LOCUS       sequence-65165-e        5335 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 5335)
  TITLE     Craig lab unpublished plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5335)
  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            "Unpublished"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: pEGFPC3/GgVcl 884-1066 (Vt)
FEATURES             Location/Qualifiers
     source          1..5335
                     /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(658..679)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     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"
     primer_bind     complement(1965..1981)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1966..1982)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     polyA_signal    2123..2244
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2160..2179)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2214..2233
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2251..2706)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2388..2409)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2600..2619
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2733..2837
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2839..3196
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2839..3030
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2862..2882)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3047..3182
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3231..4025
                     /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             3231..4025
                     /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(3285..3304)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3895..3914
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4213..4232)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4257..4304
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4633..5221
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5122..5141
                     /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 actcagatct cgagctcaag cttcgaattc tgcaggaatt cccagagcag
     1381 aaagcaggag aagctattaa tcagcccatg atgatggctg ctaggcagtt gcatgacgag
     1441 gctcggaaat ggtctagcaa gggtaacgac atcattgctg ctgctaaacg aatggcgctg
     1501 ctcatggctg agatgtcacg cctggtgcga ggaggcagcg gaaacaagcg tgccctcatc
     1561 cagtgtgcaa aagatattgc taaggcatcg gatgaagtca ctcgattggc caaagaggtg
     1621 gccaagcaat gtactgataa acgcattaga acaaacctct tacaggtctg tgagcgaatc
     1681 ccaaccatca gcacgcagct caaaattctt tccacagtga aagctaccat gctgggcagg
     1741 actaacatca gcgatgaaga atcagaacag gcaactgaga tgttggttca taacgcccag
     1801 aacctcatgc agtctgtgaa ggaaactgtg agagaagctg aagcagcatc cattaagata
     1861 agaacagatg ccggattcac tctgcgctgg gtcagaaaga ccccatggta tcagtaaaca
     1921 ccacataagc attgtcttct gtcggaattc gatatcaagc ttatcgatac cgtcgacctc
     1981 gagggggggc ccgggatcca ccggatctag ataactgatc ataatcagcc ataccacatt
     2041 tgtagaggtt ttacttgctt taaaaaacct cccacacctc cccctgaacc tgaaacataa
     2101 aatgaatgca attgttgttg ttaacttgtt tattgcagct tataatggtt acaaataaag
     2161 caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt
     2221 gtccaaactc atcaatgtat cttaacgcgt aaattgtaag cgttaatatt ttgttaaaat
     2281 tcgcgttaaa tttttgttaa atcagctcat tttttaacca ataggccgaa atcggcaaaa
     2341 tcccttataa atcaaaagaa tagaccgaga tagggttgag tgttgttcca gtttggaaca
     2401 agagtccact attaaagaac gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg
     2461 gcgatggccc actacgtgaa ccatcaccct aatcaagttt tttggggtcg aggtgccgta
     2521 aagcactaaa tcggaaccct aaagggagcc cccgatttag agcttgacgg ggaaagccgg
     2581 cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa
     2641 gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg ccgctacagg
     2701 gcgcgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct
     2761 aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat
     2821 attgaaaaag gaagagtcct gaggcggaaa gaaccagctg tggaatgtgt gtcagttagg
     2881 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
     2941 gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat
     3001 gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac
     3061 tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga
     3121 ggccgaggcc gcctcggcct ctgagctatt ccagaagtag tgaggaggct tttttggagg
     3181 cctaggcttt tgcaaagatc gatcaagaga caggatgagg atcgtttcgc atgattgaac
     3241 aagatggatt gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact
     3301 gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc
     3361 gcccggttct ttttgtcaag accgacctgt ccggtgccct gaatgaactg caagacgagg
     3421 cagcgcggct atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg
     3481 tcactgaagc gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt
     3541 catctcacct tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc
     3601 atacgcttga tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag
     3661 cacgtactcg gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg
     3721 ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgag catgcccgac ggcgaggatc
     3781 tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt
     3841 ctggattcat cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg
     3901 ctacccgtga tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt
     3961 acggtatcgc cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct
     4021 tctgagcggg actctggggt tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg
     4081 agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga
     4141 cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccctag
     4201 ggggaggcta actgaaacac ggaaggagac aataccggaa ggaacccgcg ctatgacggc
     4261 aataaaaaga cagaataaaa cgcacggtgt tgggtcgttt gttcataaac gcggggttcg
     4321 gtcccagggc tggcactctg tcgatacccc accgagaccc cattggggcc aatacgcccg
     4381 cgtttcttcc ttttccccac cccacccccc aagttcgggt gaaggcccag ggctcgcagc
     4441 caacgtcggg gcggcaggcc ctgccatagc ctcaggttac tcatatatac tttagattga
     4501 tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat
     4561 gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat
     4621 caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa
     4681 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa
     4741 ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt agccgtagtt
     4801 aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt
     4861 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata
     4921 gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt
     4981 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
     5041 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
     5101 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
     5161 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
     5221 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
     5281 gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcca tgcat
//
