LOCUS       sequence-389913-        7669 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral transfer plasmid for doxycycline inducible expression of 
            a C-terminally TEV-SUMOEu-GFP-tagged protein in mammalian cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7669)
  AUTHORS   Stevens TA, Tomaleri GP, Hazu M, Wei S, Nguyen VN, DeKalb C, 
            Voorhees RM, Pleiner T
  TITLE     A nanobody-based strategy for rapid and scalable purification of 
            human protein complexes.
  JOURNAL   Nat Protoc. 2024 Jan;19(1):127-158. doi: 10.1038/s41596-023-00904-w.
            Epub 2023 Nov 16.
  PUBMED    37974029
REFERENCE   2  (bases 1 to 7669)
  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"; doi: 
            "10.1038/s41596-023-00904-w"; journalName: "Nat Protoc"; date: 
            "2024-01"; volume: "19"; issue: "1"; pages: "127-158"
COMMENT     Sequence Label: pTS107 pHAGE2 CMVtetO2 MCS-TEV-SUMOEu-22xGS-GFP
FEATURES             Location/Qualifiers
     source          1..7669
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             6..639
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    686..811
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1308..1541
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1726..1770
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    2063..2180
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     enhancer        2224..2603
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2604..2807
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2758..2778
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    2809..2827
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2830..2848
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     2858..2882
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      2957..2966
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3026..3046
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     misc_feature    4116..4704
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4169..4189)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4587..4598)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             4852..5032
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(5079..5097)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5165..5269
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5270..5338,5339..6130)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5488..5507)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6301..6889
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7043..7060
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7300..7321
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7336..7341,7342..7359,7360..7366)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7374..7390
                     /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     7379..7401
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7398..7414
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7398..7414
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     7464..7483
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
ORIGIN
        1 gtagttggaa gggctaattc actcccaaag aagacaagat atccttgatc tgtggatcta
       61 ccacacacaa ggctacttcc ctgattagca gaactacaca ccagggccag gggtcagata
      121 tccactgacc tttggatggt gctacaagct agtaccagtt gagccagata aggtagaaga
      181 ggccaataaa ggagagaaca ccagcttgtt acaccctgtg agcctgcatg ggatggatga
      241 cccggagaga gaagtgttag agtggaggtt tgacagccgc ctagcatttc atcacgtggc
      301 ccgagagctg catccggagt acttcaagaa ctgctgatat cgagcttgct acaagggact
      361 ttccgctggg gactttccag ggaggcgtgg cctgggcggg actggggagt ggcgagccct
      421 cagatcctgc atataagcag ctgctttttg cctgtactgg gtctctctgg ttagaccaga
      481 tctgagcctg ggagctctct ggctaactag ggaacccact gcttaagcct caataaagct
      541 tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt aactagagat
      601 ccctcagacc cttttagtca gtgtggaaaa tctctagcag tggcgcccga acagggactt
      661 gaaagcgaaa gggaaaccag aggagctctc tcgacgcagg actcggcttg ctgaagcgcg
      721 cacggcaaga ggcgaggggc ggcgactggt gagtacgcca aaaattttga ctagcggagg
      781 ctagaaggag agagatgggt gcgagagcgt cagtattaag cgggggagaa ttagatcgcg
      841 atgggaaaaa attcggttaa ggccaggggg aaagaaaaaa tataaattaa aacatatagt
      901 atgggcaagc agggagctag aacgattcgc agttaatcct ggcctgttag aaacatcaga
      961 aggctgtaga caaatactgg gacagctaca accatccctt cagacaggat cagaagaact
     1021 tagatcatta tataatacag tagcaaccct ctattgtgtg catcaaagga tagagataaa
     1081 agacaccaag gaagctttag acaagataga ggaagagcaa aacaaaagta agaccaccgc
     1141 acagcaagcg gccggccgct gatcttcaga cctggaggag gagatatgag ggacaattgg
     1201 agaagtgaat tatataaata taaagtagta aaaattgaac cattaggagt agcacccacc
     1261 aaggcaaaga gaagagtggt gcagagagaa aaaagagcag tgggaatagg agctttgttc
     1321 cttgggttct tgggagcagc aggaagcact atgggcgcag cgtcaatgac gctgacggta
     1381 caggccagac aattattgtc tggtatagtg cagcagcaga acaatttgct gagggctatt
     1441 gaggcgcaac agcatctgtt gcaactcaca gtctggggca tcaagcagct ccaggcaaga
     1501 atcctggctg tggaaagata cctaaaggat caacagctcc tggggatttg gggttgctct
     1561 ggaaaactca tttgcaccac tgctgtgcct tggaatgcta gttggagtaa taaatctctg
     1621 gaacagattt ggaatcacac gacctggatg gagtgggaca gagaaattaa caattacaca
     1681 agcttaatac actccttaat tgaagaatcg caaaaccagc aagaaaagaa tgaacaagaa
     1741 ttattggaat tagataaatg ggcaagtttg tggaattggt ttaacataac aaattggctg
     1801 tggtatataa aattattcat aatgatagta ggaggcttgg taggtttaag aatagttttt
     1861 gctgtacttt ctatagtgaa tagagttagg cagggatatt caccattatc gtttcagacc
     1921 cacctcccaa ccccgagggg acccgacagg cccgaaggaa tagaagaaga aggtggagag
     1981 agagacagag acagatccat tcgattagtg aacggatctc gacggtatcg ccgaattcac
     2041 aaatggcagt attcatccac aattttaaaa gaaaaggggg gattgggggg tacagtgcag
     2101 gggaaagaat agtagacata atagcaacag acatacaaac taaagaatta caaaaacaaa
     2161 ttacaaaaat tcaaaatttt cgggtttatt acagggacag cagagatcca gtttggacgc
     2221 gttgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
     2281 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
     2341 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
