LOCUS Exported 7425 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Constitutive expression of mTagBFP2 using PGK promoter in a lentiviral backbone.. ACCESSION . VERSION . KEYWORDS pRRlsinPGK_BFP2_WPRE SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7425) TITLE Torok-Storb lab lentiviral fluorescent protein JOURNAL Unpublished REFERENCE 2 (bases 1 to 7425) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7425 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(16..33) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(187..775) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(267..286) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(946..1806) /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 1569..1588 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(1807..1911) /gene="bla" /label=AmpR promoter rep_origin complement(1937..2392) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2074..2095) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 2286..2305 /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 2519..2541 /label=M13/pUC Forward /note="In lacZ gene" primer_bind 2533..2550 /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind 2534..2550 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind 2560..2579 /label=T7 /note="T7 promoter, forward primer" promoter 2560..2578 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" rep_origin 2599..2734 /label=SV40 ori /note="SV40 origin of replication" primer_bind complement(2653..2672) /label=SV40pro-F /note="SV40 promoter/origin, forward primer" polyA_signal 2774..2895 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2785..2804) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 2839..2858 /label=SV40pA-R /note="SV40 polyA, reverse primer" LTR 2967..3200 /label=3' LTR (Delta-U3) /note="self-inactivating 3' long terminal repeat (LTR) from HIV-1" misc_feature 3287..3875 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" CDS 3393..3404 /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" primer_bind 3802..3822 /label=WPRE-R /note="WPRE, reverse primer" CDS complement(3937..4647) /codon_start=1 /product="enhanced monomeric blue fluorescent protein (Subach et al., 2011)" /label=mTagBFP2 /note="mammalian codon-optimized" /translation="MVSKGEELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTM RIKVVEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGG VLTATQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRND MALKLVGGSHLIANAKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAV ARYCDLPSKLGHKLN" regulatory 4644..4653 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" promoter complement(4669..5179) /label=hPGK promoter /note="human phosphoglycerate kinase 1 promoter" primer_bind complement(4732..4750) /label=hPGK-F /note="Human PGK promoter, forward primer" misc_feature 5225..5342 /label=cPPT/CTS /note="central polypurine tract and central termination sequence of HIV-1" CDS complement(5609..5653) /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 5838..6071 /label=RRE /note="The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm." misc_feature 6564..6689 /label=HIV-1 Psi /note="packaging signal of human immunodeficiency virus type 1" LTR 6736..6916 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from HIV-1" promoter complement(6917..7143) /label=RSV promoter /note="Rous sarcoma virus enhancer/promoter" primer_bind complement(7171..7191) /label=T3 /note="T3 promoter, forward primer" promoter complement(7171..7189) /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" primer_bind complement(7210..7226) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(7210..7226) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(7223..7245) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 7234..7250 /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(7258..7288) /label=lac promoter /note="promoter for the E. coli lac operon" ORIGIN 1 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc 61 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg 121 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct 