LOCUS       sequence-465433-        9069 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9069)
  TITLE     Hurley Lab Plasmids 2025
REFERENCE   2  (bases 1 to 9069)
  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"
COMMENT     Sequence Label: pCAG/VPS15(R36D)-TSF
FEATURES             Location/Qualifiers
     source          1..9069
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             3177..3197
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             3210..3233
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             3270..3293
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             3300..3323
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(3416..3435)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3481..3536
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3535..3554)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3897..3913)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3897..3913)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3910..3932)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3921..3937
                     /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(3945..3951,3952..3969,3970..3975))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3990..4011
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4070..4265
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      4116..4251
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    4271..4405
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4308..4327)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4362..4381
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4473..4490)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4644..5232)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4724..4743)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5403..6194,6195..6263))
                     /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     6026..6045
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6264..6368)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        6399..6778
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          7070..8086
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     8010..8032
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     8094..8113
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      8160..8169
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 tggaggctga agattacctg aagcagcaga gaggcaacgc cttcccagag atcttctaca
       61 catttctgca gccctatatg gcccagttcg ctaaagagac ttttctgagc gccgacgaac
      121 ggatcctggt catcagaaag gatctgggaa acatcattca taatctgtgc ggccacgacc
      181 tgcccgaaaa agccgagggc gaacctaagg agaatgggct ggtcatcctg gtgtctgtga
      241 ttacaagttg cctgcagact ctgaagtatt gtgatagcaa actggccgct ctggaactga
      301 tcctgcatct ggctccaagg ctgtccgtgg agatcctgct ggacagaatt accccttacc
      361 tgctgcactt ctccaacgat tctgtgccac gggtgagagc cgaggctctg aggacactga
      421 ctaaagtgct ggccctggtg aaggaagtgc ctcgcaacga catcaatatc taccccgagt
      481 atatcctgcc tggcattgcc cacctggctc aggacgatgc cactattgtg aggctggcct
      541 atgctgagaa tatcgccctg ctggctgaaa ccgccctgcg ctttctggag ctggtgcagc
      601 tgaagaacct gaatatggaa aacgacccca acaatgagga aatcgatgag gtgacccatc
      661 ctaacgggaa ttacgacaca gagctgcagg ccctgcacga aatggtgcag cagaaagtgg
      721 tgaccctgct gtccgatcca gaaaacattg tgaagcagac tctgatggag aatgggatca
      781 ccaggctgtg tgtgttcttt ggacgccaga aagctaacga cgtgctgctg tcacatatga
      841 tcacattcct gaacgacaag aatgattggc acctgcgggg cgccttcttt gatagcatcg
      901 tgggagtggc cgcttatgtg ggctggcaga gctcctctat tctgaagcct ctgctgcagc
      961 aggggctgtc tgacgccgag gaatttgtga tcgtgaaagc tctgtatgcc ctgacctgca
     1021 tgtgtcagct gggactgctg cagaagccac atgtgtacga gttcgctagc gatatcgccc
     1081 catttctgtg ccaccccaat ctgtggattc ggtatggggc cgtgggattc atcactgtgg
     1141 tggctagaca gattagcacc gccgacgtgt actgtaaact gatgccatac ctggatccct
     1201 atatcacaca gcccatcatt cagattgaac ggaaactggt gctgctgtcc gtgctgaagg
     1261 agcctgtgtc taggagtatt ttcgactacg ccctgcgcag caaggatatc acttccctgt
     1321 ttagacacct gcatatgagg cagaagaaac gcaacggatc cctgcctgac tgcccacctc
     1381 cagaggatcc agccatcgct cagctgctga agaaactgct gtctcagggg atgaccgagg
     1441 aagaggaaga caagctgctg gccctgaaag atttcatgat gaagtctaac aaggccaaag
     1501 ctaatatcgt ggaccagagt cacctgcatg atagttcaca gaagggcgtg atcgacctgg
     1561 ccgctctggg cattacaggg aggcaggtgg atctggtgaa gactaaacag gagcccgacg
     1621 ataaaagggc ccgcaagcac gtgaaacagg acagcaacgt gaatgaggaa tggaagtcaa
     1681 tgtttggaag cctggacccc cctaatatgc ctcaggctct gccaaaaggc agcgatcagg
     1741 aagtgatcca gaccgggaag ccaccaagat cagagagctc cgccggcatt tgtgtgcccc
     1801 tgagcacatc tagtcaggtg cctgaggtga ccacagtgca gaacaagaaa cctgtgatcc
     1861 cagtgctgtc aagcaccatt ctgccaagca cataccagat caggattact acctgcaaaa
     1921 ccgaactgca gcagctgatc cagcagaagc gggagcagtg taacgctgaa agaattgcca
     1981 agcagatgat ggagaatgcc gagtgggaaa gtaaacctcc accacctgga tggaggccaa
     2041 agggactgct ggtggctcac ctgcatgaac acaaaagcgc cgtgaatcgg atcagagtgt
     2101 ccgacgagca ctctctgttc gccacatgct ccaacgatgg aactgtgaag atttggaatt
     2161 ctcagaagat ggagggcaaa acaactacca caaggtctat cctgacctat agtcggattg
     2221 gaggcagagt gaagaccctg acattttgtc agggcagcca ttacctggcc atcgcttccg
     2281 acaacggggc tgtgcagctg ctgggaattg aagcctcaaa gctgcccaaa agccctaaga
     2341 tccaccccct gcagtccagg attctggacc agaaggagga tggctgcgtg gtggacatgc
     2401 accatttcaa cagtggggcc cagtcagtgc tggcctacgc tacagtgaat gggtctctgg
     2461 tgggatggga tctgcgctcc tctagtaacg cttggactct gaagcatgac ctgaaatccg
     2521 gcctgatcac ctctttcgcc gtggatattc accagtgctg gctgtgtatc ggcacatcaa
     2581 gcgggactat ggcttgctgg gacatgcgct ttcagctgcc aatctcctct cattgtcacc
     2641 caagcagggc taggattagg aggctgtcta tgcaccctct gtatcagagt tgggtcatcg
     2701 ccgctgtgca ggggaacaat gaagtgtcca tgtgggacat ggagaccgga gatcggagat
     2761 tcacactgtg ggctagttca gccccacccc tgtctgagct gcagcctagt ccacattcag
     2821 tgcacgggat ctactgttcc cctgctgatg gaaatccaat cctgctgacc gccggctctg
     2881 acatgaaaat taggttttgg gatctggctt acccagagcg cagttatgtg gtggccggct
     2941 caaccagctc cccctcagtg agctactatc ggaagatcat tgagggcaca gaagtggtgc
     3001 aggaaatcca gaacaagcag aaagtgggac caagtgacga tactccaagg aggggacctg
     3061 agtcactgcc agtgggacac catgacatca ttaccgatgt ggctacattc cagactaccc
     3121 agggcttcat cgtgaccgcc tccagagacg gcatcgtgaa agtgtggaag ggtaccgaaa
     3181 acctgtactt ccaatccggt atggcggcct ggtcacaccc gcagttcgag aaaggtggag
     3241 gtgcccgagg tggatcggga ggtggatcgt ggagccaccc gcagttcgaa aaaggattcg
     3301 attacaagga tgacgatgac aagtagctcg aggaattcac tcctcaggtg caggctgcct
     3361 atcagaaggt ggtggctggt gtggccaatg ccctggctca caaataccac tgagatcttt
     3421 ttccctctgc caaaaattat ggggacatca tgaagcccct tgagcatctg acttctggct
     3481 aataaaggaa atttattttc attgcaatag tgtgttggaa ttttttgtgt ctctcactcg
     3541 gaaggacata tgggagggca aatcatttaa aacatcagaa tgagtatttg gtttagagtt
     3601 tggcaacata tgcccatatg ctggctgcca tgaacaaagg ttggctataa agaggtcatc
     3661 agtatatgaa acagccccct gctgtccatt ccttattcca tagaaaagcc ttgacttgag
     3721 gttagatttt ttttatattt tgttttgtgt tatttttttc tttaacatcc ctaaaatttt
     3781 ccttacatgt tttactagcc agatttttcc tcctctcctg actactccca gtcatagctg
     3841 tccctcttct cttatggaga tccctcgacc tgcagcccaa gcttggcgta atcatggtca
     3901 tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga
     3961 agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg
     4021 cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agcggatccg catctcaatt
     4081 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt
     4141 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg
     4201 cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt
     4261 gcaaaaagct aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac
     4321 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat
     4381 caatgtatct tatcatgtct ggatccgctg cattaatgaa tcggccaacg cgcggggaga
     4441 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     4501 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     4561 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     4621 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     4681 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     4741 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     4801 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     4861 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     4921 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     4981 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     5041 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     5101 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     5161 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     5221 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     5281 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     5341 