     2401 ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac
     2461 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
     2521 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
     2581 tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat
     2641 agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt
     2701 tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc
     2761 aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctctc cctatcagtg
     2821 atagagatct ccctatcagt gatagagatc gtcgacgagc tcgtttagtg aaccgtcaga
     2881 tcgcctggag acgccatcca cgctgttttg acctccatag aagacaccgg gaccgatcca
     2941 gcctccggct agagcggccg ccatgggatc cggttctgct agcggttcag gttctaccgg
     3001 tggatcatct agaactagcg gtagcgagaa tttgtatttt cagggtggta gcggatcttc
     3061 tgctgcgggt ggcgaagaag ataagaaacc ggcaggtggc gaaggtggcg gtgcccatat
     3121 caacctgaaa gtgaaaggtc aagacggcaa cgaagtcttt ttccgcatca aacgttctac
     3181 ccagctgaaa aagctgatga acgcatactg tgaccgtcag tctgtagaca tgaaggcaat
     3241 tgctttcctc tttaagggtc gtcgcctacg tgcggaaagg accccggatg aactggaaat
     3301 ggaagatggc gacgaaatcg acgcaatgct gcatcagact ggtggcggat ctggaagtgg
     3361 atctggatca ggaagtggtt ctggatcagg tagtggttct ggtagtggat caatggtgtc
     3421 taaaggtgaa gaacttttta cgggtgtcgt ccccatcctg gttgaattgg acggagacgt
     3481 gaatggccat aaattttcag tgagcggtga aggagaggga gacgccactt acggtaaact
     3541 taccttgaaa tttatctgca ctaccgggaa actgcccgta ccctggccga cgctcgtaac
     3601 aacgcttacc tacggcgtgc aatgttttag cagataccca gaccatatga agcagcacga
     3661 ttttttcaag agtgcaatgc ctgaggggta cgtacaggaa aggacgatct ttttcaaaga
     3721 cgatggtaat tataaaacgc gggctgaggt taaatttgaa ggagacacgc tcgtgaatcg
     3781 aatagagctg aagggaatcg acttcaagga agacggtaac atcctgggcc ataaactcga
     3841 atataattac aattcacaca acgtctatat catggcagat aaacagaaga atggcatcaa
     3901 agtcaatttt aagatccggc ataatatcga ggacggatca gtacagcttg ccgaccatta
     3961 ccaacagaat acaccgatag gagacggtcc cgtcctcttg ccagataatc actatttgag
     4021 cacacaatcc gctctttcta aagatccgaa tgaaaagcgc gatcatatgg tcttgaaaga
     4081 gttcgtcact gccgccggca taacctaact cgagtaatca acctctggat tacaaaattt
     4141 gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt ggatacgctg
     4201 ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc tcctccttgt
     4261 ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg caacgtggcg
     4321 tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc accacctgtc
     4381 agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa ctcatcgccg
     4441 cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat tccgtggtgt
     4501 tgttggggaa atcatcgtcc tttccttggc tgctcgcctg tgttgccacc tggattctgc
     4561 gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg
     4621 gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag acgagtcgga
     4681 tctccctttg ggccgcctcc ccgcctggta cctttaagac caatgactta caaggcagct
     4741 gtagatctta gccacttttt aaaagaaaag gggggactgg aagggctaat tcactcccaa
     4801 cgaagacaag atcacctgca ggacaggcgc gccctgcttt ttgcttgtac tgggtctctc
     4861 tggttagacc agatctgagc ctgggagctc tctggctaac tagggaaccc actgcttaag
     4921 cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct
     4981 ggtaactaga gatccctcag acccttttag tcagtgtgga aaatctctag cacccgggcg
     5041 attaaggaaa gggctagatc attcttgaag acgaaagggc ctcgtgatac gcctattttt
     5101 ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa
     5161 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     5221 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     5281 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     5341 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     5401 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     5461 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtgttgacgc
     5521 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     5581 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     5641 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     5701 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     5761 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat
     5821 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     5881 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     5941 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     6001 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     6061 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     6121 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     6181 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     6241 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     6301 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     6361 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     6421 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
     6481 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     6541 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     6601 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     6661 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     6721 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     6781 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     6841 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     6901 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     6961 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
     7021 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
     7081 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagcaagc tcatggctga
     7141 ctaatttttt ttatttatgc agaggccgag gccgcctcgg cctctgagct attccagaag
     7201 tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctccccgt ggcacgacag
     7261 gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt agctcactca
     7321 ttaggcaccc caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag
     7381 cggataacaa tttcacacag gaaacagcta tgacatgatt acgaatttca caaataaagc
     7441 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt
     7501 ctggatcaac tggataactc aagctaacca aaatcatccc aaacttccca ccccataccc
     7561 tattaccact gccaattacc tgtggtttca tttactctaa acctgtgatt cctctgaatt
     7621 attttcattt taaagaaatt gtatttgtta aatatgtact acaaactta
//
