LOCUS Exported 8473 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Lentiviral expression of Tet inducible TagBFP. ACCESSION . VERSION . KEYWORDS pLVX TRE 3G BFP SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 8473) TITLE Emory Custom Cloning Core Plasmids - Oskar Laur REFERENCE 2 (bases 1 to 8473) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..8473 /organism="synthetic DNA construct" /mol_type="other DNA" LTR 1..634 /label=3' LTR /note="3' long terminal repeat (LTR) from HIV-1" misc_feature 681..806 /label=HIV-1 Psi /note="packaging signal of human immunodeficiency virus type 1" misc_feature 1303..1536 /label=RRE /note="The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm." CDS 1721..1765 /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 2028..2143 /label=cPPT/CTS /note="central polypurine tract and central termination sequence of HIV-1 (lacking the first T)" promoter 2206..2555 /label=TRE3GV promoter /note="3rd-generation Tet-responsive promoter that can be activated by binding of Tet-On(R) 3G, optimized for retroviral and lentiviral vectors" protein_bind 2214..2232 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 2250..2268 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 2286..2304 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 2322..2340 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 2358..2376 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 2394..2412 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 2430..2448 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" primer_bind 2494..2518 /label=LNCX /note="Human CMV promoter, forward primer" CDS 2594..3295 /codon_start=1 /product="monomeric blue fluorescent protein" /label=TagBFP /note="mammalian codon-optimized" /translation="MSELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKV VEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTA TQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALK LVGGSHLIANIKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYC DLPSKLGHKLN" promoter 3306..3805 /label=PGK promoter /note="mouse phosphoglycerate kinase 1 promoter" primer_bind complement(3341..3364) /label=MSCV-rev /note="Murine stem cell virus, reverse primer" primer_bind 3738..3757 /label=mPGK-F /note="Mouse PGK promoter, forward primer" CDS 3826..4425 /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(3826..3845) /label=Puro-R /note="Puromycin resistance gene, reverse primer. Also called puro-variant-R" primer_bind 4322..4342 /label=Puro-F /note="Puromycin resistance gene, forward primer" misc_feature 4442..5030 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(4495..4515) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(4913..4924) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" LTR 5237..5870 /label=3' LTR /note="3' long terminal repeat (LTR) from HIV-1" primer_bind complement(5998..6014) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(5998..6014) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(6011..6033) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 6022..6038 /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)." promoter complement(6046..6076) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 6091..6112 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(6229..6246) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(6400..6988) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(6480..6499) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(7159..8019) /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 7782..7801 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(8020..8124) /gene="bla" /label=AmpR promoter polyA_signal 8172..8306 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(8209..8228) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 8263..8282 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" ORIGIN 1 tggaagggct aattcactcc caaagaagac aagatatcct tgatctgtgg atctaccaca 61 cacaaggcta cttccctgat tagcagaact acacaccagg gccaggggtc agatatccac 121 tgacctttgg atggtgctac aagctagtac cagttgagcc agataaggta gaagaggcca 181 ataaaggaga gaacaccagc ttgttacacc ctgtgagcct gcatgggatg gatgacccgg 241 agagagaagt gttagagtgg aggtttgaca gccgcctagc atttcatcac gtggcccgag 301 agctgcatcc ggagtacttc aagaactgct gatatcgagc ttgctacaag ggactttccg 361 ctggggactt tccagggagg cgtggcctgg gcgggactgg ggagtggcga gccctcagat 421 cctgcatata agcagctgct ttttgcctgt actgggtctc tctggttaga ccagatctga 481 gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct 541 tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc 601 agaccctttt agtcagtgtg gaaaatctct agcagtggcg cccgaacagg gacttgaaag 661 cgaaagggaa accagaggag ctctctcgac gcaggactcg gcttgctgaa gcgcgcacgg 721 caagaggcga ggggcggcga ctggtgagta cgccaaaaat tttgactagc ggaggctaga 781 aggagagaga tgggtgcgag agcgtcagta ttaagcgggg gagaattaga tcgcgatggg 841 aaaaaattcg gttaaggcca gggggaaaga aaaaatataa attaaaacat atagtatggg 901 caagcaggga gctagaacga ttcgcagtta atcctggcct gttagaaaca tcagaaggct 961 gtagacaaat actgggacag ctacaaccat cccttcagac aggatcagaa gaacttagat 1021 cattatataa tacagtagca accctctatt gtgtgcatca aaggatagag ataaaagaca 1081 ccaaggaagc tttagacaag atagaggaag agcaaaacaa aagtaagacc accgcacagc 1141 aagcggccgg ccgctgatct tcagacctgg aggaggagat atgagggaca attggagaag 1201 tgaattatat aaatataaag tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc 1261 aaagagaaga gtggtgcaga gagaaaaaag agcagtggga ataggagctt tgttccttgg 1321 gttcttggga gcagcaggaa gcactatggg cgcagcgtca atgacgctga cggtacaggc 1381 cagacaatta ttgtctggta tagtgcagca gcagaacaat ttgctgaggg ctattgaggc 1441 gcaacagcat ctgttgcaac tcacagtctg gggcatcaag cagctccagg caagaatcct 1501 ggctgtggaa agatacctaa aggatcaaca gctcctgggg atttggggtt gctctggaaa 1561 actcatttgc accactgctg tgccttggaa tgctagttgg agtaataaat ctctggaaca 1621 gatttggaat cacacgacct ggatggagtg ggacagagaa attaacaatt acacaagctt 1681 aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt 1741 ggaattagat aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt ggctgtggta 1801 tataaaatta ttcataatga tagtaggagg cttggtaggt ttaagaatag tttttgctgt 1861 actttctata gtgaatagag ttaggcaggg atattcacca ttatcgtttc agacccacct 1921 cccaaccccg aggggacccg acaggcccga aggaatagaa gaagaaggtg gagagagaga 1981 cagagacaga tccattcgat tagtgaacgg atctcgacgg tatcgccttt aaaagaaaag 2041 gggggattgg ggggtacagt gcaggggaaa gaatagtaga cataatagca acagacatac 2101 aaactaaaga attacaaaaa caaattacaa aaattcaaaa ttttcgggtt tattacaggg 2161 acagcagaga tccagtttat cgatgaggcc ctttcgtctt cactcgagtt tactccctat 2221 cagtgataga gaacgtatga agagtttact ccctatcagt gatagagaac gtatgcagac 2281 tttactccct atcagtgata gagaacgtat aaggagttta ctccctatca gtgatagaga 2341 acgtatgacc agtttactcc ctatcagtga tagagaacgt atctacagtt tactccctat 2401 cagtgataga gaacgtatat ccagtttact ccctatcagt gatagagaac gtatgtcgag 2461 gtaggcgtgt acggtgggcg cctataaaag cagagctcgt ttagtgaacc gtcagatcgc 2521 ctggagcaat tccacaacac ttttgtctta tacttggatc cgggcccgcg gccgccccgg 2581 gacgcgtgcc accatgagcg agctgattaa ggagaacatg cacatgaagc tgtacatgga 2641 gggcaccgtg gacaaccatc acttcaagtg cacatccgag ggcgaaggca agccctacga 2701 gggcacccag accatgagaa tcaaggtggt cgagggcggc cctctcccct tcgccttcga 2761 catcctggct actagcttcc tctacggcag caagaccttc atcaaccaca cccagggcat 2821 ccccgacttc ttcaagcagt ccttccctga gggcttcaca tgggagagag tcaccacata 2881 cgaagacggg ggcgtgctga ccgctaccca ggacaccagc ctccaggacg gctgcctcat 2941 ctacaacgtc aagatcagag gggtgaactt cacatccaac ggccctgtga tgcagaagaa 3001 aacactcggc tgggaggcct tcaccgagac gctgtacccc gctgacggcg gcctggaagg 3061 cagaaacgac atggccctga agctcgtggg cgggagccat ctgatcgcaa acatcaagac 3121 cacatataga tccaagaaac ccgctaagaa cctcaagatg cctggcgtct actatgtgga 3181 ctacagactg gaaagaatca aggaggccaa caacgagacc tacgtcgagc agcacgaggt 3241 ggcagtggcc agatactgcg acctccctag caaactgggg cacaagctta attaagaatt 3301 ctaccgggta ggggaggcgc ttttcccaag gcagtctgga gcatgcgctt tagcagcccc 3361 gctgggcact tggcgctaca caagtggcct ctggcctcgc acacattcca catccaccgg 3421 taggcgccaa ccggctccgt tctttggtgg ccccttcgcg ccaccttcta ctcctcccct 3481 agtcaggaag ttcccccccg ccccgcagct cgcgtcgtgc aggacgtgac aaatggaagt 3541 agcacgtctc actagtctcg tgcagatgga cagcaccgct gagcaatgga agcgggtagg 3601 cctttggggc agcggccaat agcagctttg ctccttcgct ttctgggctc agaggctggg 3661 aaggggtggg tccgggggcg ggctcagggg cgggctcagg ggcggggcgg gcgcccgaag 3721 gtcctccgga ggcccggcat tctgcacgct tcaaaagcgc acgtctgccg cgctgttctc 3781 ctcttcctca tctccgggcc tttcgacctg cagcccaagc ttaccatgac cgagtacaag 3841 cccacggtgc gcctcgccac ccgcgacgac gtccccaggg ccgtacgcac cctcgccgcc 3901 gcgttcgccg actaccccgc cacgcgccac accgtcgatc cggaccgcca catcgagcgg 3961 gtcaccgagc tgcaagaact cttcctcacg cgcgtcgggc tcgacatcgg caaggtgtgg 4021 gtcgcggacg acggcgccgc ggtggcggtc tggaccacgc cggagagcgt cgaagcgggg 4081 gcggtgttcg ccgagatcgg cccgcgcatg gccgagttga gcggttcccg gctggccgcg 4141 cagcaacaga tggaaggcct cctggcgccg caccggccca aggagcccgc gtggttcctg 4201 gccaccgtcg gcgtctcgcc cgaccaccag ggcaagggtc tgggcagcgc cgtcgtgctc 4261 cccggagtgg aggcggccga gcgcgccggg gtgcccgcct tcctggagac ctccgcgccc 4321 cgcaacctcc ccttctacga gcggctcggc ttcaccgtca ccgccgacgt cgaggtgccc 