LOCUS       Exported                7997 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  3rd gen lentiviral empty control vector, CMV/TO promoter, Puro.
ACCESSION   .
VERSION     .
KEYWORDS    pLenti CMV/TO Puro empty (w175-1)
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7997)
  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 7997)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7997
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        21..324
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        325..528
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     479..499
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    530..548
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    551..569
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    610..634
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(703..727)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    758..1346
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(811..831)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(1229..1240)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     promoter        1360..1859
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(1395..1418)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     1792..1811
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             1880..2479
                     /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(1880..1899)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     2376..2396
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             2617..2850
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    2922..3043
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2959..2978)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3013..3032
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      3062..3197
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3124..3143
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     complement(3217..3236)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3218..3236)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3246..3263)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3246..3262)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3255..3277)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3404..3859
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3491..3510)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3701..3722
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3885..3989
                     /gene="bla"
                     /label=AmpR promoter
     CDS             3990..4850
                     /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     complement(4208..4227)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5021..5609
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5510..5529
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5763..5780
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5897..5918
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        5933..5963
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5971..5987
                     /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     5976..5998
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5995..6011
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5995..6011
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     6030..6050
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        6032..6050
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        6078..6304
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             6305..6485
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    6532..6657
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    7150..7383
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             7568..7612
                     /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    7879..7995
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
ORIGIN
        1 cgataagctt gggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg
       61 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata
      121 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta
      181 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc
      241 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac
      301 gtattagtca tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga
      361 tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg
      421 ttttggcacc aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg
      481 caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt
      541 gatagagatc tccctatcag tgatagagat cgtcgactag tccagtgtgg tggaattctg
      601 cagatatcaa caagtttgta caaaaaagca ggctccacca tgggaaccaa ttcagtcgac
      661 tggatccggt accgaattcg cggccgcact cgagatatct agacccagct ttcttgtaca
      721 aagtggttga tatccagcac agtggcggcc gctcgacaat caacctctgg attacaaaat
      781 ttgtgaaaga ttgactggta ttcttaacta tgttgctcct tttacgctat gtggatacgc
      841 tgctttaatg cctttgtatc atgctattgc ttcccgtatg gctttcattt tctcctcctt
      901 gtataaatcc tggttgctgt ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg
      961 cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt tggggcattg ccaccacctg
     1021 tcagctcctt tccgggactt tcgctttccc cctccctatt gccacggcgg aactcatcgc
     1081 cgcctgcctt gcccgctgct ggacaggggc tcggctgttg ggcactgaca attccgtggt
     1141 gttgtcgggg aagctgacgt cctttccatg gctgctcgcc tgtgttgcca cctggattct
     1201 gcgcgggacg tccttctgct acgtcccttc ggccctcaat ccagcggacc ttccttcccg
     1261 cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg
     1321 gatctccctt tgggccgcct ccccgcctgg aattctaccg ggtaggggag gcgcttttcc
     1381 caaggcagtc tggagcatgc gctttagcag ccccgctggg cacttggcgc tacacaagtg
     1441 gcctctggcc tcgcacacat tccacatcca ccggtaggcg ccaaccggct ccgttctttg
     1501 gtggcccctt cgcgccacct tctactcctc ccctagtcag gaagttcccc cccgccccgc
     1561 agctcgcgtc gtgcaggacg tgacaaatgg aagtagcacg tctcactagt ctcgtgcaga
     1621 tggacagcac cgctgagcaa tggaagcggg taggcctttg gggcagcggc caatagcagc
     1681 tttgctcctt cgctttctgg gctcagaggc tgggaagggg tgggtccggg ggcgggctca
     1741 ggggcgggct caggggcggg gcgggcgccc gaaggtcctc cggaggcccg gcattctgca
     1801 cgcttcaaaa gcgcacgtct gccgcgctgt tctcctcttc ctcatctccg ggcctttcga
     1861 cctgcagccc aagcttacca tgaccgagta caagcccacg gtgcgcctcg ccacccgcga
     1921 cgacgtcccc agggccgtac gcaccctcgc cgccgcgttc gccgactacc ccgccacgcg
     1981 ccacaccgtc gatccggacc gccacatcga gcgggtcacc gagctgcaag aactcttcct
     2041 cacgcgcgtc gggctcgaca tcggcaaggt gtgggtcgcg gacgacggcg ccgcggtggc
     2101 ggtctggacc acgccggaga gcgtcgaagc gggggcggtg ttcgccgaga tcggcccgcg
     2161 catggccgag ttgagcggtt cccggctggc cgcgcagcaa cagatggaag gcctcctggc
     2221 gccgcaccgg cccaaggagc ccgcgtggtt cctggccacc gtcggcgtct cgcccgacca
