agttaagaag
     1441 ctgggcgcgt cgaggactgg gatttcaaat atgcgtgcat tagagaatga ctttttcaat
     1501 tctcccccaa gaaaaactgt tcggtttggt ggaactgtga cagaagtctt gctgaagtac
     1561 aaaaagggtg aaacaaatga ctttgagttg ttgaagaacc agctgttaga tccagacata
     1621 aaggatgacc agatcatcaa ctggctgcta gaattccgtt cttctatcat gtacttgaca
     1681 aaagactttg agcaacttat cagtattata ttaagattgc cttggttgaa tagaagtcaa
     1741 acagtagtgg aagagtattt ggcttttctt ggtaatcttg tatcagcaca gactgttttc
     1801 ctcagaccgt gtctcagcat gattgcttcc cattttgtgc ctccccgagt gatcattaag
     1861 gaaggcgatg tagatgtttc agattctgat gatgaagatg ataatcttcc tgcaaatttt
     1921 gacacatgtc acagagcctt gcaaataata gcaagatatg taccatcgac accgtggttt
     1981 ctcatgccaa tactggtgga aaaatttcca tttgttcgaa aatcagagag aacactggaa
     2041 tgttacgttc ataacttact aaggattagt gtatatttac caaccttgag gcatgaaatt
     2101 ctggagctta ttattgaaaa actactcaag ttggatgtga atgcatcccg gcagggtatt
     2161 gaagatgctg aagaaacagc aactcaaact tgtggtggga cagattccac ggaaggattg
     2221 tttaatatgg atgaagatga agaaactgaa catgaaacaa aggctggtcc tgaacggctc
     2281 gaccagatgg tgcatcctgt agccgagcgc ctggacatcc tgatgtcttt ggttttgtcc
     2341 tacatgaagg atgtctgcta tgtagatggt aaggttgata acggcaaaac aaaggatcta
     2401 tatcgcgacc tgataaacat ctttgacaaa ctcctgttgc ccacccatgc ctcctgccat
     2461 gtacagtttt tcatgtttta cctctgtagt ttcaaattgg gattcgcaga ggcatttttg
     2521 gaacatctct ggaaaaaatt gcaggaccca agtaatcctg ccatcatcag gcaggctgct
     2581 ggaaattata ttggaagctt tttggcaaga gctaaattta ttcctcttat tactgtaaaa
     2641 tcatgcctag atcttttggt taactggctg cacatatacc ttaataacca ggattcggga
     2701 acaaaggcat tctgcgatgt tgctctccat gggccatttt actcagcctg ccaagctgtg
     2761 ttctacacct ttgtttttag acacaagcag cttttgagcg gaaacctgaa agaaggtttg
     2821 cagtatcttc agagtctgaa ttttgagcgg atagtgatga gccagctaaa tcccctgaag
     2881 atttgcctgc cctcagtggt taactttttt gctgcaatca caaataagta ccagctcgtc
     2941 ttctgctaca ccatcattga gaggaacaat cgccagatgc tgccagtcat taggagtacc
     3001 gctggaggag actcagtgca gatctgcaca aacccgctgg acaccttctt cccctttgat
     3061 ccctgtgtgc tgaagaggtc aaagaaattc attgatccta tttatcaggt gtgggaagac
     3121 atgagtgctg aagagctaca ggagttcaag aaacccatga aaaaggacat agtggaagat
     3181 gaagatgatg actttctgaa aggcgaagtg ccccagaatg ataccgtgat tgggatcaca
     3241 ccaagctcct ttgacacgca tttccgaagt ccttcaagta gtgtgggctc cccacccgtg
     3301 ttgtacatgc aacccagtcc cctctgacgg cagaaatttg tgactgagat gtgacatttg
     3361 ggattcccca tcacttgtca tgccctcagc acccagctgg atccaccgga tctagataac
     3421 tgatcataat cagccatacc acatttgtag aggttttact tgctttaaaa aacctcccac
     3481 acctccccct gaacctgaaa cataaaatga atgcaattgt tgttgttaac ttgtttattg
     3541 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
     3601 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttaa cgcgtaaatt
     3661 gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag ctcatttttt
     3721 aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac cgagataggg
     3781 ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga ctccaacgtc
     3841 aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc accctaatca
     3901 agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg gagcccccga
     3961 tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa
     4021 ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc
     4081 gccgcgctta atgcgccgct acagggcgcg tcaggtggca cttttcgggg aaatgtgcgc
     4141 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     4201 taaccctgat aaatgcttca ataatattga aaaaggaaga gtcctgaggc ggaaagaacc
     4261 agctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa
     4321 gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc
     4381 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc
     4441 taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct
     4501 gactaatttt ttttatttat gcagaggccg aggccgcctc ggcctctgag ctattccaga
     4561 agtagtgagg aggctttttt ggaggcctag gcttttgcaa agatcgatca agagacagga
     4621 tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg
     4681 gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc
     4741 gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt
     4801 gccctgaatg aactgcaaga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt
     4861 ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc
     4921 gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc
     4981 atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac
     5041 caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag
     5101 gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag
     5161 gcgagcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat
     5221 atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg
     5281 gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa
     5341 tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc
     5401 ttctatcgcc ttcttgacga gttcttctga gcgggactct ggggttcgaa atgaccgacc
     5461 aagcgacgcc caacctgcca tcacgagatt tcgattccac cgccgccttc tatgaaaggt
     5521 tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca
     5581 tgctggagtt cttcgcccac cctaggggga ggctaactga aacacggaag gagacaatac
     5641 cggaaggaac ccgcgctatg acggcaataa aaagacagaa taaaacgcac ggtgttgggt
     5701 cgtttgttca taaacgcggg gttcggtccc agggctggca ctctgtcgat accccaccga
     5761 gaccccattg gggccaatac gcccgcgttt cttccttttc cccaccccac cccccaagtt
     5821 cgggtgaagg cccagggctc gcagccaacg tcggggcggc aggccctgcc atagcctcag
     5881 gttactcata tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg
     5941 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact
     6001 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     6061 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     6121 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     6181 ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     6241 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     6301 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     6361 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     6421 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     6481 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     6541 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     6601 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     6661 ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     6721 accgtattac cgccatgcat
//
