LOCUS Exported 8928 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Lentiviral vector to express ERK KTR mClover under CMV promoter (With Puromycin Resistance). ACCESSION . VERSION . KEYWORDS pLentiCMV Puro DEST ERKKTRClover SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 8928) AUTHORS Regot S, Hughey JJ, Bajar BT, Carrasco S, Covert MW TITLE High-sensitivity measurements of multiple kinase activities in live single cells. JOURNAL Cell. 2014 Jun 19;157(7):1724-34. doi: 10.1016/j.cell.2014.04.039. PUBMED 24949979 REFERENCE 2 (bases 1 to 8928) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..8928 /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" promoter 2220..2446 /label=RSV promoter /note="Rous sarcoma virus enhancer/promoter" LTR 2447..2627 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from HIV-1" misc_feature 2674..2799 /label=HIV-1 Psi /note="packaging signal of human immunodeficiency virus type 1" misc_feature 3292..3525 /label=RRE /note="The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm." CDS 3710..3754 /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 4021..4137 /label=cPPT/CTS /note="central polypurine tract and central termination sequence of HIV-1" enhancer 4160..4463 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 4464..4667 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 4618..4638 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" primer_bind 4664..4688 /label=LNCX /note="Human CMV promoter, forward primer" protein_bind 4772..4796 /gene="mutant version of attB" /label=attB1 /bound_moiety="BP Clonase(TM)" /note="recombination site for the Gateway(R) BP reaction" primer_bind complement(5099..5120) /label=EGFP-N /note="EGFP, reverse primer" primer_bind 5707..5728 /label=EGFP-C /note="EGFP, forward primer" protein_bind complement(5764..5788) /gene="mutant version of attB" /label=attB2 /bound_moiety="BP Clonase(TM)" /note="recombination site for the Gateway(R) BP reaction" misc_feature 5819..6407 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(5872..5892) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(6290..6301) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" primer_bind complement(6456..6479) /label=MSCV-rev /note="Murine stem cell virus, reverse primer" primer_bind 6841..6860 /label=mPGK-F /note="Mouse PGK promoter, forward primer" CDS 6929..7528 /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(6929..6948) /label=Puro-R /note="Puromycin resistance gene, reverse primer. Also called puro-variant-R" primer_bind 7425..7445 /label=Puro-F /note="Puromycin resistance gene, forward primer" LTR 7666..7899 /label=3' LTR (Delta-U3) /note="self-inactivating 3' long terminal repeat (LTR) from HIV-1" polyA_signal 7971..8092 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(8008..8027) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 8062..8081 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" rep_origin 8132..8267 /label=SV40 ori /note="SV40 origin of replication" primer_bind 8194..8213 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" primer_bind complement(8287..8306) /label=T7 /note="T7 promoter, forward primer" promoter complement(8288..8306) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(8316..8333) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(8316..8332) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(8325..8347) /label=M13/pUC Forward /note="In lacZ gene" rep_origin 8474..1 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(8561..8580) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 8771..8792 /label=F1ori-F /note="F1 origin, forward primer" ORIGIN 1 taggtggcac 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 aaatactgtt 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 