181 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca 241 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct 301 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc 361 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt 421 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc 481 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc 541 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg 601 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc 661 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag 721 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga 781 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat 841 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag 901 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat 961 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc 1021 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat 1081 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag 1141 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg 1201 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc 1261 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca 1321 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg 1381 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc 1441 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta 1501 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc 1561 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg 1621 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc 1681 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc 1741 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat 1801 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag 1861 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc 1921 ccgaaaagtg ccacctaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt 1981 tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca 2041 aaagaataga ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta 2101 aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta 2161 cgtgaaccat caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg 2221 aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga 2281 aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg 2341 ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtcccattcg 2401 ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc 2461 cagctggcga aagggggatg tgctgcaagg cgattaagtt gggtaacgcc agggttttcc 2521 cagtcacgac gttgtaaaac gacggccagt gagcgcgcgt aatacgactc actatagggc 2581 gaattgggta cgtccctagg cctccaaaaa agcctcctca ctacttctgg aatagctcag 2641 aggccgaggc ggcctcggcc tctgcataaa taaaaaaaat tagtcagcca tggggcggag 2701 aatgggcgga actgggcgga gttaggggcg ggatgggcgg agttaggggc gggatagcta 2761 gagccagaca tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga 2821 aaaaaatgct ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc 2881 tgcaataaac aagttcctct cactctctga tattcatttc tttgcaagtt ataaatactg 2941 aataataaga tgacatgaac tactactgct agagattttc cacactgact aaaagggtct 3001 gagggatctc tagttaccag agtcacacaa cagacgggca cacactactt gaagcactca 3061 aggcaagctt tattgaggct taagcagtgg gttccctagt tagccagaga gctcccaggc 3121 tcagatctgg tctaaccaga gagacccagt acaagcaaaa agcagatctt gtcttcgttg 3181 ggagtgaatt agcccttcca gtcccccctt ttcttttaaa aagtggctaa gatctacagc 3241 tgccttgtaa gtcattggtc ttaaaggtac cgagctcgaa ttccaggcgg ggaggcggcc 3301 caaagggaga tccgactcgt ctgagggcga aggcgaagac gcggaagagg ccgcagagcc 3361 ggcagcaggc cgcgggaagg aaggtccgct ggattgaggg ccgaagggac gtagcagaag 3421 gacgtcccgc gcagaatcca ggtggcaaca caggcgagca gccatggaaa ggacgtcagc 3481 ttccccgaca acaccacgga attgtcagtg cccaacagcc gagcccctgt ccagcagcgg 3541 gcaaggcagg cggcgatgag ttccgccgtg gcaataggga gggggaaagc gaaagtcccg 3601 gaaaggagct gacaggtggt ggcaatgccc caaccagtgg gggttgcgtc agcaaacaca 3661 gtgcacacca cgccacgttg cctgacaacg ggccacaact cctcataaag agacagcaac 3721 caggatttat acaaggagga gaaaatgaaa gccatacggg aagcaatagc atgatacaaa 3781 ggcattaaag cagcgtatcc acatagcgta aaaggagcaa catagttaag aataccagtc 3841 aatctttcac aaattttgta atccagaggt tgattgtcga ctgcagttcg aacccgggca 3901 tatgtctaga accggtggat cagtcgcggc cgcttaatta