LOCUS       sequence-271157-        7460 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  3rd gen lentiviral Gateway destination vector, expression, CMV/TO 
            promoter, Zeo.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7460)
  AUTHORS   Campeau E, Ruhl VE, Rodier F, Smith CL, Rahmberg BL, Fuss JO, 
            Campisi J, Yaswen P, Cooper PK, Kaufman PD.
  TITLE     A versatile viral system for expression and depletion of proteins in
            mammalian cells.
  JOURNAL   PLoS One. 2009 Aug 6;4(8):e6529.
  PUBMED    19657394
REFERENCE   2  (bases 1 to 7460)
  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: "PLoS One.";
            date: "2009-08-6"; volume: "4(8)"; pages: "e6529"
COMMENT     Sequence Label: pLenti CMV/TO Zeo DEST (644-1)
FEATURES             Location/Qualifiers
     source          1..7460
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    51..167
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     enhancer        190..493
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        494..697
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     648..668
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    699..717
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    720..738
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    779..903
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(928..933,934..951,952..958)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             1012..1671
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1078..1097)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             2013..2318
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     2197..2216
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(2359..2483)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     misc_feature    2514..3102
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2567..2587)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2985..2996)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3153..3173)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3158..3487
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3338..3473
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        3499..3546
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3565..3939
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     LTR             4032..4265
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    4337..4458
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4374..4393)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4428..4447
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      4653..5241
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5142..5161
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5395..5412
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5529..5550
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5565..5570,5571..5588,5589..5595)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5603..5619
                     /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     5608..5630
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5627..5643
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5627..5643
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     5662..5682
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        5664..5682
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        5710..5936
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             5937..6117
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    6164..6289
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    6782..7015
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             7200..7244
                     /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"
ORIGIN
        1 agagacagat ccattcgatt agtgaacgga tctcgacggt atcggttaac ttttaaaaga
       61 aaagggggga ttggggggta cagtgcaggg gaaagaatag tagacataat agcaacagac
      121 atacaaacta aagaattaca aaaacaaatt acaaaattca aaattttatc gataagcttg
      181 ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc
      241 ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca
      301 ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta
      361 tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta
      421 tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat
      481 cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga
      541 ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca
      601 aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg
      661 taggcgtgta cggtgggagg tctatataag cagagctctc cctatcagtg atagagatct
      721 ccctatcagt gatagagatc gtcgactagt ccagtgtggt ggaattctgc agatatcaac
      781 aagtttgtac aaaaaagctg aacgagaaac gtaaaatgat ataaatatca atatattaaa
      841 ttagattttg cataaaaaac agactacata atactgtaaa acacaacata tccagtcact
      901 atggcggccg cattaggcac cccaggcttt acactttatg cttccggctc gtataatgtg
      961 tggattttga gttaggatcc gtcgagattt tcaggagcta aggaagctaa aatggagaaa
     1021 aaaatcactg gatataccac cgttgatata tcccaatggc atcgtaaaga acattttgag
     1081 gcatttcagt cagttgctca atgtacctat aaccagaccg ttcagctgga tattacggcc
     1141 tttttaaaga ccgtaaagaa aaataagcac aagttttatc cggcctttat tcacattctt
     1201 gcccgcctga tgaatgctca tccggaattc cgtatggcaa tgaaagacgg tgagctggtg
     1261 atatgggata gtgttcaccc ttgttacacc gttttccatg agcaaactga aacgttttca
     1321 tcgctctgga gtgaatacca cgacgatttc cggcagtttc tacacatata ttcgcaagat
     1381 gtggcgtgtt acggtgaaaa cctggcctat ttccctaaag ggtttattga gaatatgttt
     1441 ttcgtctcag ccaatccctg ggtgagtttc accagttttg atttaaacgt ggccaatatg
     1501 gacaacttct tcgcccccgt tttcaccatg ggcaaatatt atacgcaagg cgacaaggtg
     1561 ctgatgccgc tggcgattca ggttcatcat gccgtttgtg atggcttcca tgtcggcaga
     1621 atgcttaatg aattacaaca gtactgcgat gagtggcagg gcggggcgta aagatctgga
     1681 tccggcttac taaaagccag ataacagtat gcgtatttgc gcgctgattt ttgcggtata
     1741 agaatatata ctgatatgta tacccgaagt atgtcaaaaa gaggtatgct atgaagcagc
     1801 gtattacagt gacagttgac agcgacagct atcagttgct caaggcatat atgatgtcaa
     1861 tatctccggt ctggtaagca caaccatgca gaatgaagcc cgtcgtctgc gtgccgaacg
     1921 ctggaaagcg gaaaatcagg aagggatggc tgaggtcgcc cggtttattg aaatgaacgg
     1981 ctcttttgct gacgagaaca ggggctggtg aaatgcagtt taaggtttac acctataaaa
     2041 gagagagccg ttatcgtctg tttgtggatg tacagagtga tattattgac acgcccgggc
     2101 gacggatggt gatccccctg gccagtgcac gtctgctgtc agataaagtc tcccgtgaac
     2161 tttacccggt ggtgcatatc ggggatgaaa gctggcgcat gatgaccacc gatatggcca
     2221 gtgtgccggt ctccgttatc ggggaagaag tggctgatct cagccaccgc gaaaatgaca
     2281 tcaaaaacgc cattaacctg atgttctggg gaatataaat gtcaggctcc cttatacaca
     2341 gccagtctgc aggtcgacca tagtgactgg atatgttgtg ttttacagta ttatgtagtc
     2401 tgttttttat gcaaaatcta atttaatata ttgatattta tatcatttta cgtttctcgt
     2461 tcagctttct tgtacaaagt ggttgatatc cagcacagtg gcggccgctc gacaatcaac
     2521 ctctggatta caaaatttgt gaaagattga ctggtattct taactatgtt gctcctttta
     2581 cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt
     2641 tcattttctc ctccttgtat aaatcctggt tgctgtctct ttatgaggag ttgtggcccg
     2701 ttgtcaggca acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc actggttggg
     2761 gcattgccac cacctgtcag ctcctttccg ggactttcgc tttccccctc cctattgcca
     2821 cggcggaact catcgccgcc tgccttgccc gctgctggac aggggctcgg ctgttgggca
     2881 ctgacaattc cgtggtgttg tcggggaagc tgacgtcctt tccatggctg ctcgcctgtg
     2941 ttgccacctg gattctgcgc gggacgtcct tctgctacgt cccttcggcc ctcaatccag
     3001 cggaccttcc ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc
     3061 gccctcagac gagtcggatc tccctttggg ccgcctcccc gcctggaatt ctgcagatat
     3121 ccggttagta atgagtttgg aattaattct gtggaatgtg tgtcagttag ggtgtggaaa
     3181 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     3241 caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     3301 ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag
     3361 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc
     3421 cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt
     3481 ttgcaaaaag ctccccctgt tgacaattaa tcatcggcat agtatatcgg catagtataa
     3541 tacgacaagg tgaggaacta aaccatggcc aagttgacca gtgccgttcc ggtgctcacc
     3601 gcgcgcgacg tcgccggagc ggtcgagttc tggaccgacc ggctcgggtt ctcccgggac
     3661 ttcgtggagg acgacttcgc cggtgtggtc cgggacgacg tgaccctgtt catcagcgcg
     3721 gtccaggacc aggtggtgcc ggacaacacc ctggcctggg tgtgggtgcg cggcctggac
     3781 gagctgtacg ccgagtggtc ggaggtcgtg tccacgaact tccgggacgc ctccgggccg
     3841 gccatgaccg agatcggcga gcagccgtgg gggcgggagt tcgccctgcg cgacccggcc
     3901 ggcaactgcg tgcacttcgt ggccgaggag caggactgac acgtgctacg agatttaaat
     3961 ggtaccttta agaccaatga cttacaaggc agctgtagat cttagccact ttttaaaaga
     4021 aaagggggga ctggaagggc tagctcactc ccaacgaaga caagatctgc tttttgcttg
     4081 tactgggtct ctctggttag accagatctg agcctgggag ctctctggct aactagggaa
     4141 cccactgctt aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct
     4201 gttgtgtgac tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc
     4261 tagcagtagt agttcatgtc atcttattat tcagtattta taacttgcaa agaaatgaat
     4321 atcagagagt gagaggaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     4381 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     4441 actcatcaat gtatcttatc atgtctggct ctagctatcc cgcccctaac tccgcccatc
     4501 ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt
     4561 atttatgcag aggccgaggc cgccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     4621 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     4681 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     4741 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt
     4801 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     4861 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     4921 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     4981 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     5041 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     5101 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     5161 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     5221 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     5281 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
     5341 attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag
     5401 tcagtgagcg aggaagcgga agagcgccca atacgcaaac cgcctctccc cgcgcgttgg
     5461 ccgattcatt aatgcagctg gcacgacagg tttcccgact ggaaagcggg cagtgagcgc
     5521 aacgcaatta atgtgagtta gctcactcat taggcacccc aggctttaca ctttatgctt
     5581 ccggctcgta tgttgtgtgg aattgtgagc ggataacaat ttcacacagg aaacagctat
     5641 gaccatgatt acgccaagcg cgcaattaac cctcactaaa gggaacaaaa gctggagctg
     5701 caagcttaat gtagtcttat gcaatactct tgtagtcttg caacatggta acgatgagtt
     5761 agcaacatgc cttacaagga gagaaaaagc accgtgcatg ccgattggtg gaagtaaggt
     5821 ggtacgatcg tgccttatta ggaaggcaac agacgggtct gacatggatt ggacgaacca
     5881 ctgaattgcc gcattgcaga gatattgtat ttaagtgcct agctcgatac ataaacgggt
     5941 ctctctggtt agaccagatc tgagcctggg agctctctgg ctaactaggg aacccactgc
     6001 ttaagcctca ataaagcttg ccttgagtgc ttcaagtagt gtgtgcccgt ctgttgtgtg
     6061 actctggtaa ctagagatcc ctcagaccct tttagtcagt gtggaaaatc tctagcagtg
     6121 gcgcccgaac agggacttga aagcgaaagg gaaaccagag gagctctctc gacgcaggac
     6181 tcggcttgct gaagcgcgca cggcaagagg cgaggggcgg cgactggtga gtacgccaaa
     6241 aattttgact agcggaggct agaaggagag agatgggtgc gagagcgtca gtattaagcg
     6301 ggggagaatt agatcgcgat gggaaaaaat tcggttaagg ccagggggaa agaaaaaata
     6361 taaattaaaa catatagtat gggcaagcag ggagctagaa cgattcgcag ttaatcctgg
     6421 cctgttagaa acatcagaag gctgtagaca aatactggga cagctacaac catcccttca
     6481 gacaggatca gaagaactta gatcattata taatacagta gcaaccctct attgtgtgca
     6541 tcaaaggata gagataaaag acaccaagga agctttagac aagatagagg aagagcaaaa
     6601 caaaagtaag accaccgcac agcaagcggc cgctgatctt cagacctgga ggaggagata
     6661 tgagggacaa ttggagaagt gaattatata aatataaagt agtaaaaatt gaaccattag
     6721 gagtagcacc caccaaggca aagagaagag tggtgcagag agaaaaaaga gcagtgggaa
     6781 taggagcttt gttccttggg ttcttgggag cagcaggaag cactatgggc gcagcgtcaa
     6841 tgacgctgac ggtacaggcc agacaattat tgtctggtat agtgcagcag cagaacaatt
     6901 tgctgagggc tattgaggcg caacagcatc tgttgcaact cacagtctgg ggcatcaagc
     6961 agctccaggc aagaatcctg gctgtggaaa gatacctaaa ggatcaacag ctcctgggga
     7021 tttggggttg ctctggaaaa ctcatttgca ccactgctgt gccttggaat gctagttgga
     7081 gtaataaatc tctggaacag atttggaatc acacgacctg gatggagtgg gacagagaaa
     7141 ttaacaatta cacaagctta atacactcct taattgaaga atcgcaaaac cagcaagaaa
     7201 agaatgaaca agaattattg gaattagata aatgggcaag tttgtggaat tggtttaaca
     7261 taacaaattg gctgtggtat ataaaattat tcataatgat agtaggaggc ttggtaggtt
     7321 taagaatagt ttttgctgta ctttctatag tgaatagagt taggcaggga tattcaccat
     7381 tatcgtttca gacccacctc ccaaccccga ggggacccga caggcccgaa ggaatagaag
     7441 aagaaggtgg agagagagac
//