ataaacgggt ctctctggtt 2461 agaccagatc tgagcctggg agctctctgg ctaactaggg aacccactgc ttaagcctca 2521 ataaagcttg ccttgagtgc ttcaagtagt gtgtgcccgt ctgttgtgtg actctggtaa 2581 ctagagatcc ctcagaccct tttagtcagt gtggaaaatc tctagcagtg gcgcccgaac 2641 agggacttga aagcgaaagg gaaaccagag gagctctctc gacgcaggac tcggcttgct 2701 gaagcgcgca cggcaagagg cgaggggcgg cgactggtga gtacgccaaa aattttgact 2761 agcggaggct agaaggagag agatgggtgc gagagcgtca gtattaagcg ggggagaatt 2821 agatcgcgat gggaaaaaat tcggttaagg ccagggggaa agaaaaaata taaattaaaa 2881 catatagtat gggcaagcag ggagctagaa cgattcgcag ttaatcctgg cctgttagaa 2941 acatcagaag gctgtagaca aatactggga cagctacaac catcccttca gacaggatca 3001 gaagaactta gatcattata taatacagta gcaaccctct attgtgtgca tcaaaggata 3061 gagataaaag acaccaagga agctttagac aagatagagg aagagcaaaa caaaagtaag 3121 accaccgcac agcaagcggc cgctgatctt cagacctgga ggaggagata tgagggacaa 3181 ttggagaagt gaattatata aatataaagt agtaaaaatt gaaccattag gagtagcacc 3241 caccaaggca aagagaagag tggtgcagag agaaaaaaga gcagtgggaa taggagcttt 3301 gttccttggg ttcttgggag cagcaggaag cactatgggc gcagcgtcaa tgacgctgac 3361 ggtacaggcc agacaattat tgtctggtat agtgcagcag cagaacaatt tgctgagggc 3421 tattgaggcg caacagcatc tgttgcaact cacagtctgg ggcatcaagc agctccaggc 3481 aagaatcctg gctgtggaaa gatacctaaa ggatcaacag ctcctgggga tttggggttg 3541 ctctggaaaa ctcatttgca ccactgctgt gccttggaat gctagttgga gtaataaatc 3601 tctggaacag atttggaatc acacgacctg gatggagtgg gacagagaaa ttaacaatta 3661 cacaagctta atacactcct taattgaaga atcgcaaaac cagcaagaaa agaatgaaca 3721 agaattattg gaattagata aatgggcaag tttgtggaat tggtttaaca taacaaattg 3781 gctgtggtat ataaaattat tcataatgat agtaggaggc ttggtaggtt taagaatagt 3841 ttttgctgta ctttctatag tgaatagagt taggcaggga tattcaccat tatcgtttca 3901 gacccacctc ccaaccccga ggggacccga caggcccgaa ggaatagaag aagaaggtgg 3961 agagagagac agagacagat ccattcgatt agtgaacgga tctcgacggt atcggttaac 4021 ttttaaaaga aaagggggga ttggggggta cagtgcaggg gaaagaatag tagacataat 4081 agcaacagac atacaaacta aagaattaca aaaacaaatt acaaaattca aaattttatc 4141 gataagcttg ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc 4201 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag 4261 ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac 4321 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg 4381 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg 4441 tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat 4501 agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt 4561 tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc 4621 aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctcgt ttagtgaacc 4681 gtcagatcgc ctggagacgc catccacgct gttttgacct ccatagaaga caccgactct 4741 agtccagtgt ggtggaattc tgcagatatc aacaagtttg tacaaaaaag caggctccac 4801 catgaagggc cgaaagcctc gggacctgga acttccactt agcccaagcc tgctgggtgg 4861 ccagggacct gaacggactc caggatcagg aacaagctct ggtcttcagg caccggggcc 4921 agcgctttcg cccagcaagc gtagcggtct cgaggaccca gcgacgccat cgaagaagcc 4981 acgtacccca tcggtgtcgt caaggctgga gcgcctgacc ctccagtcca gctttcaatt 5041 cccatccact agtatggtga gcaagggcga ggagctgttc accggggttg tgcccatcct 5101 ggtcgagctg gacggcgacg taaacggcca caagttcagc gtccgcggcg agggcgaggg 5161 cgatgccacc aacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt 5221 gccctggccc accctcgtga ccaccttcgg ctacggcgtg gcctgcttca gccgctaccc 5281 cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga 5341 gcgcaccatc tctttcaagg acgacggtac ctacaagacc cgcgccgagg tgaagttcga 5401 gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa 5461 catcctgggg cacaagctgg agtacaactt caacagccac aacgtctata tcacggccga 5521 caagcagaag aacggcatca aggctaactt caagatccgc cacaacgttg aggacggcag 5581 cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct 5641 gcccgacaac cactacctga gccatcagtc cgccctgagc aaagacccca acgagaagcg 5701 cgatcacatg gtcctgctgg agttcgtgac cgccgcctaa gcggccgcac tcgagatatc 5761 tagacccagc tttcttgtac aaagtggttg atatccagca cagtggcggc cgctcgacaa 5821 tcaacctctg gattacaaaa tttgtgaaag attgactggt attcttaact atgttgctcc 5881 ttttacgcta tgtggatacg ctgctttaat gcctttgtat catgctattg cttcccgtat 5941 ggctttcatt ttctcctcct tgtataaatc ctggttgctg tctctttatg aggagttgtg 6001 gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg 6061 ttggggcatt gccaccacct gtcagctcct ttccgggact ttcgctttcc ccctccctat 6121 tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt 6181 gggcactgac aattccgtgg tgttgtcggg gaagctgacg tcctttccat ggctgctcgc 6241 ctgtgttgcc acctggattc tgcgcgggac gtccttctgc tacgtccctt cggccctcaa 6301 tccagcggac cttccttccc gcggcctgct gccggctctg cggcctcttc cgcgtcttcg 6361 ccttcgccct cagacgagtc ggatctccct ttgggccgcc tccccgcctg gaattctacc 6421 gggtagggga ggcgcttttc ccaaggcagt ctggagcatg cgctttagca gccccgctgg 6481 gcacttggcg ctacacaagt ggcctctggc ctcgcacaca ttccacatcc accggtaggc 6541 gccaaccggc tccgttcttt ggtggcccct tcgcgccacc ttctactcct cccctagtca 6601 ggaagttccc ccccgccccg cagctcgcgt cgtgcaggac gtgacaaatg gaagtagcac 6661 gtctcactag tctcgtgcag atggacagca ccgctgagca atggaagcgg gtaggccttt 6721 ggggcagcgg ccaatagcag ctttgctcct tcgctttctg ggctcagagg ctgggaaggg 6781 gtgggtccgg gggcgggctc aggggcgggg cgggcgcccg aaggtcctcc ggaggcccgg 6841 cattctgcac gcttcaaaag cgcacgtctg ccgcgctgtt ctcctcttcc tcatctccgg 6901 gcctttcgac ctgcagccca agcttaccat gaccgagtac aagcccacgg tgcgcctcgc 6961 cacccgcgac gacgtcccca gggccgtacg caccctcgcc gccgcgttcg ccgactaccc 7021 cgccacgcgc cacaccgtcg atccggaccg ccacatcgag cgggtcaccg agctgcaaga 7081 actcttcctc acgcgcgtcg ggctcgacat cggcaaggtg tgggtcgcgg acgacggcgc 7141 cgcggtggcg gtctggacca cgccggagag cgtcgaagcg ggggcggtgt tcgccgagat 7201 cggcccgcgc atggccgagt tgagcggttc ccggctggcc gcgcagcaac agatggaagg 7261 cctcctggcg ccgcaccggc ccaaggagcc cgcgtggttc ctggccaccg tcggcgtctc 7321 gcccgaccac cagggcaagg gtctgggcag cgccgtcgtg ctccccggag tggaggcggc 7381 cgagcgcgcc ggggtgcccg ccttcctgga gacctccgcg ccccgcaacc tccccttcta 7441 cgagcggctc ggcttcaccg tcaccgccga cgtcgaggtg cccgaaggac cgcgcacctg 7501 gtgcatgacc cgcaagcccg gtgcctgacg cccgccccac gacccgcagc gcccgaccga 7561 aaggagcgca cgaccccatg catctcgagg gcccggtacc tttaagacca atgacttaca 7621 aggcagctgt agatcttagc cactttttaa aagaaaaggg gggactggaa gggctagctc 7681 actcccaacg aagacaagat ctgctttttg cttgtactgg gtctctctgg ttagaccaga 7741 tctgagcctg ggagctctct ggctaactag ggaacccact gcttaagcct caataaagct 7801 tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt aactagagat 7861 ccctcagacc cttttagtca gtgtggaaaa tctctagcag tagtagttca tgtcatctta 7921 ttattcagta tttataactt gcaaagaaat gaatatcaga gagtgagagg aacttgttta 7981 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat 8041 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct 8101 ggctctagct atcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc 8161 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc 8221 ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag ggacgtaccc 8281 aattcgccct atagtgagtc gtattacgcg cgctcactgg ccgtcgtttt acaacgtcgt 8341 gactgggaaa accctggcgt tacccaactt aatcgccttg cagcacatcc ccctttcgcc 8401 agctggcgta atagcgaaga ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg 8461 aatggcgaat gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg 8521 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct 8581 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta 8641 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt 8701 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg 8761 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat 8821 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt 8881 taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc ttacaatt //