LOCUS       sequence-273869-        5121 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  low-copy URA-selectable plasmid, ECM33 intron inserted into URA3.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5121)
  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 5121)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1038/s41586-018-0828-1"; journalName: "Nature"; date: "2019-01";
            volume: "565"; issue: "7741"; pages: "606-611"
COMMENT     Sequence Label: pRS416-ECM33_intron
FEATURES             Location/Qualifiers
     source          1..5121
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     415..433
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(475..497)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     600..619
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        643..858
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     rep_origin      complement(2123..2578)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2260..2281)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2472..2491
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     CDS             complement(2555..2833)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSAQLTLTKGNKSWVPNSPYSESYYARSLAVVLQRRDWEN
                     PGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWTRPVAAH"
     primer_bind     2704..2726
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2718..2735
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2719..2735
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2745..2764
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2745..2763
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2790..2810)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2790..2808)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2829..2845)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2829..2845)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2842..2864)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2853..2869
                     /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(join(2877..2883,2884..2901,2902..2907))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2922..2943
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3060..3077)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3231..3819)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3311..3330)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3990..4781,4782..4850))
                     /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     4613..4632
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4851..4955)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    join(4992..5121,1..374)
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
ORIGIN
        1 gatcgccaac aaatactacc ttttatcttg ctcttcctgc tctcaggtat taatgccgaa
       61 ttgtttcatc ttgtctgtgt agaagaccac acacgaaaat cctgtgattt tacattttac
      121 ttatcgttaa tcgaatgtat atctatttaa tctgcttttc ttgtctaata aatatatatg
      181 taaagtacgc tttttgttga aattttttaa acctttgttt attttttttt cttcattccg
      241 taactcttct accttcttta tttactttct aaaatccaaa tacaaaacat aaaaataaat
      301 aaacacagag taaattccca aattattcca tcattaaaag atacgaggcg cgtgtaagtt
      361 acaggcaagc gatccgtcta agaaaccatt attatcatga cattaaccta taaaaatagg
      421 cgtatcacga ggccctttcg tcgaattcgc gcgtttcggt gatgacggtg aaaacctctg
      481 acacatgcag ctcccggaga tctcggtcac agcttgtctg taagcggatg ccgggagcag
      541 acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt cggggctggc ttaactatgc
      601 ggcatcagag cagattgtac cgagagtgca ccataccacc ttttcaattc atcatttttt
      661 ttttattctt ttttttgatt tcggtttcct tgaaattttt ttgattcggt aatctccgaa
      721 cagaaggaag aacgaaggaa ggagcacaga cttagattgg tatatatacg catatgtagt
      781 gttgaagaaa catgaaattg cccagtattc ttaacccaac tgcacagaac aaaaacctgc
      841 aggaaacgaa gataaatcat gtcgaaagct acatataagg aacgtgctgc tactcatcct
      901 agtcctgttg gtatgtacac attctccttt tttttcatct ttttttcttt atttgcctct
      961 ttcctcacaa atctcgagta gattcgtggt cctcttcatt cttttctttt ttctttgtcg
     1021 attactgggc tttttttcat aggtctcgat tgacgcggac ggacaatgcg aaaaaaaaaa
     1081 atttccaaaa gaggaaacgg gtttcgagat ctcgtgagga tggttttgga atattcttca
     1141 atatttctgc ctgtcatagg attagggcga gtaatgaaac agaataatac atgtataaat
     1201 cgatcgggaa tactaacact acttttcttt atctaagcag ctgccaagct atttaatatc
     1261 atgcacgaaa agcaaacaaa cttgtgtgct tcattggatg ttcgtaccac caaggaatta
     1321 ctggagttag ttgaagcatt aggtcccaaa atttgtttac taaaaacaca tgtggatatc
     1381 ttgactgatt tttccatgga gggcacagtt aagccgctaa aggcattatc cgccaagtac
     1441 aattttttac tcttcgaaga cagaaaattt gctgacattg gtaatacagt caaattgcag
     1501 tactctgcgg gtgtatacag aatagcagaa tgggcagaca ttacgaatgc acacggtgtg
     1561 gtgggcccag gtattgttag cggtttgaag caggcggcag aagaagtaac aaaggaacct
     1621 agaggccttt tgatgttagc agaattgtca tgcaagggct ccctatctac tggagaatat
     1681 actaagggta ctgttgacat tgcgaagagc gacaaagatt ttgttatcgg ctttattgct
     1741 caaagagaca tgggtggaag agatgaaggt tacgattggt tgattatgac acccggtgtg
     1801 ggtttagatg acaagggtga tgcattgggt caacagtata gaaccgtgga tgatgtggtc
     1861 tctacaggat ctgacattat tattgttgga agaggactat ttgcaaaggg aagggatgct
     1921 aaggtagagg gtgaacgtta cagaaaagca ggctgggaag catatttgag aagatgcggc
     1981 cagcaaaact aaaaaactgt attataagta aatgcatgta tactaaactc acaaattaga
     2041 gcttcaattt aattatatca gttattaccc tatgcggtgt gaaataccgc acagatgcgt
     2101 aaggagaaaa taccgcatca ggaaattgta agcgttaata ttttgttaaa attcgcgtta
     2161 aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat
     2221 aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca
     2281 ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc
     2341 ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg taaagcacta
     2401 aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg
     2461 gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg
     2521 gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc
     2581 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
     2641 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
     2701 tttcccagtc acgacgttgt aaaacgacgg ccagtgagcg cgcgtaatac gactcactat
     2761 agggcgaatt gggtacccag cttttgttcc ctttagtgag ggttaattgc gcgcttggcg
     2821 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
     2881 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca
     2941 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     3001 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     3061 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     3121 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     3181 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     3241 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     3301 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     3361 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     3421 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     3481 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     3541 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     3601 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     3661 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     3721 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     3781 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     3841 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     3901 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     3961 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     4021 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     4081 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     4141 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     4201 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     4261 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     4321 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     4381 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     4441 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     4501 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     4561 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     4621 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     4681 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     4741 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     4801 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     4861 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     4921 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     4981 gggtcctttt catcacgtgc tataaaaata attataattt aaatttttta atataaatat
     5041 ataaattaaa aatagaaagt aaaaaaagaa attaaagaaa aaatagtttt tgttttccga
     5101 agatgtaaaa gactctaggg g
//
