LOCUS       sequence-269409-        3995 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Enhanced Green fluorescent protein under constitutive CAG Promoter 
            for mammalian expression in minimal backbone plasmid.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3995)
  TITLE     Constitutive promoter minimal backbone plasmids for optimized 
            expression in mammalian cells
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 3995)
  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: 
            "Unpublished"
COMMENT     Sequence Label: CAG-eGFP-Z1
FEATURES             Location/Qualifiers
     source          1..3995
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        88..391
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          636..1652
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1576..1598
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1660..1679
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1745..1754
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1751..1753,1754..1756,1757..2470)
                     /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(1751..1753,1754..1756,1757..2467)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1796..1817)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2057..2076)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2404..2425
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2520..2641
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2531..2550)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2585..2604
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(2800..3174)
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     promoter        complement(3184..3202)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(3194..3240)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     rep_origin      complement(3313..3901)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3393..3412)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gcggccgcct gcagaagctt tactaactag ttattaatag taatcaatta cggggtcatt
       61 agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg
      121 ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac
      181 gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt
      241 ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa
      301 atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta
      361 catctacgta ttagtcatcg ctattaccat ggtcgaggtg agccccacgt tctgcttcac
      421 tctccccatc tcccccccct ccccaccccc aattttgtat ttatttattt tttaattatt
      481 ttgtgcagcg atgggggcgg gggggggggg ggcgcgcgcg ggcgaggcgg agaggtgcgg
      541 cggcagccaa tcagagcggc gcgctccgaa agtttccttt tatggcgagg cggcggcggc
      601 ggcggcccta taaaaagcga agcgcgcggc gggcgggagt cgctgcgcgc tgccttcgcc
      661 ccgtgccccg ctccgccgcc gcctcgcgcc gcccgccccg gctctgactg accgcgttac
      721 tcccacaggt gagcgggcgg gacggccctt ctcctccggg ctgtaattag cgcttggttt
      781 aatgacggct tgtttctttt ctgtggctgc gtgaaagcct tgaggggctc cgggagggcc
      841 ctttgtgcgg ggggagcggc tcggggggtg cgtgcgtgtg tgtgtgcgtg gggagcgccg
      901 cgtgcggctc cgcgctgccc ggcggctgtg agcgctgcgg gcgcggcgcg gggctttgtg
      961 cgctccgcag tgtgcgcgag gggagcgcgg ccgggggcgg tgccccgcgg tgcggggggg
     1021 gctgcgaggg gaacaaaggc tgcgtgcggg gtgtgtgcgt gggggggtga gcagggggtg
     1081 tgggcgcgtc ggtcgggctg caaccccccc tgcacccccc tccccgagtt gctgagcacg
     1141 gcccggcttc gggtgcgggg ctccgtacgg ggcgtggcgc ggggctcgcc gtgccgggcg
     1201 gggggtggcg gcaggtgggg gtgccgggcg gggcggggcc gcctcgggcc ggggagggct
     1261 cgggggaggg gcgcggcggc ccccggagcg ccggcggctg tcgaggcgcg gcgagccgca
     1321 gccattgcct tttatggtaa tcgtgcgaga gggcgcaggg acttcctttg tcccaaatct
     1381 gtgcggagcc gaaatctggg aggcgccgcc gcaccccctc tagcgggcgc ggggcgaagc
     1441 ggtgcggcgc cggcaggaag gaaatgggcg gggagggcct tcgtgcgtcg ccgcgccgcc
     1501 gtccccttct ccctctccag cctcggggct gtccgcgggg ggacggctgc cttcgggggg
     1561 gacggggcag ggcggggttc ggcttctggc gtgtgaccgg cggctctaga gcctctgcta
     1621 accatgttca tgccttcttc tttttcctac agctcctggg caacgtgctg gttattgtgc
     1681 tgtctcatca ttttggcaaa gaattctgca gtcgacggta ccgcgggccc gggatccacc
     1741 ggtcgccacc atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt
     1801 cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga
     1861 tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc
     1921 ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc gctaccccga
     1981 ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg
     2041 caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg
     2101 cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat
     2161 cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca tggccgacaa
     2221 gcagaagaac ggcatcaagg tgaacttcaa gatccgccac aacatcgagg acggcagcgt
     2281 gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc
     2341 cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg agaagcgcga
     2401 tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct
     2461 gtacaagtaa agcggccgcg actctagatg tacacatatg gctagctggc cagacatgat
     2521 aagatacatt gatgagtttg gacaaaccac aactagaatg cagtgaaaaa aatgctttat
     2581 ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca ataaacaagt
     2641 taacaacaac aattgcattc attttatgtt tcaggttcag ggggaggtgt gggaggtttt
     2701 ttaaagcaag taaaacctct acaaatgtgg tatggaattc tggactcaca aagaaaaaac
     2761 gcccggtgtg caagaccgag cgttctgaac aatttatcct cagtcctgct cctctgccac
     2821 aaagtgcacg cagttgccgg ccgggtcgcg cagggcgaac tcccgccccc acggctgctc
     2881 gccgatctcg gtcatggccg gcccggaggc gtcccggaag ttcgtggaca cgacctccga
     2941 ccactcggcg tacagctcgt ccaggccgcg cacccacacc caggccaggg tgttgtccgg
     3001 caccacctgg tcctggaccg cgctgatgaa cagggtcacg tcgtcccgga ccacaccggc
     3061 gaagtcgtcc tccacgaagt cccgggagaa cccgagccgg tcggtccaga actcgaccgc
     3121 tccggcgacg tcgcgcgcgg tgagcaccgg aacggcactg gtcaacttgg ccatgatggc
     3181 cctcctatag tgagtcgtat tatactatgc cgatatacta tgccgatgat taattgtcaa
     3241 ggatcctaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     3301 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     3361 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     3421 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     3481 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     3541 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     3601 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     3661 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     3721 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     3781 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     3841 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     3901 agaagatcct ttgatctttt ctacggggtc tgacgctcag tgaaaactca cgttaaggga
     3961 ttttggtcat ggctagttaa ttaacattta aatca
//
