LOCUS       sequence-471570-        4865 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian HA-EGFP expression plasmid, control plasmid for Addgene 
            #237879.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4865)
  AUTHORS   Addgene Research Program
  TITLE     Addgene empty and control mammalian expression backbones
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 4865)
  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            "Unpublished"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 HA-EGFP Puro
FEATURES             Location/Qualifiers
     source          1..4865
                     /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             475..501
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             join(508..510,511..513,514..1224)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(508..510,511..513,514..1224)
                     /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(553..574)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(814..833)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1161..1182
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1234..1290
                     /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             1291..1890
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTLTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(1291..1310)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1787..1807
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    1974..2095
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2011..2030)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2065..2084
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2192..2212)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2192..2210)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2231..2247)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2231..2247)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2244..2266)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2255..2271
                     /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(2279..2285,2286..2303,2304..2309))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2324..2345
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2462..2479)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2633..3221)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2713..2732)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3392..4183,4184..4252))
                     /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     4015..4034
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4253..4357)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4383..4838)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4520..4541)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4732..4751
                     /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 catgtaccca
      481 tacgatgttc cggattatgc tttcgaaatg gtgagcaagg gcgaggagct gttcaccggg
      541 gtggtgccca tcctggtcga gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc
      601 ggcgagggcg agggcgatgc cacctacggc aagctgaccc tgaagttcat ctgcaccacc
      661 ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc
      721 ttcagccgct accccgacca catgaagcag cacgacttct tcaagtccgc catgcccgaa
      781 ggctacgtcc aggagcgcac catcttcttc aaggacgacg gcaactacaa gacccgcgcc
      841 gaggtgaagt tcgagggcga caccctggtg aaccgcatcg agctgaaggg catcgacttc
      901 aaggaggacg gcaacatcct ggggcacaag ctggagtaca actacaacag ccacaacgtc
      961 tatatcatgg ccgacaagca gaagaacggc atcaaggtga acttcaagat ccgccacaac
     1021 atcgaggacg gcagcgtgca gctcgccgac cactaccagc agaacacccc catcggcgac
     1081 ggccccgtgc tgctgcccga caaccactac ctgagcaccc agtccgccct gagcaaagac
     1141 cccaacgaga agcgcgatca catggtcctg ctggagttcg tgaccgccgc cgggatcact
     1201 ctcggcatgg acgagctgta caagggatcc ggcgcaacaa acttctctct gctgaaacaa
     1261 gccggagatg tcgaagagaa tcctggaccg atgaccgagt acaagcccac ggtgcgcctc
     1321 gccacccgcg acgacgtccc cagggccgta cgcaccctcg ccgccgcgtt cgccgactac
     1381 cccgccacgc gccacaccgt cgatccggac cgccacatcg agcgggtcac cgagctgcaa
     1441 gaactcttcc tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc
     1501 gcagcggtgg cggtctggac cacgccggag agcgtcgaag cgggggcggt gttcgccgag
     1561 atcggcccgc gcatggccga gttgagcggt tcccggctgg ccgcgcagca acagatggaa
     1621 ggcctcctgg caccgcaccg gcccaaggag cccgcgtggt tcctggccac cgtcggcgtc
     1681 tcgcccgacc accagggcaa gggtctgggc agcgccgtcg tgctccccgg agtggaggcg
     1741 gccgagcgcg ccggggtgcc cgccttcctg gagacctccg cgccccgcaa cctccccttc
     1801 tacgagcggc tcggcttcac cctcaccgcc gacgtcgagg tgcccgaagg accgcgcacc
     1861 tggtgcatga cccgcaagcc cggtgcctga acgcgtgtag tagttcaagt catcttatta
     1921 ttcagtattt ataacttgca aagaaatgaa tatcagagag tgagaggtgt acaaacttgt
     1981 ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag
     2041 catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta tcttaactag
     2101 tggcgccaag ccgaattctg cagatatcca tcacactggc ggccgctcga ctagagcggc
     2161 cgccaccgcg gtggagctcc agcttttgtt ccctttagtg agggttaatt gcgcgcttgg
     2221 cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca
     2281 acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca
     2341 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
     2401 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
     2461 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     2521 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     2581 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     2641 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     2701 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     2761 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     2821 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     2881 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     2941 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     3001 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     3061 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     3121 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     3181 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     3241 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     3301 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     3361 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     3421 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     3481 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
     3541 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     3601 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     3661 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
     3721 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     3781 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     3841 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     3901 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     3961 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
     4021 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
     4081 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
     4141 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
     4201 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
     4261 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
     4321 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
     4381 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
     4441 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
     4501 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
     4561 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
     4621 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
     4681 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
     4741 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
     4801 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg
     4861 caact
//