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     5401 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     5461 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     5521 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     5581 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     5641 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     5701 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     5761 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     5821 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     5881 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     5941 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     6001 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     6061 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     6121 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     6181 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     6241 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     6301 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     6361 gttccgcgca catttccccg aaaagtgcca cctgggtcga cattgattat tgactagtta
     6421 ttaatagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt tccgcgttac
     6481 ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc cattgacgtc
     6541 aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac gtcaatgggt
     6601 ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtac
     6661 gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc agtacatgac
     6721 cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatggt
     6781 cgaggtgagc cccacgttct gcttcactct ccccatctcc cccccctccc cacccccaat
     6841 tttgtattta tttatttttt aattattttg tgcagcgatg ggggcggggg gggggggggg
     6901 gggggggggg gggggggcgc gggggggggg gggggggggg gaggggcggg gcggggcgag
     6961 gcggagaggt gcggcggcag ccaatcagag cggcgcgctc cgaaagtttc cttttatggc
     7021 gaggcggcgg cggcggcggc cctataaaaa gcgaagcgcg cggcgggcgg gagtcgctgc
     7081 gcgctgcctt cgccccgtgc cccgctccgc cgccgcctcg cgccgcccgc cccggctctg
     7141 actgaccgcg ttactcccac aggtgagcgg gcgggacggc ccttctcctc cgggctgtaa
     7201 ttagcgcttg gtttaatgac ggcttgtttc ttttctgtgg ctgcgtgaaa gccttgaggg
     7261 gctccgggag ggccctttgt gcggggggag cggctcgggg ggtgcgtgcg tgtgtgtgtg
     7321 cgtggggagc gccgcgtgcg gctccgcgct gcccggcggc tgtgagcgct gcgggcgcgg
     7381 cgcggggctt tgtgcgctcc gcagtgtgcg cgaggggagc gcggccgggg gcggtgcccc
     7441 gcggtgcggg gggggctgcg aggggaacaa aggctgcgtg cggggtgtgt gcgtgggggg
     7501 gtgagcaggg ggtgtgggcg cgtcggtcgg gctgcaaccc cccctgcacc cccctccccg
     7561 agttgctgag cacggcccgg cttcgggtgc ggggctccgt acggggcgtg gcgcggggct
     7621 cgccgtgccg ggcggggggt ggcggcaggt gggggtgccg ggcggggcgg ggccgcctcg
     7681 ggccggggag ggctcggggg aggggcgcgg cggcccccgg agcgccggcg gctgtcgagg
     7741 cgcggcgagc cgcagccatt gccttttatg gtaatcgtgc gagagggcgc agggacttcc
     7801 tttgtcccaa atctgtgcgg agccgaaatc tgggaggcgc cgccgcaccc cctctagcgg
     7861 gcgcggggcg aagcggtgcg gcgccggcag gaaggaaatg ggcggggagg gccttcgtgc
     7921 gtcgccgcgc cgccgtcccc ttctccctct ccagcctcgg ggctgtccgc ggggggacgg
     7981 ctgccttcgg gggggacggg gcagggcggg gttcggcttc tggcgtgtga ccggcggctc
     8041 tagagcctct gctaaccatg ttcatgcctt cttctttttc ctacagctcc tgggcaacgt
     8101 gctggttatt gtgctgtctc atcattttgg caaagaattc ctcgatcgac ggtatcgaag
     8161 ccaccatggg aaatcagctg gccggcatcg ctcctagcca gattctgtcc gtggaatctt
     8221 acttcagtga catccatgat tttgagtatg acaaatcact gggaagcacc gacttcttta
     8281 aggtggcccg ggctaaacac agagaaggcc tggtggtggt gaaggtgttc gctatccagg
     8341 atcccactct gcctctgacc agttataaac aggagctgga ggaactgaag attagactga
     8401 actcagccca gaattgcctg ccattccaga aagctagcga gaaggcctcc gaaaaagccg
     8461 ctatgctgtt taggcagtac gtgcgcgaca acctgtatga tcggatttcc acaagaccct
     8521 tcctgaacaa tatcgagaag cggtggatcg cctttcagat tctgactgct gtggaccagg
     8581 cccataaatc tggagtgaga cacggcgata tcaagactga gaacgtgatg gtgaccagct
     8641 ggaattgggt gctgctgaca gacttcgctt cctttaagcc aacttacctg cccgaagaca
     8701 accctgccga tttcaattat ttctttgaca ctagccggag aaggacctgc tacatcgctc
     8761 ctgagcggtt cgtggatggc gggatgtttg ccaccgagct ggaatacatg agagacccat
     8821 caacacccct ggtggatctg aacagcaatc agaggaggag aggagagctg aagagggcta
     8881 tggacatctt ttctgctggg tgtgtgattg ccgagctgtt caccgaagga gtgcctctgt
     8941 ttgatctgag tcagctgctg gcctatagga acggacactt ctttccagag caggtgctga
     9001 ataagatcga ggatcatagc attcgcgaac tggtgacaca gatgatccac agggaaccag
     9061 acaaacgcc
//