agcttgtgcc ccagtttgct 3961 agggaggtcg cagtatctgg ccactgccac ctcgtgctgc tcgacgtagg tctcgttgtt 4021 ggcctccttg attctttcca gtctgtagtc cacatagtag acgccaggca tcttgaggtt 4081 cttagcgggt ttcttggatc tatatgtggt cttggcgttt gcgatcagat ggctcccgcc 4141 cacgagcttc agggccatgt cgtttctgcc ttccaggccg ccgtcagcgg ggtacagcgt 4201 ctcggtgaag gcctcccagc cgagtgtttt cttctgcatc acagggccgt tggatgtgaa 4261 gttcacccct ctgatcttga cgttgtagat gaggcagccg tcctggaggc tggtgtcctg 4321 ggtagcggtc agcacgcccc cgtcttcgta tgtggtgact ctctcccatg tgaagccctc 4381 agggaaggac tgcttgaaga agtcggggat gccctgggtg tggttgatga aggtcttgct 4441 gccgtagagg aagctagtag ccaggatgtc gaaggcgaag gggagagggc cgccctcgac 4501 caccttgatt ctcatggtct gggtgccctc gtagggcttg ccttcgccct cggatgtgca 4561 cttgaagtga tggttgtcca cggtgccctc catgtacagc ttcatgtgca tgttctcctt 4621 aatcagctct tcgcccttag acaccatggt ggcgaccggt ggatccccct ggggagagag 4681 gtcggtgatt cggtcaacga gggagccgac tgccgacgtg cgctccggag gcttgcagaa 4741 tgcggaacac cgcgcgggca ggaacagggc ccacactacc gccccacacc ccgcctcccg 4801 caccgcccct tcccggccgc tgctctcggc gcgccctgct gagcagccgc tattggccac 4861 agcccatcgc ggtcggcgcg ctgccattgc tccctggcgc tgtccgtctg cgagggtact 4921 agtgagacgt gcggcttccg tttgtcacgt ccggcacgcc gcgaaccgca aggaaccttc 4981 ccgacttagg ggcggagcag gaagcgtcgc cggggggccc acaagggtag cggcgaagat 5041 ccgggtgacg ctgcgaacgg acgtgaagaa tgtgcgagac ccagggtcgg cgccgctgcg 5101 tttcccggaa ccacgcccag agcagccgcg tccctgcgca aacccagggc tgccttggaa 5161 aaggcgcaac cccaaccccg tggaattcga tatcaagctt ctcgaggcta gtctcgtgat 5221 cgataaaatt ttgaattttt gtaatttgtt tttgtaattc tttagtttgt atgtctgttg 5281 ctattatgtc tactattctt tcccctgcac tgtacccccc aatcccccct tttcttttaa 5341 aagttaaccg ataccgtcga gatccgttca ctaatcgaat ggatctgtct ctgtctctct 5401 ctccaccttc ttcttctatt ccttcgggcc tgtcgggtcc cctcggggtt gggaggtggg 5461 tctgaaacga taatggtgaa tatccctgcc taactctatt cactatagaa agtacagcaa 5521 aaactattct taaacctacc aagcctccta ctatcattat gaataatttt atataccaca 5581 gccaatttgt tatgttaaac caattccaca aacttgccca tttatctaat tccaataatt 5641 cttgttcatt cttttcttgc tggttttgcg attcttcaat taaggagtgt attaagcttg 5701 tgtaattgtt aatttctctg tcccactcca tccaggtcgt gtgattccaa atctgttcca 5761 gagatttatt actccaacta gcattccaag gcacagcagt ggtgcaaatg agttttccag 5821 agcaacccca aatccccagg agctgttgat cctttaggta tctttccaca gccaggattc 5881 ttgcctggag ctgcttgatg ccccagactg tgagttgcaa cagatgctgt tgcgcctcaa 5941 tagccctcag caaattgttc tgctgctgca ctataccaga caataattgt ctggcctgta 6001 ccgtcagcgt cattgacgct gcgcccatag tgcttcctgc tgctcccaag aacccaagga 6061 acaaagctcc tattcccact gctctttttt ctctctgcac cactcttctc tttgccttgg 6121 tgggtgctac tcctaatggt tcaattttta ctactttata tttatataat tcacttctcc 6181 aattgtccct catatctcct cctccaggtc tgaagatcag cggccgcttg ctgtgcggtg 6241 gtcttacttt tgttttgctc ttcctctatc ttgtctaaag cttccttggt gtcttttatc 6301 tctatccttt gatgcacaca atagagggtt gctactgtat tatataatga tctaagttct 6361 tctgatcctg tctgaaggga tggttgtagc tgtcccagta tttgtctaca gccttctgat 6421 gtttctaaca ggccaggatt aactgcgaat cgttctagct ccctgcttgc ccatactata 6481 tgttttaatt tatatttttt ctttccccct ggccttaacc gaattttttc ccatcgcgat 6541 ctaattctcc cccgcttaat actgacgctc tcgcacccat ctctctcctt ctagcctccg 6601 ctagtcaaaa tttttggcgt actcaccagt cgccgcccct cgcctcttgc cgtgcgcgct 6661 tcagcaagcc gagtcctgcg tcgagagagc tcctctggtt tccctttcgc tttcaagtcc 6721 ctgttcgggc gccactgcta gagattttcc acactgacta aaagggtctg agggatctct 6781 agttaccaga gtcacacaac agacgggcac acactacttg aagcactcaa ggcaagcttt 6841 attgaggctt aagcagtggg ttccctagtt agccagagag ctcccaggct cagatctggt 6901 ctaaccagag agacccgttt atgtatcgag ctaggcactt aaatacaata tctctgcaat 6961 gcggcaattc agtggttcgt ccaatccatg tcagacccgt ctgttgcctt cctaataagg 7021 cacgatcgta ccaccttact tccaccaatc ggcatgcacg gtgctttttc tctccttgta 7081 aggcatgttg ctaactcatc gttaccatgt tgcaagacta caagagtatt gcataagact 7141 acattaagct tgcagctcca gcttttgttc cctttagtga gggttaattg cgcgcttggc 7201 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa 7261 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac 7321 cttaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca 7381 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgg //