LOCUS Exported 8410 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION 3rd gen lentiviral Gateway destination vector, expression, CMV/TO promoter. ACCESSION . VERSION . KEYWORDS pLenti CMV/TO DEST (775-1) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 8410) 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 8410) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..8410 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 27..131 /gene="bla" /label=AmpR promoter CDS 132..992 /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(350..369) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 1163..1751 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 1652..1671 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 1905..1922 /label=L4440 /note="L4440 vector, forward primer" protein_bind 2039..2060 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 2075..2105 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 2113..2129 /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 2118..2140 /label=M13/pUC Reverse /note="In lacZ gene" primer_bind 2137..2153 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 2137..2153 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind 2172..2192 /label=T3 /note="T3 promoter, forward primer" promoter 2174..2192 /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" LTR 2448..2628 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from HIV-1" misc_feature 2672..2797 /label=HIV-1 Psi /note="packaging signal of human immunodeficiency virus type 1" misc_feature 3290..3523 /label=RRE /note="The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm." CDS 3707..3751 /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 4012..4129 /label=cPPT/CTS /note="central polypurine tract and central termination sequence of HIV-1" enhancer 4152..4455 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 4456..4659 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 4610..4630 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" protein_bind 4661..4679 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 4682..4700 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 4741..4865 /gene="mutant version of attR" /label=attR1 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" promoter 4890..4920 /label=lac UV5 promoter /note="E. coli lac promoter with an ""up"" mutation" CDS 4974..5633 /codon_start=1 /gene="cat" /product="chloramphenicol acetyltransferase" /label=CmR /note="confers resistance to chloramphenicol" /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA" primer_bind complement(5040..5059) /label=CAT-R /note="Chloramphenicol resistance gene, reverse primer" CDS 5975..6280 /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 6159..6178 /label=ccdB-fwd /note="ccdB gene, forward primer" protein_bind complement(6321..6445) /gene="mutant version of attR" /label=attR2 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" misc_feature 6476..7064 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(6529..6549) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(6947..6958) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" LTR 7151..7384 /label=3' LTR (Delta-U3) /note="self-inactivating 3' long terminal repeat (LTR) from HIV-1" polyA_signal 7456..7577 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(7493..7512) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 7547..7566 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" rep_origin 7596..7731 /label=SV40 ori /note="SV40 origin of replication" primer_bind 7658..7677 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" primer_bind complement(7763..7782) /label=T7 /note="T7 promoter, forward primer" promoter complement(7764..7782) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(7792..7809) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(7792..7808) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(7801..7823) /label=M13/pUC Forward /note="In lacZ gene" rep_origin 7953..8408 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(8040..8059) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 8250..8271 /label=F1ori-F /note="F1 origin, forward primer" ORIGIN 1 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac 61 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa 121 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat 181 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc 241 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga 301 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg 361 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc 421 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag 481 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc 541 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg 601 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg 661 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac 721 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac 781 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg 841 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg 901 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg 961 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac 1021 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg 1081 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg 1141 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc 1201 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc 1261 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt 1321 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc 1381 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact 1441 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac 1501 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag 1561 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg 1621 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg 1681 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga 1741 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt 1801 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct 1861 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg 1921 aggaagcgga agagcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt 1981 aatgcagctg gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta 2041 atgtgagtta gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta 2101 tgttgtgtgg aattgtgagc ggataacaat ttcacacagg aaacagctat gaccatgatt 2161 acgccaagcg cgcaattaac cctcactaaa gggaacaaaa gctggagctg caagcttaat 2221 gtagtcttat gcaatactct tgtagtcttg caacatggta acgatgagtt agcaacatgc 2281 cttacaagga gagaaaaagc accgtgcatg ccgattggtg gaagtaaggt ggtacgatcg 2341 tgccttatta ggaaggcaac agacgggtct gacatggatt ggacgaacca ctgaattgcc 2401 gcattgcaga gatattgtat ttaagtgcct agctcgatac aataaacggg tctctctggt 2461 tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg cttaagcctc 2521 aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt gactctggta 2581 actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagt ggcgcccgaa 2641 cagggacctg aaagcgaaag ggaaaccaga gctctctcga cgcaggactc ggcttgctga 2701 agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa ttttgactag 2761 cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg ggagaattag 2821 atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata aattaaaaca 2881 tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc tgttagaaac 2941 atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga caggatcaga 3001 agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc aaaggataga 3061 gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca aaagtaagac 3121 caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg agggacaatt 3181 ggagaagtga attatataaa tataaagtag taaaaattga accattagga gtagcaccca 3241 ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata ggagctttgt 3301 tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcctcaatg acgctgacgg 3361 tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg ctgagggcta 3421 ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag ctccaggcaa 3481 gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt tggggttgct 3541 ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt aataaatctc 3601 tggaacagat tggaatcaca cgacctggat ggagtgggac agagaaatta acaattacac 3661 aagcttaata cactccttaa ttgaagaatc gcaaaaccag caagaaaaga atgaacaaga 3721 attattggaa ttagataaat gggcaagttt gtggaattgg tttaacataa caaattggct 3781 gtggtatata aaattattca taatgatagt aggaggcttg gtaggtttaa gaatagtttt 3841 tgctgtactt tctatagtga atagagttag gcagggatat tcaccattat cgtttcagac 3901 ccacctccca accccgaggg gacccgacag gcccgaagga atagaagaag aaggtggaga 3961 gagagacaga gacagatcca ttcgattagt gaacggatct cgacggttaa cttttaaaag 4021 aaaagggggg attggggggt acagtgcagg ggaaagaata gtagacataa tagcaacaga 4081 catacaaact aaagaattac aaaaacaaat tacaaaaatt caaaatttta tcgataagct 4141 tgggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac 4201 ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc 4261 cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg 4321 tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat 4381 tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc 4441 atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt 4501 gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac 4561 caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc 4621 ggtaggcgtg tacggtggga ggtctatata agcagagctc tccctatcag tgatagagat 4681 ctccctatca gtgatagaga tcgtcgacta gtccagtgtg gtggaattct gcagatatca 4741 acaagtttgt acaaaaaagc tgaacgagaa acgtaaaatg atataaatat caatatatta 4801 aattagattt tgcataaaaa acagactaca taatactgta aaacacaaca tatccagtca 4861 ctatggcggc cgcattaggc accccaggct ttacacttta tgcttccggc tcgtataatg 4921 tgtggatttt gagttaggat ccggcgagat tttcaggagc taaggaagct aaaatggaga 4981 aaaaaatcac tggatatacc accgttgata tatcccaatg gcatcgtaaa gaacattttg 5041 aggcatttca gtcagttgct caatgtacct ataaccagac cgttcagctg gatattacgg 5101 cctttttaaa gaccgtaaag aaaaataagc acaagtttta tccggccttt attcacattc 5161 ttgcccgcct gatgaatgct catccggaat tccgtatggc aatgaaagac ggtgagctgg 5221 tgatatggga tagtgttcac ccttgttaca ccgttttcca tgagcaaact gaaacgtttt 5281 catcgctctg gagtgaatac cacgacgatt tccggcagtt tctacacata tattcgcaag 5341 atgtggcgtg ttacggtgaa aacctggcct atttccctaa agggtttatt gagaatatgt 5401 ttttcgtctc agccaatccc tgggtgagtt tcaccagttt tgatttaaac gtggccaata 5461 tggacaactt cttcgccccc gttttcacca tgggcaaata ttatacgcaa ggcgacaagg 5521 tgctgatgcc gctggcgatt caggttcatc atgccgtctg tgatggcttc catgtcggca 5581 gaatgcttaa tgaattacaa cagtactgcg atgagtggca gggcggggcg taaagatctg 5641 gatccggctt actaaaagcc agataacagt atgcgtattt gcgcgctgat ttttgcggta 5701 taagaatata tactgatatg tatacccgaa gtatgtcaaa aagaggtgtg ctatgaagca 5761 gcgtattaca gtgacagttg acagcgacag ctatcagttg ctcaaggcat atatgatgtc 5821 aatatctccg gtctggtaag cacaaccatg cagaatgaag cccgtcgtct gcgtgccgaa 5881 cgctggaaag cggaaaatca ggaagggatg gctgaggtcg cccggtttat tgaaatgaac 5941 ggctcttttg ctgacgagaa cagggactgg tgaaatgcag tttaaggttt acacctataa 6001 aagagagagc cgttatcgtc tgtttgtgga tgtacagagt gatattattg acacgcccgg 6061 gcgacggatg gtgatccccc tggccagtgc acgtctgctg tcagataaag tctcccgtga 6121 actttacccg gtggtgcata tcggggatga aagctggcgc atgatgacca ccgatatggc 6181 cagtgtgccg gtctccgtta tcggggaaga agtggctgat ctcagccacc gcgaaaatga 6241 catcaaaaac gccattaacc tgatgttctg gggaatataa atgtcaggct ccgttataca 6301 cagccagtct gcaggtcgac catagtgact ggatatgttg tgttttacag tattatgtag 6361 tctgtttttt atgcaaaatc taatttaata tattgatatt tatatcattt tacgtttctc 6421 gttcagcttt cttgtacaaa gtggttgata tccagcacag tggcggccgc tcgacaatca 6481 acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt 6541 tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gctattgctt cccgtatggc 6601 tttcattttc tcctccttgt ataaatcctg gttgctgtct ctttatgagg agttgtggcc 6661 cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg 6721 gggcattgcc accacctgtc agctcctttc cgggactttc gctttccccc tccctattgc 6781 cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg acaggggctc ggctgttggg 6841 cactgacaat tccgtggtgt tgtcggggaa gctgacgtcc tttccatggc tgctcgcctg 6901 tgttgccacc tggattctgc gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc 6961 agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct 7021 tcgccctcag acgagtcgga tctccctttg ggccgcctcc ccgcctggaa ttcgagctcg 7081 gtacctttaa gaccaatgac ttacaaggca gctgtagatc ttagccactt tttaaaagaa 7141 aaggggggac tggaagggct agctcactcc caacgaagac aagatctgct ttttgcttgt 7201 actgggtctc tctggttaga ccagatctga gcctgggagc tctctggctg cctagggaac 7261 ccactgctta agcctcaata aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg 7321 ttgtgtgact ctggtaacta gagatccctc agaccctttt agtcagtgtg gaaaatctct 7381 agcagtagta gttcatgtca tcttattatt cagtatttat aacttgcaaa gaaatgaata 7441 tcagagagtg agaggaactt gtttattgca gcttataatg gttacaaata aagcaatagc 7501 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa 7561 ctcatcaatg tatcttatca tgtctggctc tagctatccc gcccctaact ccgcccagtt 7621 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg 7681 cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt 7741 gcgtcgagac gtacccaatt cgccctatag tgagtcgtat tacgcgcgct cactggccgt 7801 cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc 7861 acatccccct ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca 7921 acagttgcgc agcctgaatg gcgaatggcg cgacgcgccc tgtagcggcg cattaagcgc 7981 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc 8041 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct 8101 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa 8161 acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc 8221 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact 8281 caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg 8341 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgtt 8401 tacaatttcc //