LOCUS       sequence-377582-        6742 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression of HPV16 E7 Mutant 2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6742)
  AUTHORS   Nouel J, White EA
  TITLE     ZER1 Contributes to the Carcinogenic Activity of High-Risk HPV E7 
            Proteins.
  JOURNAL   mBio. 2022 Nov 8:e0203322. doi: 10.1128/mbio.02033-22.
  PUBMED    36346242
REFERENCE   2  (bases 1 to 6742)
  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: "mBio. 2022 
            Nov 8:e0203322. doi: 10.1128/mbio.02033-22."
COMMENT     Sequence Label: MSCV-P C-FlagHA HPV16 E7 Mutant 2
FEATURES             Location/Qualifiers
     source          1..6742
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             1..517
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
     misc_feature    581..922
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             989..1405
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1335..1357
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1373..1389
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     primer_bind     1417..1436
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1417..1435
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1465..1489
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(1784..1808)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1840..1863
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1879..1905
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YPYDVPDYA"
     CDS             1879..1905
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     promoter        1918..2417
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(1953..1976)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     2350..2369
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             2438..3037
                     /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(2438..2457)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     2934..2954
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             3137..3651
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    3813..3829
                     /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(3837..3843,3844..3861,3862..3867))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3882..3903
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4020..4037)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4191..4779)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4271..4290)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4950..5741,5742..5810))
                     /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     5573..5592
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5811..5915)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5983..6001
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6039..6061)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6161..6180
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6374..6396
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6524..6543
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 aatgaaagac cccacctgta ggtttggcaa gctagcttaa gtaacgccat tttgcaaggc
       61 atggaaaata cataactgag aatagagaag ttcagatcaa ggttaggaac agagagacag
      121 cagaatatgg gccaaacagg atatctgtgg taagcagttc ctgccccggc tcagggccaa
      181 gaacagatgg tccccagatg cggtcccgcc ctcagcagtt tctagagaac catcagatgt
      241 ttccagggtg ccccaaggac ctgaaatgac cctgtgcctt atttgaacta accaatcagt
      301 tcgcttctcg cttctgttcg cgcgcttctg ctccccgagc tcaataaaag agcccacaac
      361 ccctcactcg gcgcgccagt cctccgatag actgcgtcgc ccgggtaccc gtattcccaa
      421 taaagcctct tgctgtttgc atccgaatcg tggactcgct gatccttggg agggtctcct
      481 cagattgatt gactgcccac ctcgggggtc tttcatttgg aggttccacc gagatttgga
      541 gacccctgcc cagggaccac cgaccccccc gccgggaggt aagctggcca gcggtcgttt
      601 cgtgtctgtc tctgtctttg tgcgtgtttg tgccggcatc taatgtttgc gcctgcgtct
      661 gtactagtta gctaactagc tctgtatctg gcggacccgt ggtggaactg acgagttctg
      721 aacacccggc cgcaaccctg ggagacgtcc cagggacttt gggggccgtt tttgtggccc
      781 gacctgagga agggagtcga tgtggaatcc gaccccgtca ggatatgtgg ttctggtagg
      841 agacgagaac ctaaaacagt tcccgcctcc gtctgaattt ttgctttcgg tttggaaccg
      901 aagccgcgcg tcttgtctgc tgcagcgctg cagcatcgtt ctgtgttgtc tctgtctgac
      961 tgtgtttctg tatttgtctg aaaattaggg ccagactgtt accactccct taagtttgac
     1021 cttaggtcac tggaaagatg tcgagcggat cgctcacaac cagtcggtag atgtcaagaa
     1081 gagacgttgg gttaccttct gctctgcaga atggccaacc tttaacgtcg gatggccgcg
     1141 agacggcacc tttaaccgag acctcatcac ccaggttaag atcaaggtct tttcacctgg
     1201 cccgcatgga cacccagacc aggtccccta catcgtgacc tgggaagcct tggcttttga
     1261 cccccctccc tgggtcaagc cctttgtaca ccctaagcct ccgcctcctc ttcctccatc
     1321 cgccccgtct ctcccccttg aacctcctcg ttcgaccccg cctcgatcct ccctttatcc
     1381 agccctcact ccttctctag gcgccggaat tagatctaat acgactcact atagggagac
     1441 ccaagctggc tagttaagct atcaacaagt ttgtacaaaa aagttggcac catgcatgga
     1501 gatacaccta cattgcatga atatatgtta gatttgcaac cagagacaac tgatctctac
     1561 tgttatgagc aattaaatga cagctcagag gaggaggatg aaatagatgg tccagctgga
     1621 caagcagaac cggacagagc ccattacaat attgtaacct tttgttgcaa gtgtgactct
     1681 acgcttcggt tgtgcgtaca aagcacacac gtagacattc gtactttgga agacctgtta
     1741 atgggcacac taggaattgt gtgccccgcc tgttctcagg cagcaccaac tttcttgtac
     1801 aaagtggttg atctagaggg cccgcggttc gaactcgagg actacaagga cgacgatgac
     1861 aagtcggccg ctggaggata cccctacgac gtgcccgact acgcctagaa ttctaccggg
     1921 taggggaggc gcttttccca aggcagtctg gagcatgcgc tttagcagcc ccgctgggca
