LOCUS Exported 8955 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Lentiviral expression of SARS-CoV-2 protein; transient expression and generate lentivirus. ACCESSION . VERSION . KEYWORDS pLVX-EF1alpha-SARS-CoV-2-nsp11-2xStrep-IRES-Puro SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 8955) AUTHORS Gordon DE, et al. TITLE A SARS-CoV-2-Human Protein-Protein Interaction Map Reveals Drug Targets and Potential Drug Repurposing JOURNAL BioRxiv March 2020 REFERENCE 2 (bases 1 to 8955) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..8955 /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 2338..3519 /label=EF-1-alpha promoter /note="strong constitutive promoter for human elongation factor EF-1-alpha" intron 2568..3510 /label=EF-1-alpha intron A /note="intron upstream of the start codon of human EF-1-alpha" primer_bind 3467..3487 /label=EF1a-F /note="Human elongation factor-1a promoter, forward primer" CDS 3610..3633 /codon_start=1 /product="peptide that binds Strep-Tactin(R), an engineered form of streptavidin" /label=Strep-Tag II /translation="WSHPQFEK" CDS 3670..3693 /codon_start=1 /product="peptide that binds Strep-Tactin(R), an engineered form of streptavidin" /label=Strep-Tag II /translation="WSHPQFEK" misc_feature 3705..4278 /label=IRES /note="internal ribosome entry site (IRES) of the encephalomyocarditis virus (EMCV)" primer_bind complement(3872..3889) /label=IRES reverse /note="IRES internal ribosome entry site, reverse primer. Also called pCDH-rev" primer_bind 4099..4118 /label=IRES-F /note="IRES internal ribosome entry site, forward primer" CDS 4311..4910 /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(4311..4330) /label=Puro-R /note="Puromycin resistance gene, reverse primer. Also called puro-variant-R" primer_bind 4807..4827 /label=Puro-F /note="Puromycin resistance gene, forward primer" misc_feature 4924..5512 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(4977..4997) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(5395..5406) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" LTR 5719..6352 /label=3' LTR /note="3' long terminal repeat (LTR) from HIV-1" primer_bind complement(6480..6496) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(6480..6496) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(6493..6515) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 6504..6520 /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(6528..6558) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 6573..6594 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(6711..6728) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(6882..7470) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(6962..6981) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(7641..8501) /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 8264..8283 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(8502..8606) /gene="bla" /label=AmpR promoter polyA_signal 8654..8788 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(8691..8710) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 8745..8764 /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 cgatgagtaa ttcatacaaa aggactcgcc cctgccttgg 2221 ggaatcccag ggaccgtcgt taaactccca ctaacgtaga acccagagat cgctgcgttc 2281 ccgccccctc acccgcccgc tctcgtcatc actgaggtgg agaagagcat gcgtgaggct 2341 ccggtgcccg tcagtgggca gagcgcacat cgcccacagt ccccgagaag ttggggggag 2401 gggtcggcaa ttgaaccggt gcctagagaa ggtggcgcgg ggtaaactgg gaaagtgatg 2461 tcgtgtactg gctccgcctt tttcccgagg gtgggggaga accgtatata agtgcagtag 2521 tcgccgtgaa cgttcttttt cgcaacgggt ttgccgccag aacacaggta agtgccgtgt 2581 gtggttcccg cgggcctggc ctctttacgg gttatggccc ttgcgtgcct tgaattactt 2641 ccacgcccct ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg 2701 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg 2761 cttgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct 2821 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg 2881 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc 2941 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga 3001 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct 3061 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca 3121 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg 3181 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg 3241 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat 3301 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg 3361 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa 3421 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca 3481 gtggttcaaa gtttttttct tccatttcag gtgtcgtgag gatctatttc cggtgaattc 3541 gccgccacca tgtcagccga tgcgcagtct ttcttgaacg gtttcgccgt gctcgaaggc 3601 ggcgggggat ggagccatcc acaattcgag aaaggcggtg gttcaggagg aggtagcggg 3661 ggtggatcat ggtcacatcc gcagtttgaa aagtaaggat cccgcccctc tccctccccc 3721 ccccctaacg ttactggccg aagccgcttg gaataaggcc ggtgtgcgtt tgtctatatg 3781 ttattttcca ccatattgcc gtcttttggc aatgtgaggg cccggaaacc tggccctgtc 3841 ttcttgacga gcattcctag gggtctttcc cctctcgcca aaggaatgca aggtctgttg 3901 aatgtcgtga aggaagcagt tcctctggaa gcttcttgaa gacaaacaac gtctgtagcg 3961 accctttgca ggcagcggaa ccccccacct ggcgacaggt gcctctgcgg ccaaaagcca 4021 cgtgtataag atacacctgc aaaggcggca caaccccagt gccacgttgt gagttggata 4081 gttgtggaaa gagtcaaatg gctctcctca agcgtattca acaaggggct gaaggatgcc 4141 cagaaggtac cccattgtat