LOCUS Exported 5865 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION low-copy URA-selectable plasmid, 1xFLAG-MCP under TEF1 promoter. ACCESSION . VERSION . KEYWORDS pRS416-PTEF1-MCP-no_intron SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5865) AUTHORS Morgan JT, Fink GR, Bartel DP TITLE Excised linear introns regulate growth in yeast. JOURNAL Nature. 2019 Jan;565(7741):606-611. doi: 10.1038/s41586-018-0828-1. Epub 2019 Jan 16. PUBMED 30651636 REFERENCE 2 (bases 1 to 5865) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5865 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind 51..73 /label=M13/pUC Forward /note="In lacZ gene" primer_bind 65..82 /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind 66..82 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind 92..111 /label=T7 /note="T7 promoter, forward primer" promoter 92..110 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" promoter 124..522 /gene="S. cerevisiae TEF1" /label=TEF1 promoter /note="promoter for EF-1-alpha" CDS 541..564 /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" primer_bind complement(967..985) /label=CYC1 /note="CYC1 transcription termination signal, reverse primer" terminator 997..1186 /gene="S. cerevisiae CYC1" /label=CYC1 terminator /note="transcription terminator for CYC1" primer_bind complement(1211..1231) /label=T3 /note="T3 promoter, forward primer" promoter complement(1211..1229) /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" primer_bind complement(1250..1266) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(1250..1266) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(1263..1285) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 1274..1290 /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(1298..1328) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 1343..1364 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(1481..1498) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(1652..2240) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(1732..1751) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(2411..3271) /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 3034..3053 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(3272..3376) /gene="bla" /label=AmpR promoter misc_feature 3413..3916 /label=CEN/ARS /note="S. cerevisiae CEN6 centromere fused to an autonomously replicating sequence" primer_bind 3957..3975 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" primer_bind complement(4017..4039) /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind 4142..4161 /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" promoter 4185..4400 /gene="S. cerevisiae URA3" /label=URA3 promoter CDS 4401..5204 /codon_start=1 /gene="S. cerevisiae URA3" /product="orotidine-5'-phosphate decarboxylase, required for uracil biosynthesis" /label=URA3 /note="yeast auxotrophic marker, counterselectable with 5-fluoroorotic acid (5-FOA)" /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN" rep_origin complement(5335..5790) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(5472..5493) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 5684..5703 /label=F1ori-R /note="F1 origin, reverse primer" ORIGIN 1 gaaaggggga tgtgctgcaa ggcgattaag ttgggtaacg ccagggtttt cccagtcacg 61 acgttgtaaa acgacggcca gtgagcgcgc gtaatacgac tcactatagg gcgaattggg 121 taccatagct tcaaaatgtt tctactcctt ttttactctt ccagattttc tcggactccg 181 cgcatcgccg taccacttca aaacacccaa gcacagcata ctaaatttcc cctctttctt 241 cctctagggt gtcgttaatt acccgtacta aaggtttgga aaagaaaaaa gagaccgcct 301 cgtttctttt tcttcgtcga aaaaggcaat aaaaattttt atcacgtttc tttttcttga 361 aaattttttt tttgattttt ttctctttcg atgacctccc attgatattt aagttaataa 421 acggtcttca atttctcaag tttcagtttc atttttcttg ttctattaca acttttttta 481 cttcttgctc attagaaaga aagcatagca atctaatcta agttttctag aactagtatg 541 gactacaagg acgacgatga caagatggct tctaacttta ctcagttcgt tctcgtcgac 601 aatggcggaa ctggcgacgt gactgtcgcc ccaagcaact tcgctaacgg ggtcgctgaa 661 tggatcagct ctaactcgcg atcacaggct tacaaagtaa cctgtagcgt tcgtcagagc 721 tctgcgcaga atcgcaaata caccatcaaa gtcgaggtgc ctaaagtggc aacccagact 781 gttggtggtg aagagcttcc tgtagccgga tggagatctt acttaaatat ggaactaacc 841 attccaattt tcgccacgaa ttccgactgc gagcttattg ttaaggcaat gcaaggtctc 901 ctaaaagatg gaaacccgat tccctcggcc atcgcggcca actccggcat ctactagatt 961 agttatgtca cgcttacatt cacgccctcc ccccacatcc gctctaaccg aaaaggaagg 1021 agttagacaa cctgaagtct aggtccctat ttattttttt atagttatgt tagtattaag 1081 aacgttattt atatttcaaa tttttctttt ttttctgtac agacgcgtgt acgcatgtaa 1141 cattatactg aaaaccttgc ttgagaaggt tttgggacgc tcgaaggctt taatttgcca 1201 gcttttgttc cctttagtga gggttaattg cgcgcttggc gtaatcatgg tcatagctgt 1261 ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa 1321 agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac 1381 tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg 1441 cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc 1501 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat 1561 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca 1621 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc 1681 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc 1741 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg 1801 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta 1861 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg 1921 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac 1981 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag 2041 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat 2101 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat 2161 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc 2221 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt 2281 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct 2341 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt 2401 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc 2461 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac 2521 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat 2581 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg 2641 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata 2701 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta 2761 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt 2821 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag 2881 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa 2941 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc 3001 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt 3061 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc 3121 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta 3181 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa 3241 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca 3301 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac 3361 aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgggtccttt tcatcacgtg 3421 ctataaaaat aattataatt taaatttttt aatataaata tataaattaa aaatagaaag 3481 taaaaaaaga aattaaagaa aaaatagttt ttgttttccg aagatgtaaa agactctagg 3541 gggatcgcca acaaatacta ccttttatct tgctcttcct gctctcaggt attaatgccg 3601 aattgtttca tcttgtctgt gtagaagacc acacacgaaa atcctgtgat tttacatttt 3661 acttatcgtt aatcgaatgt atatctattt aatctgcttt tcttgtctaa taaatatata 3721 tgtaaagtac gctttttgtt gaaatttttt aaacctttgt ttattttttt ttcttcattc 3781 cgtaactctt ctaccttctt tatttacttt ctaaaatcca aatacaaaac ataaaaataa 3841 ataaacacag agtaaattcc caaattattc catcattaaa agatacgagg cgcgtgtaag 3901 ttacaggcaa gcgatccgtc taagaaacca ttattatcat gacattaacc tataaaaata 3961 ggcgtatcac gaggcccttt cgtcgaattc gcgcgtttcg gtgatgacgg tgaaaacctc 4021 tgacacatgc agctcccgga gatctcggtc acagcttgtc tgtaagcgga tgccgggagc 4081 agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggctg gcttaactat 4141 gcggcatcag agcagattgt accgagagtg caccatacca ccttttcaat tcatcatttt 4201 ttttttattc ttttttttga tttcggtttc cttgaaattt ttttgattcg gtaatctccg 4261 aacagaagga agaacgaagg aaggagcaca gacttagatt ggtatatata cgcatatgta 4321 gtgttgaaga aacatgaaat tgcccagtat tcttaaccca actgcacaga acaaaaacct 4381 gcaggaaacg aagataaatc atgtcgaaag ctacatataa ggaacgtgct gctactcatc 4441 ctagtcctgt tgctgccaag ctatttaata tcatgcacga aaagcaaaca aacttgtgtg 4501 cttcattgga tgttcgtacc accaaggaat tactggagtt agttgaagca ttaggtccca 4561 aaatttgttt actaaaaaca catgtggata tcttgactga tttttccatg gagggcacag 4621 ttaagccgct aaaggcatta tccgccaagt acaatttttt actcttcgaa gacagaaaat 4681 ttgctgacat tggtaataca gtcaaattgc agtactctgc gggtgtatac agaatagcag 4741 aatgggcaga cattacgaat gcacacggtg tggtgggccc aggtattgtt agcggtttga 4801 agcaggcggc agaagaagta acaaaggaac ctagaggcct tttgatgtta gcagaattgt 4861 catgcaaggg ctccctatct actggagaat atactaaggg tactgttgac attgcgaaga 4921 gcgacaaaga ttttgttatc ggctttattg ctcaaagaga catgggtgga agagatgaag 4981 gttacgattg gttgattatg acacccggtg tgggtttaga tgacaagggt gatgcattgg 5041 gtcaacagta tagaaccgtg gatgatgtgg tctctacagg atctgacatt attattgttg 5101 gaagaggact atttgcaaag ggaagggatg ctaaggtaga gggtgaacgt tacagaaaag 5161 caggctggga agcatatttg agaagatgcg gccagcaaaa ctaaaaaact gtattataag 5221 taaatgcatg tatactaaac tcacaaatta gagcttcaat ttaattatat cagttattac 5281 cctatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat caggaaattg 5341 taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta 5401 accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt 5461 tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca 5521 aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa 5581 gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat 5641 ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag 5701 gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg 5761 ccgcgcttaa tgcgccgcta cagggcgcgt ccattcgcca ttcaggctgc gcaactgttg 5821 ggaagggcga tcggtgcggg cctcttcgct attacgccag ctggc //