LOCUS       sequence-438461-        4731 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 4731)
  TITLE     Monomeric EGFP Plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 4731)
  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            "Unpublished"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: pmEGFP-C1
FEATURES             Location/Qualifiers
     source          1..4731
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(238..257)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     585..606
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    649..714
                     /label=MCS
                     /note="multiple cloning site"
     polyA_signal    838..959
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(875..894)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     929..948
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(966..1421)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1103..1124)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1315..1334
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        1448..1552
                     /gene="bla"
                     /label=AmpR promoter
     promoter        1554..1911
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        1554..1745
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(1577..1597)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      1762..1897
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             1946..2740
                     /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             1946..2740
                     /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(2000..2019)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2610..2629
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(2928..2947)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    2972..3019
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      3348..3936
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3837..3856
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        4111..4414
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4415..4618
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4569..4589
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      4657..4666
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(4663..4665,4666..4668,4669..4731,1..648)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4708..4729)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
ORIGIN
        1 aacggccaca agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg
       61 accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc
      121 accctgacct acggcgtgca gtgcttcagc cgctaccccg accacatgaa gcagcacgac
      181 ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac
      241 gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc
      301 atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag
      361 tacaactaca acagccacaa cgtctatatc atggccgaca agcagaagaa cggcatcaag
      421 gtgaacttca agatccgcca caacatcgag gacggcagcg tgcagctcgc cgaccactac
      481 cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc
      541 acccagtcca agctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag
      601 ttcgtgaccg ccgccgggat cactctcggc atggacgagc tgtacaagtc cggactcaga
      661 tctcgagctc aagcttcgaa ttctgcagtc gacggtaccg cgggcccggg atccaccgga
      721 tctagataac tgatcataat cagccatacc acatttgtag aggttttact tgctttaaaa
      781 aacctcccac acctccccct gaacctgaaa cataaaatga atgcaattgt tgttgttaac
      841 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
      901 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttaa
      961 cgcgtaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag
     1021 ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac
     1081 cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga
     1141 ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc
     1201 accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg
     1261 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa
     1321 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac
     1381 caccacaccc gccgcgctta atgcgccgct acagggcgcg tcaggtggca cttttcgggg
     1441 aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
     1501 catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga gtcctgaggc
     1561 ggaaagaacc agctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca
     1621 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc
     1681 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata
     1741 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
     1801 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc ggcctctgag
     1861 ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa agatcgatca
     1921 agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc
     1981 ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc
     2041 tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga
     2101 cctgtccggt gccctgaatg aactgcaaga cgaggcagcg cggctatcgt ggctggccac
     2161 gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct
     2221 gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa
     2281 agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc
     2341 attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct
     2401 tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc
     2461 caggctcaag gcgagcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg
     2521 cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct
     2581 gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct
     2641 tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca
     2701 gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gcgggactct ggggttcgaa
     2761 atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac cgccgccttc
     2821 tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc
     2881 ggggatctca tgctggagtt cttcgcccac cctaggggga ggctaactga aacacggaag
     2941 gagacaatac cggaaggaac ccgcgctatg acggcaataa aaagacagaa taaaacgcac
     3001 ggtgttgggt cgtttgttca taaacgcggg gttcggtccc agggctggca ctctgtcgat
     3061 accccaccga gaccccattg gggccaatac gcccgcgttt cttccttttc cccaccccac
     3121 cccccaagtt cgggtgaagg cccagggctc gcagccaacg tcggggcggc aggccctgcc
     3181 atagcctcag gttactcata tatactttag attgatttaa aacttcattt ttaatttaaa
     3241 aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt
     3301 tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt
     3361 tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt
     3421 ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag
     3481 ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta
     3541 gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat
     3601 aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg
     3661 ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg
     3721 agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac
     3781 aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga
     3841 aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt
     3901 ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta
     3961 cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat
     4021 tctgtggata accgtattac cgccatgcat tagttattaa tagtaatcaa ttacggggtc
     4081 attagttcat agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc
     4141 tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt
     4201 aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca
     4261 cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg
     4321 taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca
     4381 gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc agtacatcaa
     4441 tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa
     4501 tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta acaactccgc
     4561 cccattgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctgg
     4621 tttagtgaac cgtcagatcc gctagcgcta ccggtcgcca ccatggtgag caagggcgag
     4681 gagctgttca ccggggtggt gcccatcctg gtcgagctgg acggcgacgt a
//
