LOCUS       sequence-389912-        7672 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral transfer plasmid for doxycycline inducible expression of 
            a C-terminally 3C-SUMOEu-GFP-tagged protein in mammalian cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7672)
  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 7672)
  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: pTS106 pHAGE2 CMVtetO2 MCS-3C-SUMOEu-22xGS-GFP
FEATURES             Location/Qualifiers
     source          1..7672
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             12..645
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    692..817
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1314..1547
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1732..1776
                     /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    2069..2186
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     enhancer        2230..2609
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2610..2813
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2764..2784
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    2815..2833
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2836..2854
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     2864..2888
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      2963..2972
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3032..3055
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     misc_feature    4125..4713
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4178..4198)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4596..4607)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             4861..5041
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(5088..5106)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5174..5278
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5279..5347,5348..6139)
                     /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(5497..5516)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6310..6898
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7052..7069
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7309..7330
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7345..7350,7351..7368,7369..7375)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7383..7399
                     /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     7388..7410
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7407..7423
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7407..7423
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     7473..7492
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
ORIGIN
        1 aacttagtag ttggaagggc taattcactc ccaaagaaga caagatatcc ttgatctgtg
       61 gatctaccac acacaaggct acttccctga ttagcagaac tacacaccag ggccaggggt
      121 cagatatcca ctgacctttg gatggtgcta caagctagta ccagttgagc cagataaggt
      181 agaagaggcc aataaaggag agaacaccag cttgttacac cctgtgagcc tgcatgggat
      241 ggatgacccg gagagagaag tgttagagtg gaggtttgac agccgcctag catttcatca
      301 cgtggcccga gagctgcatc cggagtactt caagaactgc tgatatcgag cttgctacaa
      361 gggactttcc gctggggact ttccagggag gcgtggcctg ggcgggactg gggagtggcg
      421 agccctcaga tcctgcatat aagcagctgc tttttgcctg tactgggtct ctctggttag
      481 accagatctg agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat
      541 aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact
      601 agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc gcccgaacag
      661 ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc ggcttgctga
      721 agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa ttttgactag
      781 cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg ggagaattag
      841 atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata aattaaaaca
      901 tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc tgttagaaac
      961 atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga caggatcaga
     1021 agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc aaaggataga
     1081 gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca aaagtaagac
     1141 caccgcacag caagcggccg gccgctgatc ttcagacctg gaggaggaga tatgagggac
     1201 aattggagaa gtgaattata taaatataaa gtagtaaaaa ttgaaccatt aggagtagca
     1261 cccaccaagg caaagagaag agtggtgcag agagaaaaaa gagcagtggg aataggagct
     1321 ttgttccttg ggttcttggg agcagcagga agcactatgg gcgcagcgtc aatgacgctg
     1381 acggtacagg ccagacaatt attgtctggt atagtgcagc agcagaacaa tttgctgagg
     1441 gctattgagg cgcaacagca tctgttgcaa ctcacagtct ggggcatcaa gcagctccag
     1501 gcaagaatcc tggctgtgga aagataccta aaggatcaac agctcctggg gatttggggt
     1561 tgctctggaa aactcatttg caccactgct gtgccttgga atgctagttg gagtaataaa
     1621 tctctggaac agatttggaa tcacacgacc tggatggagt gggacagaga aattaacaat
     1681 tacacaagct taatacactc cttaattgaa gaatcgcaaa accagcaaga aaagaatgaa
     1741 caagaattat tggaattaga taaatgggca agtttgtgga attggtttaa cataacaaat
     1801 tggctgtggt atataaaatt attcataatg atagtaggag gcttggtagg tttaagaata
     1861 gtttttgctg tactttctat agtgaataga gttaggcagg gatattcacc attatcgttt
     1921 cagacccacc tcccaacccc gaggggaccc gacaggcccg aaggaataga agaagaaggt
     1981 ggagagagag acagagacag atccattcga ttagtgaacg gatctcgacg gtatcgccga
     2041 attcacaaat ggcagtattc atccacaatt ttaaaagaaa aggggggatt ggggggtaca
     2101 gtgcagggga aagaatagta gacataatag caacagacat acaaactaaa gaattacaaa
     2161 aacaaattac aaaaattcaa aattttcggg tttattacag ggacagcaga gatccagttt
     2221 ggacgcgttg acattgatta ttgactagtt attaatagta atcaattacg gggtcattag
     2281 ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct
     2341 gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc
     2401 caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg
     2461 cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat
     2521 ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca
     2581 tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc
     2641 gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga
     2701 gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat
     2761 tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctctcccta
     2821 tcagtgatag agatctccct atcagtgata gagatcgtcg acgagctcgt ttagtgaacc
     2881 gtcagatcgc ctggagacgc catccacgct gttttgacct ccatagaaga caccgggacc
     2941 gatccagcct ccggctagag cggccgccat gggatccggt tctgctagcg gttcaggttc
     3001 taccggtgga tcatctagaa ctagcggtag cttagaagtt ttgtttcaag gtccaggtag
     3061 cggatcttct gctgcgggtg gcgaagaaga taagaaaccg gcaggtggcg aaggtggcgg
     3121 tgcccatatc aacctgaaag tgaaaggtca agacggcaac gaagtctttt tccgcatcaa
     3181 acgttctacc cagctgaaaa agctgatgaa cgcatactgt gaccgtcagt ctgtagacat
     3241 gaaggcaatt gctttcctct ttaagggtcg tcgcctacgt gcggaaagga ccccggatga
     3301 actggaaatg gaagatggcg acgaaatcga cgcaatgctg catcagactg gtggcggatc
     3361 tggaagtgga tctggatcag gaagtggttc tggatcaggt agtggttctg gtagtggatc
     3421 aatggtgtct aaaggtgaag aactttttac gggtgtcgtc cccatcctgg ttgaattgga
     3481 cggagacgtg aatggccata aattttcagt gagcggtgaa ggagagggag acgccactta
     3541 cggtaaactt accttgaaat ttatctgcac taccgggaaa ctgcccgtac cctggccgac
     3601 gctcgtaaca acgcttacct acggcgtgca atgttttagc agatacccag accatatgaa
     3661 gcagcacgat tttttcaaga gtgcaatgcc tgaggggtac gtacaggaaa ggacgatctt
     3721 tttcaaagac gatggtaatt ataaaacgcg ggctgaggtt aaatttgaag gagacacgct
     3781 cgtgaatcga atagagctga agggaatcga cttcaaggaa gacggtaaca tcctgggcca
     3841 taaactcgaa tataattaca attcacacaa cgtctatatc atggcagata aacagaagaa
     3901 tggcatcaaa gtcaatttta agatccggca taatatcgag gacggatcag tacagcttgc
     3961 cgaccattac caacagaata caccgatagg agacggtccc gtcctcttgc cagataatca
     4021 ctatttgagc acacaatccg ctctttctaa agatccgaat gaaaagcgcg atcatatggt
     4081 cttgaaagag ttcgtcactg ccgccggcat aacctaactc gagtaatcaa cctctggatt
     4141 acaaaatttg tgaaagattg actggtattc ttaactatgt tgctcctttt acgctatgtg
     4201 gatacgctgc tttaatgcct ttgtatcatg ctattgcttc ccgtatggct ttcattttct
     4261 cctccttgta taaatcctgg ttgctgtctc tttatgagga gttgtggccc gttgtcaggc
     4321 aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc cactggttgg ggcattgcca
     4381 ccacctgtca gctcctttcc gggactttcg ctttccccct ccctattgcc acggcggaac
     4441 tcatcgccgc ctgccttgcc cgctgctgga caggggctcg gctgttgggc actgacaatt
     4501 ccgtggtgtt gttggggaaa tcatcgtcct ttccttggct gctcgcctgt gttgccacct
     4561 ggattctgcg cgggacgtcc ttctgctacg tcccttcggc cctcaatcca gcggaccttc
     4621 cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga
     4681 cgagtcggat ctccctttgg gccgcctccc cgcctggtac ctttaagacc aatgacttac
     4741 aaggcagctg tagatcttag ccacttttta aaagaaaagg ggggactgga agggctaatt
     4801 cactcccaac gaagacaaga tcacctgcag gacaggcgcg ccctgctttt tgcttgtact
     4861 gggtctctct ggttagacca gatctgagcc tgggagctct ctggctaact agggaaccca
     4921 ctgcttaagc ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg
     4981 tgtgactctg gtaactagag atccctcaga cccttttagt cagtgtggaa aatctctagc
     5041 acccgggcga ttaaggaaag ggctagatca ttcttgaaga cgaaagggcc tcgtgatacg
     5101 cctattttta taggttaatg tcatgataat aatggtttct tagacgtcag gtggcacttt
     5161 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta
     5221 tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat
     5281 gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt
     5341 ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg
     5401 agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga
     5461 agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg
     5521 tgttgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt
     5581 tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg
     5641 cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg
     5701 aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga
     5761 tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc
     5821 tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc
     5881 ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc
     5941 ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg
     6001 cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac
     6061 gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc
     6121 actgattaag cattggtaac tgtcagacca agtttactca tatatacttt agattgattt
     6181 aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac
     6241 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa
     6301 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc
     6361 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt
     6421 aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg
     6481 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc
     6541 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt
     6601 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga
     6661 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct
     6721 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
     6781 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
     6841 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
     6901 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt
     6961 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
     7021 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
     7081 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagcaagct
     7141 catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta
     7201 ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctccccgtg
     7261 gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atgtgagtta
     7321 gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta tgttgtgtgg
     7381 aattgtgagc ggataacaat ttcacacagg aaacagctat gacatgatta cgaatttcac
     7441 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
     7501 ttatcatgtc tggatcaact ggataactca agctaaccaa aatcatccca aacttcccac
     7561 cccataccct attaccactg ccaattacct gtggtttcat ttactctaaa cctgtgattc
     7621 ctctgaatta ttttcatttt aaagaaattg tatttgttaa atatgtacta ca
//
