LOCUS       sequence-469908-        4868 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian EGFP-HA expression plasmid, control plasmid for Addgene 
            #237880.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4868)
  AUTHORS   Addgene Research Program
  TITLE     Addgene empty and control mammalian expression backbones
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 4868)
  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            "Unpublished"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: pAG CMV EGFP-HA Puro
FEATURES             Location/Qualifiers
     source          1..4868
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     100..122
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     114..131
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     115..131
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     141..160
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        141..159
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     183..199
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     184..200
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        218..421
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     372..392
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             join(478..480,481..483,484..1194)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(478..480,481..483,484..1194)
                     /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(523..544)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(784..803)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1131..1152
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1201..1227
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1237..1293
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             1294..1893
                     /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(1294..1313)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1790..1810
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    1977..2098
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2014..2033)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2068..2087
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2195..2215)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2195..2213)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2234..2250)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2234..2250)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2247..2269)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2258..2274
                     /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(2282..2288,2289..2306,2307..2312))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2327..2348
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2465..2482)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2636..3224)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2716..2735)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3395..4186,4187..4255))
                     /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     4018..4037
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4256..4360)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4386..4841)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4523..4544)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4735..4754
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagctggcg aaagggggat
       61 gtgctgcaag gcgattaagt tgggtaacgc cagggttttc ccagtcacga cgttgtaaaa
      121 cgacggccag tgagcgcgcg taatacgact cactataggg cgaatgggta ccggaccccc
      181 cctcgaggtc gacggtatcg ggggcatgct taattaagtg atgcggtttt ggcagtacat
      241 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      301 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaacaactc
      361 cgccccattg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
      421 ttctagaggt ttagtgaacc gtcagatccg ctagcgctac cggtcgccac cttcgaaatg
      481 gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc
      541 gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc
      601 aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc
      661 gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct accccgacca catgaagcag
      721 cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc
      781 aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg
      841 aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag
      901 ctggagtaca actacaacag ccacaacgtc tatatcatgg ccgacaagca gaagaacggc
      961 atcaaggtga acttcaagat ccgccacaac atcgaggacg gcagcgtgca gctcgccgac
     1021 cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac
     1081 ctgagcaccc agtccgccct gagcaaagac cccaacgaga agcgcgatca catggtcctg
     1141 ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta caaggggccc
     1201 tacccatacg atgttccgga ttacgctgga tccggcgcaa caaacttctc tctgctgaaa
     1261 caagccggag atgtcgaaga gaatcctgga ccgatgaccg agtacaagcc cacggtgcgc
     1321 ctcgccaccc gcgacgacgt ccccagggcc gtacgcaccc tcgccgccgc gttcgccgac
     1381 taccccgcca cgcgccacac cgtcgatccg gaccgccaca tcgagcgggt caccgagctg
     1441 caagaactct tcctcacgcg cgtcgggctc gacatcggca aggtgtgggt cgcggacgac
     1501 ggcgcagcgg tggcggtctg gaccacgccg gagagcgtcg aagcgggggc ggtgttcgcc
     1561 gagatcggcc cgcgcatggc cgagttgagc ggttcccggc tggccgcgca gcaacagatg
     1621 gaaggcctcc tggcaccgca ccggcccaag gagcccgcgt ggttcctggc caccgtcggc
     1681 gtctcgcccg accaccaggg caagggtctg ggcagcgccg tcgtgctccc cggagtggag
     1741 gcggccgagc gcgccggggt gcccgccttc ctggagacct ccgcgccccg caacctcccc
     1801 ttctacgagc ggctcggctt caccgtcacc gccgacgtcg aggtgcccga aggaccgcgc
     1861 acctggtgca tgacccgcaa gcccggtgcc tgaacgcgtg tagtagttca agtcatctta
     1921 ttattcagta tttataactt gcaaagaaat gaatatcaga gagtgagagg tgtacaaact
     1981 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     2041 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttaac
     2101 tagtggcgcc aagccgaatt ctgcagatat ccatcacact ggcggccgct cgactagagc
     2161 ggccgccacc gcggtggagc tccagctttt gttcccttta gtgagggtta attgcgcgct
     2221 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     2281 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     2341 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     2401 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
     2461 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     2521 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     2581 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     2641 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     2701 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     2761 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     2821 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     2881 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     2941 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     3001 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     3061 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     3121 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     3181 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     3241 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     3301 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     3361 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     3421 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     3481 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     3541 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     3601 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     3661 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     3721 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     3781 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     3841 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     3901 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     3961 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     4021 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     4081 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     4141 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     4201 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     4261 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     4321 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     4381 cacctaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag
     4441 ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac
     4501 cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga
     4561 ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc
     4621 accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg
     4681 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa
     4741 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac
     4801 caccacaccc gccgcgctta atgcgccgct acagggcgcg tcccattcgc cattcaggct
     4861 gcgcaact
//