4381 gaaggaccgc gcacctggtg catgacccgc aagcccggtg cctgacgggc gcgtctggaa 4441 caatcaacct ctggattaca aaatttgtga aagattgact ggtattctta actatgttgc 4501 tccttttacg ctatgtggat acgctgcttt aatgcctttg tatcatgcta ttgcttcccg 4561 tatggctttc attttctcct ccttgtataa atcctggttg ctgtctcttt atgaggagtt 4621 gtggcccgtt gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg caacccccac 4681 tggttggggc attgccacca cctgtcagct cctttccggg actttcgctt tccccctccc 4741 tattgccacg gcggaactca tcgccgcctg ccttgcccgc tgctggacag gggctcggct 4801 gttgggcact gacaattccg tggtgttgtc ggggaagctg acgtcctttc catggctgct 4861 cgcctgtgtt gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc cttcggccct 4921 caatccagcg gaccttcctt cccgcggcct gctgccggct ctgcggcctc ttccgcgtct 4981 tcgccttcgc cctcagacga gtcggatctc cctttgggcc gcctccccgc ctggaattaa 5041 ttctgcagtc gagacctaga aaaacatgga gcaatcacaa gtagcaatac agcagctacc 5101 aatgctgatt gtgcctggct agaagcacaa gaggaggagg aggtgggttt tccagtcaca 5161 cctcaggtac ctttaagacc aatgacttac aaggcagctg tagatcttag ccacttttta 5221 aaagaaaaga ggggactgga agggctaatt cactcccaac gaagacaaga tatccttgat 5281 ctgtggatct accacacaca aggctacttc cctgattagc agaactacac accagggcca 5341 ggggtcagat atccactgac ctttggatgg tgctacaagc tagtaccagt tgagccagat 5401 aaggtagaag aggccaataa aggagagaac accagcttgt tacaccctgt gagcctgcat 5461 gggatggatg acccggagag agaagtgtta gagtggaggt ttgacagccg cctagcattt 5521 catcacgtgg cccgagagct gcatccggag tacttcaaga actgctgata tcgagcttgc 5581 tacaagggac tttccgctgg ggactttcca gggaggcgtg gcctgggcgg gactggggag 5641 tggcgagccc tcagatcctg catataagca gctgcttttt gcctgtactg ggtctctctg 5701 gttagaccag atctgagcct gggagctctc tggctaacta gggaacccac tgcttaagcc 5761 tcaataaagc ttgccttgag tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg 5821 taactagaga tccctcagac ccttttagtc agtgtggaaa atctctagca gtagtagttc 5881 atgtcatctt attattcagt atttataact tgcaaagaaa tgaatatcag agagtgagag 5941 gccttgacat tgctagcgtt taccgtcgac ctctagctag agcttggcgt aatcatggtc 6001 atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg 6061 aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt 6121 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg 6181 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga 6241 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat 6301 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca 6361 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc 6421 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata 6481 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc 6541 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc 6601 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga 6661 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc 6721 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag 6781 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag 6841 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag 6901 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca 6961 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga 7021 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat 7081 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga 7141 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg 7201 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga 7261 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc 7321 agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac 7381 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc 7441 agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc 7501 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc 7561 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt 7621 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc 7681 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg 7741 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag 7801 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat 7861 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc 7921 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa 7981 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta 8041 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa 8101 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtcgacgg 8161 atcgggagat caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca 8221 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca 8281 tcaatgtatc ttatcatgtc tggatcaact ggataactca agctaaccaa aatcatccca 8341 aacttcccac cccataccct attaccactg ccaattacct gtggtttcat ttactctaaa 8401 cctgtgattc ctctgaatta ttttcatttt aaagaaattg tatttgttaa atatgtacta 8461 caaacttagt agt //