LOCUS       sequence-222555-        5167 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 5167)
  AUTHORS   Abbate J, Lacayo JC, Prichard M, Pari G, McVoy MA
  TITLE     Bifunctional protein conferring enhanced green fluorescence and 
            puromycin resistance.
  JOURNAL   Biotechniques. 2001 Aug;31(2):336-40.
  PUBMED    11515370
REFERENCE   2  (bases 1 to 5167)
  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: 
            "Biotechniques."; date: "2001-08"; volume: "31(2)"; pages: "336-40"
COMMENT     Sequence Label: pEGFP-puro
FEATURES             Location/Qualifiers
     source          1..5167
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        71..374
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        375..578
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     529..549
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      617..626
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(623..625,626..628,629..1339)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(623..625,626..628,629..1339)
                     /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(668..689)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(929..948)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1276..1297
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1370..1966
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(1370..1389)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1866..1886
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    2096..2217
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2133..2152)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2187..2206
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2234..2250)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2235..2251)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     complement(2275..2294)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2276..2294)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2304..2321)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2304..2320)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2313..2335)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      complement(2461..2917)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2599..2620)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2811..2830
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2944..3048
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3049..3117,3118..3909)
                     /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     complement(3267..3286)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4080..4668
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4569..4588
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4822..4839
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4956..4977
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4992..4997,4998..5015,5016..5022)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5030..5046
                     /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)."
     primer_bind     5035..5057
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5054..5070
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5054..5070
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     5089..5109
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        5091..5109
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     5146..5162
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5146..5162
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
ORIGIN
        1 ggctgcagga attcgattaa tagtaatcaa ttacggggtc attagttcat agcccatata
       61 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      121 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      181 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      241 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      301 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      361 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      421 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      481 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      541 gtaggcgtgt acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc
      601 gctagcgcta ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt
      661 gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga
      721 gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa
      781 gctgcccgtg ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag
      841 ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta
      901 cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt
      961 gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga
     1021 ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat
     1081 catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga
     1141 ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc
     1201 cgtgctgctg cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa
     1261 cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg
     1321 catggacgag ctgtacaagt ccggactcag atctaggaga cgaccttcca tgaccgagta
     1381 caagcccacg gtgcgcctcg ccacccgcga cgacgtcccc cgggccgtac gcaccctcgc
     1441 cgccgcgttc gccgactacc ccgccacgcg ccacaccgtc gacccggacc gccacatcga
     1501 gcgggtcacc gagctgcaag aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt
     1561 gtgggtcgcg gacgacggcg ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc
     1621 gggggcggtg ttcgccgaga tcggcccgcg catggccgag ttgagcggtt cccggctggc
     1681 cgcgcagcaa cagatggaag gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt
     1741 cctggccacc gtcggcgtct cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt
     1801 gctccccgga gtggaggcgg ccgagcgcgc cggggtgccc gccttcctgg agacctccgc
     1861 gccccgcaac ctccccttct acgagcggct cggcttcacc gtcaccgccg acgtcgaggt
     1921 gcccgaagga ccgcgcacct ggtgcatgac ccgcaagccc ggtgcgggat ccaccggatc
     1981 tagataactg atcataatca gccataccac atttgtagag gttttacttg ctttaaaaaa
     2041 cctcccacac ctccccctga acctgaaaca taaaatgaat gcaattgttg ttgttaactt
     2101 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
     2161 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttaacg
     2221 cgatcaagct tatcgatacc gtcgacctcg agggggggcc cggtacccaa ttcgccctat
     2281 agtgagtcgt attacgcgcg ctcactggcc gtcgttttac aacgtcgtga ctgggaaaac
     2341 cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag ctggcgtaat
     2401 agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg
     2461 aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat
     2521 ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag
     2581 atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc
     2641 aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc
     2701 taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc
     2761 ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa
     2821 gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc
     2881 acacccgccg cgcttaatgc gccgctacag ggcgcgtcag gtggcacttt tcggggaaat
     2941 gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
     3001 agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa
     3061 catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac
     3121 ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac
     3181 atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt
     3241 ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc
     3301 gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca
     3361 ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc
     3421 ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag
     3481 gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa
     3541 ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg
     3601 gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa
     3661 ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg
     3721 gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt
     3781 gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt
     3841 caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag
     3901 cattggtaac tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat
     3961 ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct
     4021 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     4081 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     4141 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     4201 agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc
     4261 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     4321 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     4381 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     4441 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
     4501 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     4561 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     4621 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     4681 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg
     4741 ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc
     4801 cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcccaata
     4861 cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt
     4921 cccgactgga aagcgggcag tgagcgcaac gcaattaatg tgagttagct cactcattag
     4981 gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga
     5041 taacaatttc acacaggaaa cagctatgac catgattacg ccaagcgcgc aattaaccct
     5101 cactaaaggg aacaaaagct ggagctccac cgcggtggcg gccgctctag aactagtgga
     5161 tcccccg
//
