LOCUS Exported 7305 bp ds-DNA circular SYN 01-JAN-2020 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS . SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7305) AUTHORS . TITLE Direct Submission JOURNAL Exported Tuesday, Nov 24, 2020 from SnapGene Viewer 5.0.8 https://www.snapgene.com FEATURES Location/Qualifiers source 1..7305 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind 142..158 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" misc_feature 202..1081 /label=sali arm primer_bind complement(1064..1101) /label=Primer 1 primer_bind complement(1064..1087) /label=Primer 2 Bleo 3' CDS 1088..1879 /codon_start=1 /gene="aph(3')-II (or nptII)" /product="aminoglycoside phosphotransferase from Tn5" /label=NeoR/KanR /note="confers resistance to neomycin, kanamycin, and G418 (Geneticin(R))" /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(1859..1900) /label=Neo 3' int CDS 1889..1945 /codon_start=1 /product="2A peptide from porcine teschovirus-1 polyprotein" /label=P2A /note="Eukaryotic ribosomes fail to insert a peptide bond between the Gly and Pro residues, yielding separate polypeptides." /translation="ATNFSLLKQAGDVEENPGP" CDS 1946..1972 /codon_start=1 /product="HA (human influenza hemagglutinin) epitope tag" /label=HA /translation="YPYDVPDYA" CDS 1976..2002 /codon_start=1 /product="HA (human influenza hemagglutinin) epitope tag" /label=HA /translation="YPYDVPDYA" CDS 2012..2332 /codon_start=1 /product="F36V mutant of FK506-binding protein FKBP12" /label=FKBP-V /note="binds synthetic ligands that do not bind wild-type FKBP" /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKVDSSRDRNKP FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL KLE" CDS 2363..3058 /codon_start=1 /product="bright monomeric red fluorescent protein evolved from a synthetic template (Bindels et al., 2016)" /label=mScarlet-I /note="matures faster than mScarlet" /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA KLKVTKGGPLPFSWDILSPQFMYGSRAFIKHPADIPDYYKQSFPEGFKWERVMNFEDGG AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH STGGMDELYK" primer_bind 2902..2922 /label=mcherry int5' seq primer_bind 3090..3127 /label=Primer 2 misc_feature 3110..4717 /label=kpni arm CDS 3110..3170 /codon_start=1 /translation="NGDMPHVPITTLAGIASLTD" promoter complement(4808..4826) /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" primer_bind complement(4847..4863) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" protein_bind 4871..4887 /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(4895..4925) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 4940..4961 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." rep_origin complement(5249..5837) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" CDS complement(6008..6868) /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" promoter complement(6869..6973) /gene="bla" /label=AmpR promoter ORIGIN 1 gccattcgcc attcaggctg cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc 61 tattacgcca gctggcgaaa gggggatgtg ctgcaaggcg attaagttgg gtaacgccag 121 ggttttccca gtcacgacgt tgtaaaacga cggccagtgc cactagtggc cgcggcggcc 181 gcagcttgcc tgcaggtcga ccagactctt gtgctgaggg aaagatccta gttccgcaga 241 ttatcttctg atctgcacac acatcagtca tagcaggcac acaacaatac acgtggacac 301 taagtgaaca aatagtaaca aattacaaat ggggcagggg aggaggaggg acatatcaag 361 tgtgtcacaa aggaatatac agcaatttac tgagaaaata ttgggcaata aatgtcataa 421 attcagttat cctaaggtaa aatggctgtg gaatgaaagt gttcttgaat tgtaggattt 481 agaagagagg agaataaaag tctcacactg caaaccacaa ggatccaaaa gaatctgaag 541 agatactctt ggtaggaggg gcaagtccat ttagagagta aaaagggaac tagacaagaa 601 attgaagcca gattattaag taaaaatttt attgtattta gagtttagaa agagtaccct 661 gccctagcag tatatacacc agaagttaag aaacaagatt ctggagctag attgtgtcat 721 tagaattaca gctccagtag ttactttaaa ctcactgtaa ttttctcatt agtaaaatgc 781 agaaaagagt aatactctgc tgtacctggg ttgttgtgag acctaaatct taatgtatac 841 aaagcatttt gcagtgttta atgtatggca catgcttgtt aaacacattg aacaaataga 901 agctaagagt tttgtttcta aatgaataag tatctctctg ctactcatat atttgacaaa 961 gttctacaaa taattatatg gttttctttg tgtgtgttgc agagtttggg aaatggaaag 1021 taatgacagc tggcacctga actaaatact ttcatagaca acaccattcc agaacttcag 1081 ggtcgacatg attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag 1141 gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg 1201 gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa 1261 tgaactgcag gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc 1321 agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc 1381 ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga 1441 tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa 1501 acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct 1561 ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgcgcat 1621 gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt 1681 ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta 1741 tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga 1801 ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg 1861 ccttcttgac gagttcttcg gttccggagc tactaacttc agcctgctga agcaggctgg 1921 agacgtggag gagaaccctg gaccttaccc ctacgacgtg cccgactacg ccggctaccc 1981 atacgatgtt ccagattacg ctggaagcgg aggagtgcag gtggaaacca tctccccagg 2041 agacgggcgc accttcccca agcgcggcca gacctgcgtg gtgcactaca ccgggatgct 2101 tgaagatgga aagaaagttg attcctcccg ggacagaaac aagcccttta agtttatgct 2161 aggcaagcag gaggtgatcc gaggctggga agaaggggtt gcccagatga gtgtgggtca 2221 gagagccaaa ctgactatat ctccagatta tgcctatggt gccactgggc acccaggcat 2281 catcccacca catgccactc tcgtcttcga