LOCUS       sequence-467524-        5561 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes GFP. For use in transfections as a negative control without 
            epitope tags..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5561)
  AUTHORS   Zhang Z, Xu A, Bai Y, Chen Y, Cates K, Kerr C, Bermudez A, Susanto 
            TT, Wysong K, Garcia Marques FJ, Nolan GP, Pitteri S, Barna M
  TITLE     A subcellular map of translational machinery composition and 
            regulation at the single-molecule level.
  JOURNAL   Science. 2025 Mar 7;387(6738):eadn2623. doi: 
            10.1126/science.adn2623. Epub 2025 Mar 7.
  PUBMED    40048539
REFERENCE   2  (bases 1 to 5561)
  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."; 
            date: "2025-03-7"; volume: "387(6738):eadn2623. doi"; pages: " 
            10.1126/science.adn2623. Epub 2025 Mar 7"
COMMENT     Sequence Label: pCAGEN_GFP
FEATURES             Location/Qualifiers
     source          1..5561
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        5..384
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        386..661
                     /label=chicken beta-actin promoter
     intron          662..1678
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1602..1624
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1686..1705
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1771..1780
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1777..1779,1780..1782,1783..2496)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1777..1779,1780..1782,1783..2493)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1822..1843)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2083..2102)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2430..2451
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2583..2602)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2648..2703
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2702..2721)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3064..3080)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3064..3080)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3077..3099)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3088..3104
                     /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(3112..3118,3119..3136,3137..3142))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3157..3178
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3237..3432
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3283..3418
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    3438..3572
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3475..3494)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3529..3548
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3640..3657)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3811..4399)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3891..3910)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4570..5361,5362..5430))
                     /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     5193..5212
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5431..5535)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggtcgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat
       61 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg
      121 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata
      181 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta
      241 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc
      301 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac
      361 gtattagtca tcgctattac catggtcgag gtgagcccca cgttctgctt cactctcccc
      421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
      481 gcgatggggg cggggggggg gggggggcgc gccaggcggg gcggggcggg gcgaggggcg
      541 gggcggggcg aggcggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
      601 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
      661 gggagtcgct gcgcgctgcc ttcgccccgt gccccgctcc gccgccgcct cgcgccgccc
      721 gccccggctc tgactgaccg cgttactccc acaggtgagc gggcgggacg gcccttctcc
      781 tccgggctgt aattagcgct tggtttaatg acggcttgtt tcttttctgt ggctgcgtga
      841 aagccttgag gggctccggg agggcccttt gtgcgggggg agcggctcgg ggggtgcgtg
      901 cgtgtgtgtg tgcgtgggga gcgccgcgtg cggctccgcg ctgcccggcg gctgtgagcg
      961 ctgcgggcgc ggcgcggggc tttgtgcgct ccgcagtgtg cgcgagggga gcgcggccgg
     1021 gggcggtgcc ccgcggtgcg gggggggctg cgaggggaac aaaggctgcg tgcggggtgt
     1081 gtgcgtgggg gggtgagcag ggggtgtggg cgcgtcggtc gggctgcaac cccccctgca
     1141 cccccctccc cgagttgctg agcacggccc ggcttcgggt gcggggctcc gtacggggcg
     1201 tggcgcgggg ctcgccgtgc cgggcggggg gtggcggcag gtgggggtgc cgggcggggc
     1261 ggggccgcct cgggccgggg agggctcggg ggaggggcgc ggcggccccc ggagcgccgg
     1321 cggctgtcga ggcgcggcga gccgcagcca ttgcctttta tggtaatcgt gcgagagggc
     1381 gcagggactt cctttgtccc aaatctgtgc ggagccgaaa tctgggaggc gccgccgcac
     1441 cccctctagc gggcgcgggg cgaagcggtg cggcgccggc aggaaggaaa tgggcgggga
     1501 gggccttcgt gcgtcgccgc gccgccgtcc ccttctccct ctccagcctc ggggctgtcc
     1561 gcggggggac ggctgccttc gggggggacg gggcagggcg gggttcggct tctggcgtgt
     1621 gaccggcggc tctagagcct ctgctaacca tgttcatgcc ttcttctttt tcctacagct
     1681 cctgggcaac gtgctggtta ttgtgctgtc tcatcatttt ggcaaagaat tctgcagtcg
     1741 acggtaccgc gggcccggga tccaccggtc gccaccatgg tgagcaaggg cgaggagctg
     1801 ttcaccgggg tggtgcccat cctggtcgag ctggacggcg acgtaaacgg ccacaagttc
     1861 agcgtgtccg gcgagggcga gggcgatgcc acctacggca agctgaccct gaagttcatc
     1921 tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct gacctacggc
     1981 gtgcagtgct tcagccgcta ccccgaccac atgaagcagc acgacttctt caagtccgcc
     2041 atgcccgaag gctacgtcca ggagcgcacc atcttcttca aggacgacgg caactacaag
     2101 acccgcgccg aggtgaagtt cgagggcgac accctggtga accgcatcga gctgaagggc
     2161 atcgacttca aggaggacgg caacatcctg gggcacaagc tggagtacaa ctacaacagc
     2221 cacaacgtct atatcatggc cgacaagcag aagaacggca tcaaggtgaa cttcaagatc
     2281 cgccacaaca tcgaggacgg cagcgtgcag ctcgccgacc actaccagca gaacaccccc
     2341 atcggcgacg gccccgtgct gctgcccgac aaccactacc tgagcaccca gtccgccctg
     2401 agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc
     2461 gggatcactc tcggcatgga cgagctgtac aagtaaagcg gccgcactcc tcaggtgcag
     2521 gctgcctatc agaaggtggt ggctggtgtg gccaatgccc tggctcacaa ataccactga
     2581 gatctttttc cctctgccaa aaattatggg gacatcatga agccccttga gcatctgact
     2641 tctggctaat aaaggaaatt tattttcatt gcaatagtgt gttggaattt tttgtgtctc
     2701 tcactcggaa ggacatatgg gagggcaaat catttaaaac atcagaatga gtatttggtt
     2761 tagagtttgg caacatatgc ccatatgctg gctgccatga acaaaggttg gctataaaga
     2821 ggtcatcagt atatgaaaca gccccctgct gtccattcct tattccatag aaaagccttg
     2881 acttgaggtt agattttttt tatattttgt tttgtgttat ttttttcttt aacatcccta
     2941 aaattttcct tacatgtttt actagccaga tttttcctcc tctcctgact actcccagtc
     3001 atagctgtcc ctcttctctt atggagatcc ctcgacctgc agcccaagct tggcgtaatc
     3061 atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg
     3121 agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat
     3181 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc ggatccgcat
     3241 ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg
     3301 cccagttccg cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc
     3361 gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt tggaggccta
     3421 ggcttttgca aaaagctaac ttgtttattg cagcttataa tggttacaaa taaagcaata
     3481 gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca
     3541 aactcatcaa tgtatcttat catgtctgga tccgctgcat taatgaatcg gccaacgcgc
     3601 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
     3661 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     3721 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     3781 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     3841 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     3901 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     3961 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     4021 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     4081 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     4141 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     4201 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     4261 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     4321 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     4381 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     4441 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     4501 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     4561 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     4621 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     4681 atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc
     4741 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     4801 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     4861 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     4921 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     4981 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     5041 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     5101 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     5161 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     5221 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     5281 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     5341 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     5401 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     5461 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     5521 aataggggtt ccgcgcacat ttccccgaaa agtgccacct g
//