     1981 cttggcgcta cacaagtggc ctctggcctc gcacacattc cacatccacc ggtaggcgcc
     2041 aaccggctcc gttctttggt ggccccttcg cgccaccttc tactcctccc ctagtcagga
     2101 agttcccccc cgccccgcag ctcgcgtcgt gcaggacgtg acaaatggaa gtagcacgtc
     2161 tcactagtct cgtgcagatg gacagcaccg ctgagcaatg gaagcgggta ggcctttggg
     2221 gcagcggcca atagcagctt tgctccttcg ctttctgggc tcagaggctg ggaaggggtg
     2281 ggtccggggg cgggctcagg ggcgggctca ggggcggggc gggcgcccga aggtcctccg
     2341 gaggcccggc attctgcacg cttcaaaagc gcacgtctgc cgcgctgttc tcctcttcct
     2401 catctccggg cctttcgacc tgcagcccaa gcttaccatg accgagtaca agcccacggt
     2461 gcgcctcgcc acccgcgacg acgtccccag ggccgtacgc accctcgccg ccgcgttcgc
     2521 cgactacccc gccacgcgcc acaccgtcga tccggaccgc cacatcgagc gggtcaccga
     2581 gctgcaagaa ctcttcctca cgcgcgtcgg gctcgacatc ggcaaggtgt gggtcgcgga
     2641 cgacggcgcc gcggtggcgg tctggaccac gccggagagc gtcgaagcgg gggcggtgtt
     2701 cgccgagatc ggcccgcgca tggccgagtt gagcggttcc cggctggccg cgcagcaaca
     2761 gatggaaggc ctcctggcgc cgcaccggcc caaggagccc gcgtggttcc tggccaccgt
     2821 cggcgtctcg cccgaccacc agggcaaggg tctgggcagc gccgtcgtgc tccccggagt
     2881 ggaggcggcc gagcgcgccg gggtgcccgc cttcctggag acctccgcgc cccgcaacct
     2941 ccccttctac gagcggctcg gcttcaccgt caccgccgac gtcgaggtgc ccgaaggacc
     3001 gcgcacctgg tgcatgaccc gcaagcccgg tgcctgacgc ccgccccacg acccgcagcg
     3061 cccgaccgaa aggagcgcac gaccccatgc atcgataaaa taaaagattt tatttagtct
     3121 ccagaaaaag gggggaatga aagaccccac ctgtaggttt ggcaagctag cttaagtaac
     3181 gccattttgc aaggcatgga aaatacataa ctgagaatag agaagttcag atcaaggtta
     3241 ggaacagaga gacagcagaa tatgggccaa acaggatatc tgtggtaagc agttcctgcc
     3301 ccggctcagg gccaagaaca gatggtcccc agatgcggtc ccgccctcag cagtttctag
     3361 agaaccatca gatgtttcca gggtgcccca aggacctgaa atgaccctgt gccttatttg
     3421 aactaaccaa tcagttcgct tctcgcttct gttcgcgcgc ttctgctccc cgagctcaat
     3481 aaaagagccc acaacccctc actcggcgcg ccagtcctcc gatagactgc gtcgcccggg
     3541 tacccgtgta tccaataaac cctcttgcag ttgcatccga cttgtggtct cgctgttcct
     3601 tgggagggtc tcctctgagt gattgactac ccgtcagcgg gggtctttca tgggtaacag
     3661 tttcttgaag ttggagaaca acattctgag ggtaggagtc gaatattaag taatcctgac
     3721 tcaattagcc actgttttga atccacatac tccaatactc ctgaaatagt tcattatgga
     3781 cagcgcagaa agagctgggg agaattgtga aattgttatc cgctcacaat tccacacaac
     3841 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca
     3901 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     3961 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     4021 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     4081 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     4141 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     4201 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     4261 acaggactat aaagatacca ggcgtttccc cttggaagct ccctcgtgcg ctctcctgtt
     4321 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     4381 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     4441 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     4501 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     4561 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     4621 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     4681 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     4741 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     4801 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     4861 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     4921 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     4981 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     5041 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     5101 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     5161 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     5221 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     5281 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     5341 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     5401 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     5461 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     5521 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     5581 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     5641 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     5701 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     5761 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     5821 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     5881 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     5941 gacgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg tatcacgagg
     6001 ccctttcgtc tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg
     6061 gagacggtca cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg
     6121 tcagcgggtg ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta
     6181 ctgagagtgc accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc
     6241 atcaggcgcc attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc
     6301 tcttcgctat tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta
     6361 acgccagggt tttcccagtc acgacgttgt aaaacgacgg cgcaaggaat ggtgcatgca
     6421 aggagatggc gcccaacagt cccccggcca cggggcctgc caccataccc acgccgaaac
     6481 aagcgctcat gagcccgaag tggcgagccc gatcttcccc atcggtgatg tcggcgatat
     6541 aggcgccagc aaccgcacct gtggcgccgg tgatgccggc cacgatgcgt ccggcgtaga
     6601 ggcgattagt ccaatttgtt aaagacagga tatcagtggt ccaggctcta gttttgactc
     6661 aacaatatca ccagctgaag cctatagagt acgagccata gataaaataa aagattttat
     6721 ttagtctcca gaaaaagggg gg
//