tgtggagctt ctaaaactgg aaggtggagg 2341 cgggagtgga ggtggaggat cgatggtgag caagggcgag gcagtgatca aggagttcat 2401 gcggttcaag gtgcacatgg agggctccat gaacggccac gagttcgaga tcgagggcga 2461 gggcgagggc cgcccctacg agggcaccca gaccgccaag ctgaaggtga ccaagggtgg 2521 ccccctgccc ttctcctggg acatcctgtc ccctcagttc atgtacggct ccagggcctt 2581 catcaagcac cccgccgaca tccccgacta ctataagcag tccttccccg agggcttcaa 2641 gtgggagcgc gtgatgaact tcgaggacgg cggcgccgtg accgtgaccc aggacacctc 2701 cctggaggac ggcaccctga tctacaaggt gaagctccgc ggcaccaact tccctcctga 2761 cggccccgta atgcagaaga agacaatggg ctgggaagcg tccaccgagc ggttgtaccc 2821 cgaggacggc gtgctgaagg gcgacattaa gatggccctg cgcctgaagg acggcggccg 2881 ctacctggcg gacttcaaga ccacctacaa ggccaagaag cccgtgcaga tgcccggcgc 2941 ctacaacgtg gaccgcaagt tggacatcac ctcccacaac gaggactaca ccgtggtgga 3001 acagtacgaa cgctccgagg gccgccactc caccggcggc atggacgagc tgtacaaggg 3061 tggcggtggc tcgggcggtg gtgggtcggg tggcggcgga tctggtacca atggggatat 3121 gccccatgtc cccataacta cgcttgcggg gattgctagt cttacggacc gtaagtttgc 3181 ataattttcc taagttcttt actttgctaa tactcatttt tagagacaaa aaattacctt 3241 taatgtcaca tataacagaa aatatgtatt tgattagtgt ggggggagta gcaaaattaa 3301 tgtttccacc taatacataa atttaatttc ttcatccttc tgtgaactcc taaagaaatg 3361 atagtgtgat aatttttata aagaccttaa aaatcaataa ggactttaga aaaatgacac 3421 aaagctttaa gagatgttaa aacttaaata aacatgggtt aataaacaaa actatggtct 3481 tttttttaag aaagcgttta caaaaacagt acctgaaagt gtcctccaag aatgagaaat 3541 tgatcgccca cattacctaa actgtagtgg tcctgatttt tttaatatca ggttgaattg 3601 ttagttagaa tttgttaaca ggtatgagat atgtcatcac tatgagagcc cactttactc 3661 tgaacactaa ggtattaaaa attattgtca atacttagaa atttagttaa tagaagtttt 3721 aaggttcata ttttggtaga cacataacta ttcttttcct tgggtaatgg cacagagtct 3781 gcccaagtgt ttgtccctat gtcatcctcc ttttagtata ggaatagaga taaaggaaag 3841 gccagaaaag aaaaaacaaa atgaaaacat aaataccttt ttcttatagc agtatgtaag 3901 atatggaact ttgcaataat ctatgaaaaa ataacttcca gaactgtttt caactgtgct 3961 cagaatccct ccctgggttt tgtttttgtg tattttcagt cgagtgtaag acacttgtat 4021 cactagaaag taggtggttg gattatccta aatctgcagg gaaactttga aacatgtttg 4081 tttttgttac tcgttttaag gcaaaaacac aacctgtatg gatggaaaat tatatatgct 4141 gctagacttc cctttatctt ttctgccttt tatctctgat atacctcaca tttttgatgc 4201 ttatcttcct ggtaaactct gtgaagtatg gtaattatac tatggggtcg actagaaatt 4261 aaccccaaaa catgatacct atattttagt tgggtttctt aaccttttgg caagtaaaca 4321 attcctatta agtctactta cattaaatag tgggattgaa gccacaaaat actcatttgt 4381 ttgtttgttt gtttgtttgc tgcttagagt agaatgggag aaaaaagaga actatttagg 4441 aatttatgac aagaattttg tattgttctt caggaatgtt gtactctgag tattccacta 4501 ctgtgagcta tttgtttcta tattcttaag cttttaaaat tgtattgcat atcagtataa 4561 tgtgagtcag gtattgtttt tgacttatat gtctaagagt gctactgatc ttatagataa 4621 gctactttct caccttatgt gtcaaagtca tgataataat aataaagttt tatttagtta 4681 attaaaacaa tttttaagca tgaatataaa tcagctaggt accgggcccc ccctcgaggt 4741 cgatcgacgg tatcgataag cttgatctag agcggccgcc accgcggtgg agctccagct 4801 tttgttccct ttagtgaggg ttaattccga gcttggcgta atcatggtca tagctgtttc 4861 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt 4921 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc 4981 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg 5041 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct 5101 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca 5161 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga 5221 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc 5281 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg 5341 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat 5401 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt 5461 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc 5521 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg 5581 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg 5641 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg 5701 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg 5761 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca 5821 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga 5881 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga 5941 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt 6001 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt 6061 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat 6121 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag 6181 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct 6241 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt 6301 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg 6361 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca 6421 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt 6481 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat 6541 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac 6601 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa 6661 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt 6721 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt 6781 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa 6841 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt 6901 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa 6961 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtctaagaa accattatta 7021 tcatgacatt aacctataaa aataggcgta tcacgaggcc ctttcgtctc gcgcgtttcg 7081 gtgatgacgg tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt 7141 aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc 7201 ggggctggct taactatgcg gcatcagagc agattgtact gagagtgcac catatgcggt 7261 gtgaaatacc gcacagatgc gtaaggagaa aataccgcat caggc //