LOCUS Exported 5471 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS pTK21 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5471) AUTHORS Kiyomitsu T, Cheeseman IM TITLE Chromosome- and spindle-pole-derived signals generate an intrinsic code for spindle position and orientation. JOURNAL Nat Cell Biol. 2012 Feb 12;14(3):311-7. doi: 10.1038/ncb2440. PUBMED 22327364 REFERENCE 2 (bases 1 to 5471) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5471 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 61..364 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 365..568 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 519..539 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" regulatory 607..616 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 613..1320 /codon_start=1 /product="monomeric derivative of DsRed fluorescent protein (Shaner et al., 2004)" /label=mCherry /note="mammalian codon-optimized" /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA EGRHSTGGMDELYK" primer_bind complement(758..776) /label=mCherry-R /note="mCherry, reverse primer" primer_bind 1026..1045 /label=mCherry-F /note="mCherry, forward primer" CDS 1366..1386 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQG" CDS 1399..1443 /codon_start=1 /product="affinity and epitope tag derived from pancreatic ribonuclease A" /label=S-Tag /translation="KETAAAKFERQHMDS" polyA_signal 2259..2380 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2296..2315) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 2350..2369 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" rep_origin complement(2387..2842) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2524..2545) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 2736..2755 /label=F1ori-R /note="F1 origin, reverse primer" promoter 2869..2973 /gene="bla" /label=AmpR promoter promoter 2975..3332 /label=SV40 promoter /note="SV40 enhancer and early promoter" primer_bind complement(2998..3018) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" rep_origin 3183..3318 /label=SV40 ori /note="SV40 origin of replication" primer_bind 3245..3264 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 3367..4161 /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 GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(3421..3440) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 4031..4050 /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind complement(4349..4368) /label=TK-pA-R /note="Thymidine kinase polyA, reverse primer" polyA_signal 4393..4440 /label=HSV TK poly(A) signal /note="herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)" rep_origin 4769..5357 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 5258..5277 /label=pBR322ori-F /note="pBR322 origin, forward primer" ORIGIN 1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg 61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt 121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca 181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc 241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta 301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac 361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg 421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg 481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt 541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta 601 ccggtcgcca ccatggtgag caagggcgag gaggataaca tggccatcat caaggagttc 661 atgcgcttca aggtgcacat ggagggctcc gtgaacggcc acgagttcga gatcgagggc 721 gagggcgagg gccgccccta cgagggcacc cagaccgcca agctgaaggt gaccaagggt 781 ggccccctgc ccttcgcctg ggacatcctg tcccctcagt tcatgtacgg ctccaaggcc 841 tacgtgaagc accccgccga catccccgac tacttgaagc tgtccttccc cgagggcttc 901 aagtgggagc gcgtgatgaa cttcgaggac ggcggcgtgg tgaccgtgac ccaggactcc 961 tccctgcagg acggcgagtt catctacaag gtgaagctgc gcggcaccaa cttcccctcc 1021 gacggccccg taatgcagaa gaagaccatg ggctgggagg cctcctccga gcggatgtac 1081 cccgaggacg gcgccctgaa gggcgagatc aagcagaggc tgaagctgaa ggacggcggc 1141 cactacgacg ctgaggtcaa gaccacctac aaggccaaga agcccgtgca gctgcccggc 1201 gcctacaacg tcaacatcaa gttggacatc acctcccaca acgaggacta caccatcgtg 1261 gaacagtacg aacgcgccga gggccgccac tccaccggcg gcatggacga gctgtacaag 1321 tccggagggg atcggtggag ctccaccggt ggcggccgct ctagagagaa tctttatttt 1381 cagggcgccg ccaaattcaa agaaaccgct gctgctaaat tcgaacgcca gcacatggac 1441 agcggaggtg gaggtactag tctcgagatg gctgcgcagg gagagcccca ggtccagttc 1501 aaacttgtat tggttggtga tggtggtact ggaaaaaaca ccttcgtgaa acgtcatttg 1561 actggtgaat ttgagaagaa gtatgtagcc accttgggtg ttgaggttca tcccctagtg 1621 ttccacacca acagaggacc tattaagttc aatgtatggg acacagccgg ccaggagaaa 1681 ttcggtggac tgagagatgg ctattatatc caagcccagt gtgccatcat aatgtttgat 1741 gtaacatcga gagttactta caagaatgtg cctaactggc atagagatct ggtacgagtg 1801 tgtgaaaaca tccccattgt gttgtgtggc aacaaagtgg atattaagga caggaaagtg 1861 aaggcgaaat ccattgtctt ccaccgaaag aagaatcttc agtactacga catttctgcc 1921 aaaagtaact acaactttga aaagcccttc ctctggcttg ctaggaagct cattggagac 1981 cctaacttgg aatttgttgc catgcctgct ctcgccccac cagaagttgt catggaccca 2041 gctttggcag cacagtatga gcacgactta gaggttgctc agacaactgc tctcccggat 2101 gaggatgatg acctgtgacc gcgggcccgg gatccaccgg atctagataa ctgatcataa 2161 tcagccatac cacatttgta gaggttttac ttgctttaaa aaacctccca cacctccccc 2221 tgaacctgaa acataaaatg aatgcaattg ttgttgttaa cttgtttatt gcagcttata 2281 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc 2341 attctagttg tggtttgtcc aaactcatca atgtatctta acgcgtaaat tgtaagcgtt 2401 aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag 2461 gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt 2521 gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga 2581 aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc aagttttttg 2641 gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct 2701 tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc 2761 gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt 2821 aatgcgccgc tacagggcgc gtcaggtggc acttttcggg gaaatgtgcg cggaacccct 2881 atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga 2941 taaatgcttc aataatattg aaaaaggaag agtcctgagg cggaaagaac cagctgtgga 3001 atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa 3061 gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca 3121 gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc 3181 ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt 3241 tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag 3301 gaggcttttt tggaggccta ggcttttgca aagatcgatc aagagacagg atgaggatcg 3361 tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg 3421 ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg 3481 ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat 3541 gaactgcaag acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca 3601 gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg 3661 gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat 3721 gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa 3781 catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg 3841 gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgagcatg 3901 cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg 3961 gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat 4021 caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac 4081 cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc 4141 cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac caagcgacgc 4201 ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg 4261 gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt 4321 tcttcgccca ccctaggggg aggctaactg aaacacggaa ggagacaata ccggaaggaa 4381 cccgcgctat gacggcaata aaaagacaga ataaaacgca cggtgttggg tcgtttgttc 4441 ataaacgcgg ggttcggtcc cagggctggc actctgtcga taccccaccg agaccccatt 4501 ggggccaata cgcccgcgtt tcttcctttt ccccacccca ccccccaagt tcgggtgaag 4561 gcccagggct cgcagccaac gtcggggcgg caggccctgc catagcctca ggttactcat 4621 atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc 4681 tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag 4741 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct 4801 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac 4861 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgtccttc 4921 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg 4981 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt 5041 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt 5101 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc 5161 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca 5221 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata 5281 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg 5341 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct 5401 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta 5461 ccgccatgca t //