gggatctgat ctggggcctc ggtgcacatg ctttacatgt 4201 gtttagtcga ggttaaaaaa acgtctaggc cccccgaacc acggggacgt ggttttcctt 4261 tgaaaaacac gatgataagc ttgccacaac ccacaaggag acgaccttcc atgaccgagt 4321 acaagcccac ggtgcgcctc gccacccgcg acgacgtccc ccgggccgta cgcaccctcg 4381 ccgccgcgtt cgccgactac cccgccacgc gccacaccgt cgacccggac cgccacatcg 4441 agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt cgggctcgac atcggcaagg 4501 tgtgggtcgc ggacgacggc gccgcggtgg cggtctggac cacgccggag agcgtcgaag 4561 cgggggcggt gttcgccgag atcggcccgc gcatggccga gttgagcggt tcccggctgg 4621 ccgcgcagca acagatggaa ggcctcctgg cgccgcaccg gcccaaggag cccgcgtggt 4681 tcctggccac cgtcggcgtc tcgcccgacc accagggcaa gggtctgggc agcgccgtcg 4741 tgctccccgg agtggaggcg gccgagcgcg ccggggtgcc cgccttcctg gagacctccg 4801 cgccccgcaa cctccccttc tacgagcggc tcggcttcac cgtcaccgcc gacgtcgagg 4861 tgcccgaagg accgcgcacc tggtgcatga cccgcaagcc cggtgcctag acgcgtctgg 4921 aacaatcaac ctctggatta caaaatttgt gaaagattga ctggtattct taactatgtt 4981 gctcctttta cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc tattgcttcc 5041 cgtatggctt tcattttctc ctccttgtat aaatcctggt tgctgtctct ttatgaggag 5101 ttgtggcccg ttgtcaggca acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc 5161 actggttggg gcattgccac cacctgtcag ctcctttccg ggactttcgc tttccccctc 5221 cctattgcca cggcggaact catcgccgcc tgccttgccc gctgctggac aggggctcgg 5281 ctgttgggca ctgacaattc cgtggtgttg tcggggaagc tgacgtcctt tccatggctg 5341 ctcgcctgtg ttgccacctg gattctgcgc gggacgtcct tctgctacgt cccttcggcc 5401 ctcaatccag cggaccttcc ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt 5461 cttcgccttc gccctcagac gagtcggatc tccctttggg ccgcctcccc gcctggaatt 5521 aattctgcag tcgagaccta gaaaaacatg gagcaatcac aagtagcaat acagcagcta 5581 ccaatgctga ttgtgcctgg ctagaagcac aagaggagga ggaggtgggt tttccagtca 5641 cacctcaggt acctttaaga ccaatgactt acaaggcagc tgtagatctt agccactttt 5701 taaaagaaaa gaggggactg gaagggctaa ttcactccca acgaagacaa gatatccttg 5761 atctgtggat ctaccacaca caaggctact tccctgatta gcagaactac acaccagggc 5821 caggggtcag atatccactg acctttggat ggtgctacaa gctagtacca gttgagccag 5881 ataaggtaga agaggccaat aaaggagaga acaccagctt gttacaccct gtgagcctgc 5941 atgggatgga tgacccggag agagaagtgt tagagtggag gtttgacagc cgcctagcat 6001 ttcatcacgt ggcccgagag ctgcatccgg agtacttcaa gaactgctga tatcgagctt 6061 gctacaaggg actttccgct ggggactttc cagggaggcg tggcctgggc gggactgggg 6121 agtggcgagc cctcagatcc tgcatataag cagctgcttt ttgcctgtac tgggtctctc 6181 tggttagacc agatctgagc ctgggagctc tctggctaac tagggaaccc actgcttaag 6241 cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct 6301 ggtaactaga gatccctcag acccttttag tcagtgtgga aaatctctag cagtagtagt 6361 tcatgtcatc ttattattca gtatttataa cttgcaaaga aatgaatatc agagagtgag 6421 aggccttgac attgctagcg tttaccgtcg acctctagct agagcttggc gtaatcatgg 6481 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc 6541 ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg 6601 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc 6661 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact 6721 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta 6781 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag 6841 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc 6901 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta 6961 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg 7021 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc 7081 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac 7141 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac 7201 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg 7261 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga 7321 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt 7381 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag 7441 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct 7501 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg 7561 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat 7621 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc 7681 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg 7741 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct 7801 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca 7861 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg 7921 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg 7981 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc 8041 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag 8101 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg 8161 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag 8221 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat 8281 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg 8341 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca 8401 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca 8461 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat 8521 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag 8581 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtcgac 8641 ggatcgggag atcaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat 8701 cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact 8761 catcaatgta tcttatcatg tctggatcaa ctggataact caagctaacc aaaatcatcc 8821 caaacttccc accccatacc ctattaccac tgccaattac ctgtggtttc atttactcta 8881 aacctgtgat tcctctgaat tattttcatt ttaaagaaat tgtatttgtt aaatatgtac 8941 tacaaactta gtagt //