LOCUS Exported 7615 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS pLV-EBFP2-nuc SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7615) TITLE Tsoulfas Lab plasmids JOURNAL Unpublished REFERENCE 2 (bases 1 to 7615) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7615 /organism="synthetic DNA construct" /mol_type="other DNA" misc_feature 97..685 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(150..170) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(568..579) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" LTR 772..1005 /label=3' LTR (Delta-U3) /note="self-inactivating 3' long terminal repeat (LTR) from HIV-1" polyA_signal 1077..1198 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(1114..1133) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 1168..1187 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" rep_origin 1238..1373 /label=SV40 ori /note="SV40 origin of replication" primer_bind 1300..1319 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" primer_bind complement(1393..1412) /label=T7 /note="T7 promoter, forward primer" promoter complement(1394..1412) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(1422..1439) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(1422..1438) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(1431..1453) /label=M13/pUC Forward /note="In lacZ gene" rep_origin 1580..2035 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(1667..1686) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 1877..1898 /label=F1ori-F /note="F1 origin, forward primer" promoter 2061..2165 /gene="bla" /label=AmpR promoter CDS 2166..3026 /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(2384..2403) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 3197..3785 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 3686..3705 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 3939..3956 /label=L4440 /note="L4440 vector, forward primer" protein_bind 4073..4094 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 4109..4139 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 4147..4163 /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 4152..4174 /label=M13/pUC Reverse /note="In lacZ gene" primer_bind 4171..4187 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 4171..4187 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind 4206..4226 /label=T3 /note="T3 promoter, forward primer" promoter 4208..4226 /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" promoter 4254..4480 /label=RSV promoter /note="Rous sarcoma virus enhancer/promoter" LTR 4481..4661 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from HIV-1" misc_feature 4708..4833 /label=HIV-1 Psi /note="packaging signal of human immunodeficiency virus type 1" misc_feature 5326..5559 /label=RRE /note="The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm." CDS 5744..5788 /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 6055..6171 /label=cPPT/CTS /note="central polypurine tract and central termination sequence of HIV-1" enhancer 6194..6497 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 6498..6701 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 6652..6672 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" primer_bind 6698..6722 /label=LNCX /note="Human CMV promoter, forward primer" regulatory 6777..6786 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 6783..7499 /codon_start=1 /product="enhanced blue variant of GFP (Ai et al., 2007)" /label=EBFP2 /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL KFICTTGKLPVPWPTLVTTLSHGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GTYKTRAEVKFEGDTLVNRIELKGVDFKEDGNILGHKLEYNFNSHNIYIMAVKQKNGIK VNFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDSHYLSTQSVLSKDPNEKRDHMVLL EFRTAAGITLGMDELYK" primer_bind complement(6828..6849) /label=EGFP-N /note="EGFP, reverse primer" CDS 7524..7544 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 7548..7568 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 7572..7592 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" ORIGIN 1 gatcccaccg gtgctcgaga tgcatactag tgaattcacg cgttctagac ctgcagggtc 61 gactccggat gatcagggcc ctgtacagat atcgacaatc aacctctgga ttacaaaatt 121 tgtgaaagat tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct 181 gctttaatgc ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg 241 tataaatcct ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc 301 gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt 361 cagctccttt ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc 421 gcctgccttg cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg 481 ttgtcgggga agctgacgtc ctttccatgg ctgctcgcct gtgttgccac ctggattctg 541 cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc 601 ggcctgctgc cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg 661 atctcccttt gggccgcctc cccgcctgga attcgagctc ggtaccttta agaccaatga 721 cttacaaggc agctgtagat cttagccact ttttaaaaga aaagggggga ctggaagggc 781 taattcactc ccaacgaaga caagatctgc tttttgcttg tactgggtct ctctggttag 841 accagatctg agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat 901 aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact 961 agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtagt agttcatgtc 1021 atcttattat tcagtattta taacttgcaa agaaatgaat atcagagagt gagaggaact 1081 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata 1141 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc 1201 atgtctggct ctagctatcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag 1261 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc 1321 cgcctcggcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctagggac 1381 gtacccaatt cgccctatag tgagtcgtat tacgcgcgct cactggccgt cgttttacaa 1441 cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc acatccccct 1501 ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca acagttgcgc 1561 agcctgaatg gcgaatggga cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg 1621 gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc 1681 ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc 1741 cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt 1801 gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag 1861 tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg 1921 gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag 1981 ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgcttac aatttaggtg 2041 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa 2101 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga 2161 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc 2221 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg 2281 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc 2341 gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat 2401 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg 2461 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag 2521 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa 2581 cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc 2641 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca 2701 cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc 2761 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc 2821 tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg 2881 ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta 2941 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag 3001 gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga 3061 ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc 3121 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa 3181 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa 3241 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc 3301 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt 3361 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc 3421 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac 3481 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca 3541 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg 3601 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag 3661 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt 3721 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat 3781 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc 3841 acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt 3901 gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag 3961 cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca 4021 gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga 4081 gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt 4141 gtggaattgt gagcggataa caatttcaca caggaaacag ctatgaccat gattacgcca 4201 agcgcgcaat taaccctcac taaagggaac aaaagctgga gctgcaagct taatgtagtc 4261 ttatgcaata ctcttgtagt cttgcaacat ggtaacgatg agttagcaac atgccttaca 4321 aggagagaaa aagcaccgtg catgccgatt ggtggaagta aggtggtacg atcgtgcctt 4381 attaggaagg caacagacgg gtctgacatg gattggacga accactgaat tgccgcattg 4441 cagagatatt gtatttaagt gcctagctcg atacataaac gggtctctct ggttagacca 4501 gatctgagcc tgggagctct ctggctaact agggaaccca ctgcttaagc ctcaataaag 4561 cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg gtaactagag 4621 atccctcaga cccttttagt cagtgtggaa aatctctagc agtggcgccc gaacagggac 4681 ttgaaagcga aagggaaacc agaggagctc tctcgacgca ggactcggct tgctgaagcg 4741 cgcacggcaa gaggcgaggg gcggcgactg gtgagtacgc caaaaatttt gactagcgga 4801 ggctagaagg agagagatgg gtgcgagagc gtcagtatta agcgggggag aattagatcg 4861 cgatgggaaa aaattcggtt aaggccaggg ggaaagaaaa aatataaatt aaaacatata 4921 gtatgggcaa gcagggagct agaacgattc gcagttaatc ctggcctgtt agaaacatca 4981 gaaggctgta gacaaatact gggacagcta caaccatccc ttcagacagg atcagaagaa 5041 cttagatcat tatataatac agtagcaacc ctctattgtg tgcatcaaag gatagagata 5101 aaagacacca aggaagcttt agacaagata gaggaagagc aaaacaaaag taagaccacc 5161 gcacagcaag cggccgctga tcttcagacc tggaggagga gatatgaggg acaattggag 5221 aagtgaatta tataaatata aagtagtaaa aattgaacca ttaggagtag cacccaccaa 5281 ggcaaagaga agagtggtgc agagagaaaa aagagcagtg ggaataggag ctttgttcct 5341 tgggttcttg ggagcagcag gaagcactat gggcgcagcg tcaatgacgc tgacggtaca 5401 ggccagacaa ttattgtctg gtatagtgca gcagcagaac aatttgctga gggctattga 5461 ggcgcaacag catctgttgc aactcacagt ctggggcatc aagcagctcc aggcaagaat 5521 cctggctgtg gaaagatacc taaaggatca acagctcctg gggatttggg gttgctctgg 5581 aaaactcatt tgcaccactg ctgtgccttg gaatgctagt tggagtaata aatctctgga 5641 acagatttgg aatcacacga cctggatgga gtgggacaga gaaattaaca attacacaag 5701 cttaatacac tccttaattg aagaatcgca aaaccagcaa gaaaagaatg aacaagaatt 5761 attggaatta gataaatggg caagtttgtg gaattggttt aacataacaa attggctgtg 5821 gtatataaaa ttattcataa tgatagtagg aggcttggta ggtttaagaa tagtttttgc 5881 tgtactttct atagtgaata gagttaggca gggatattca ccattatcgt ttcagaccca 5941 cctcccaacc ccgaggggac ccgacaggcc cgaaggaata gaagaagaag gtggagagag 6001 agacagagac agatccattc gattagtgaa cggatctcga cggtatcggt taacttttaa 6061 aagaaaaggg gggattgggg ggtacagtgc aggggaaaga atagtagaca taatagcaac 6121 agacatacaa actaaagaat tacaaaaaca aattacaaaa ttcaaaattt tatcgataag 6181 cttgggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg 6241 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt 6301 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag 6361 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc 6421 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag 6481 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt 6541 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc 6601 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg 6661 gcggtaggcg tgtacggtgg gaggtctata taagcagagc tcgtttagtg aaccgtcaga 6721 tcgcctggag acgccatcca cgctgttttg acctccatag aagacaccgc ccggtcgcca 6781 ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg 6841 acggcgacgt aaacggccac aagttcagcg tgaggggcga gggcgagggc gatgccacca 6901 acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg ccctggccca 6961 ccctcgtgac caccctgagc cacggcgtgc agtgcttcgc ccgctacccc gaccacatga 7021 agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct 7081 tcttcaagga cgacggcacc tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc 7141 tagtgaaccg catcgagctg aagggcgtcg acttcaagga ggacggcaac atcctggggc 7201 acaagctgga gtacaacttc aacagccaca acatctatat catggccgtc aagcagaaga 7261 acggcatcaa ggtgaacttc aagatccgcc acaacgtgga ggacggcagc gtgcagctcg 7321 ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg cccgacagcc 7381 actacctgag cacccagtcc gtgctgagca aagaccccaa cgagaagcgc gatcacatgg 7441 tcctgctgga gttccgcacc gccgccggga tcactctcgg catggacgag ctgtacaagt 7501 ccggactcag atctcgagct gatccaaaaa agaagagaaa ggtagatcca aaaaagaaga 7561 gaaaggtaga tccaaaaaag aagagaaagg taggatccac cggatctaga taact //