     2281 ccagggcaag ggtctgggca gcgccgtcgt gctccccgga gtggaggcgg ccgagcgcgc
     2341 cggggtgccc gccttcctgg agacctccgc gccccgcaac ctccccttct acgagcggct
     2401 cggcttcacc gtcaccgccg acgtcgaggt gcccgaagga ccgcgcacct ggtgcatgac
     2461 ccgcaagccc ggtgcctgac gcccgcccca cgacccgcag cgcccgaccg aaaggagcgc
     2521 acgaccccat gcatctcgag ggcccggtac ctttaagacc aatgacttac aaggcagctg
     2581 tagatcttag ccacttttta aaagaaaagg ggggactgga agggctagct cactcccaac
     2641 gaagacaaga tctgcttttt gcttgtactg ggtctctctg gttagaccag atctgagcct
     2701 gggagctctc tggctaacta gggaacccac tgcttaagcc tcaataaagc ttgccttgag
     2761 tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg taactagaga tccctcagac
     2821 ccttttagtc agtgtggaaa atctctagca gtagtagttc atgtcatctt attattcagt
     2881 atttataact tgcaaagaaa tgaatatcag agagtgagag gaacttgttt attgcagctt
     2941 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     3001 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tggctctagc
     3061 tatcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt
     3121 ttttatttat gcagaggccg aggccgcctc ggcctctgag ctattccaga agtagtgagg
     3181 aggctttttt ggaggcctag ggacgtaccc aattcgccct atagtgagtc gtattacgcg
     3241 cgctcactgg ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt tacccaactt
     3301 aatcgccttg cagcacatcc ccctttcgcc agctggcgta atagcgaaga ggcccgcacc
     3361 gatcgccctt cccaacagtt gcgcagcctg aatggcgaat gggacgcgcc ctgtagcggc
     3421 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     3481 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     3541 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     3601 gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     3661 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     3721 ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt
     3781 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
     3841 atattaacgc ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     3901 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     3961 tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta
     4021 ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     4081 taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca
     4141 gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta
     4201 aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc
     4261 gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc
     4321 ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca
     4381 ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc
     4441 acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
     4501 taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac
     4561 tattaactgg cgaactactt actctagctt cccggcaaca attaatagac tggatggagg
     4621 cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg
     4681 ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg
     4741 gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
     4801 gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc
     4861 aagtttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     4921 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     4981 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     5041 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     5101 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     5161 atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     5221 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     5281 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     5341 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     5401 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     5461 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     5521 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     5581 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     5641 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     5701 ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc
     5761 gcagcgagtc agtgagcgag gaagcggaag agcgcccaat acgcaaaccg cctctccccg
     5821 cgcgttggcc gattcattaa tgcagctggc acgacaggtt tcccgactgg aaagcgggca
     5881 gtgagcgcaa cgcaattaat gtgagttagc tcactcatta ggcaccccag gctttacact
     5941 ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa
     6001 acagctatga ccatgattac gccaagcgcg caattaaccc tcactaaagg gaacaaaagc
     6061 tggagctgca agcttaatgt agtcttatgc aatactcttg tagtcttgca acatggtaac
     6121 gatgagttag caacatgcct tacaaggaga gaaaaagcac cgtgcatgcc gattggtgga
     6181 agtaaggtgg tacgatcgtg ccttattagg aaggcaacag acgggtctga catggattgg
     6241 acgaaccact gaattgccgc attgcagaga tattgtattt aagtgcctag ctcgatacat
     6301 aaacgggtct ctctggttag accagatctg agcctgggag ctctctggct aactagggaa
     6361 cccactgctt aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct
     6421 gttgtgtgac tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc
     6481 tagcagtggc gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga
     6541 cgcaggactc ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt
     6601 acgccaaaaa ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt
     6661 attaagcggg ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag
     6721 aaaaaatata aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt
     6781 aatcctggcc tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca
     6841 tcccttcaga caggatcaga agaacttaga tcattatata atacagtagc aaccctctat
     6901 tgtgtgcatc aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa
     6961 gagcaaaaca aaagtaagac caccgcacag caagcggccg ctgatcttca gacctggagg
     7021 aggagatatg agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga
     7081 accattagga gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc
     7141 agtgggaata ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc
     7201 agcgtcaatg acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca
     7261 gaacaatttg ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg
     7321 catcaagcag ctccaggcaa gaatcctggc tgtggaaaga tacctaaagg atcaacagct
     7381 cctggggatt tggggttgct ctggaaaact catttgcacc actgctgtgc cttggaatgc
     7441 tagttggagt aataaatctc tggaacagat ttggaatcac acgacctgga tggagtggga
     7501 cagagaaatt aacaattaca caagcttaat acactcctta attgaagaat cgcaaaacca
     7561 gcaagaaaag aatgaacaag aattattgga attagataaa tgggcaagtt tgtggaattg
     7621 gtttaacata acaaattggc tgtggtatat aaaattattc ataatgatag taggaggctt
     7681 ggtaggttta agaatagttt ttgctgtact ttctatagtg aatagagtta ggcagggata
     7741 ttcaccatta tcgtttcaga cccacctccc aaccccgagg ggacccgaca ggcccgaagg
     7801 aatagaagaa gaaggtggag agagagacag agacagatcc attcgattag tgaacggatc
     7861 tcgacggtat cggttaactt ttaaaagaaa aggggggatt ggggggtaca gtgcagggga
     7921 aagaatagta gacataatag caacagacat acaaactaaa gaattacaaa aacaaattac
     7981 aaaattcaaa attttat
//
