LOCUS       sequence-376534-        6373 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Test effect of extending -Eco1 a1/a2 in yeast.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6373)
  AUTHORS   Lopez SC, Crawford KD, Lear SK, Bhattarai-Kline S, Shipman SL
  TITLE     Precise genome editing across kingdoms of life using retron-derived 
            DNA.
  JOURNAL   Nat Chem Biol. 2022 Feb;18(2):199-206. doi: 
            10.1038/s41589-021-00927-y. Epub 2021 Dec 23.
  PUBMED    34949838
REFERENCE   2  (bases 1 to 6373)
  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/s41589-021-00927-y"; journalName: "Nat Chem Biol"; date: 
            "2022-02"; volume: "18"; issue: "2"; pages: "199-206"
COMMENT     Sequence Label: pSCL.028
FEATURES             Location/Qualifiers
     source          1..6373
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        127..231
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(232..300,301..1092)
                     /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(450..469)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1263..1851
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1752..1771
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(3907..3926)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3908..3926)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3936..3953)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3936..3952)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3945..3967)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4093..4548
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4180..4199)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4390..4411
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(4679..5482)
                     /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"
     promoter        complement(5483..5698)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(5722..5741)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5841..5863
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5901..5919)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    join(5960..6373,1..90)
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
ORIGIN
        1 ttttctttaa tttctttttt tactttctat ttttaattta tatatttata ttaaaaaatt
       61 taaattataa ttatttttat agcacgtgat gaaaaggacc caggtggcac ttttcgggga
      121 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
      181 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt
      241 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct
      301 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt
      361 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt
      421 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac
      481 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac
      541 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct
      601 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg
      661 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg
      721 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca
      781 atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa
      841 caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt
      901 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc
      961 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg
     1021 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt
     1081 aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt
     1141 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc
     1201 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     1261 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     1321 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     1381 ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac
     1441 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     1501 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     1561 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     1621 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     1681 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
     1741 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
     1801 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaaggccagc
     1861 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctggcct tttgctcaag
     1921 tagagggggt aatttttccc ctttattttg ttcatacatt cttaaattgc tttgcctctc
     1981 cttttggaaa gctatacttc ggagcactgt tgagcgaagg ctcattagat atattttctg
     2041 tcattttcct taacccaaaa ataagggaaa gggtccaaaa agcgctcgga caactgttga
     2101 ccgtgatccg aaggactggc tatacagtgt tcacaaaata gccaagctga aaataatgtg
     2161 tagctatgtt cagttagttt ggctagcaaa gatataaaag caggtcggaa atatttatgg
     2221 gcattattat gcagagcatc aacatgataa aaaaaaacag ttgaatattc cctcaaaacg
     2281 ccacatgaaa tctgcagagt atctgaatac gttccgcctt aggaatttgg gcctccccgt
     2341 gatgaacaat ctccacgata tgagcaaggc gactcgaata tccgtggaaa cgctgagact
     2401 gctcatctat acagcagact ttcggtacag gatctacacg gtcgaaaaga aggggcctga
     2461 gaaacgcatg cgaacaattt atcaacctag ccgagagctc aaggcgttgc agggctgggt
     2521 tcttcgaaac atccttgaca aactctcatc atcacccttt agtattgggt ttgaaaagca
     2581 ccaaagcatc cttaacaacg cgacgccaca cataggtgcc aatttcatat tgaacatcga
     2641 cttggaggat ttttttccga gcctcacagc caataaagtg ttcggtgttt ttcacagtct
     2701 tgggtacaat cgccttatta gttccgttct taccaagatt tgttgttaca agaatctctt
     2761 gccccaggga gcacccagca gtccgaaatt ggcgaatttg atttgttcca agctcgatta
     2821 tcgaatacaa gggtacgcgg gcagccgggg actcatctat acccgctacg cagacgatct
     2881 tacgctgtct gcccaatcaa tgaagaaggt cgtaaaggcg cgggatttct tgttttctat
     2941 catcccgtcc gagggcttgg taattaattc caaaaagact tgtatctcag gaccacgatc
     3001 tcagcgaaaa gtgacaggac tcgtcatttc tcaagaaaaa gtcggtatag ggagagagaa
     3061 gtataaggaa atccgcgcga agatccacca catattctgt ggcaagagca gcgagataga
     3121 acacgtccga ggctggttgt ccttcatact gagcgtggac tcaaaaagcc accgccggtt
     3181 gatcacctat atttcaaaac tggaaaagaa atatggaaag aacccactca acaaagctaa
     3241 aacatagacc ggttagcgtg aaatgtcttt ggatttggga atcttataag ttctgtatga
     3301 gaccaccatt taactatgat aagattccgt atgcgcaccc ttagcgagag gtttatcatt
     3361 aaggtcaacc tctggatgtt gtttcggcat cctgcattga atctgagtta ctgtctgttt
     3421 tccttgttgg aacggagagc atcgcctgat gctctccgag ccaaccagga aacccgttat
     3481 ttctgacgta agggtgcgca tacggaatct tatcaaccgg ttagtaatga gtttaaacaa
     3541 agaaagtgga atattcattc atatcatatt ttttctatta actgcctggt ttcttttaaa
     3601 ttttttattg gttgtcgact tgaacggagt gacaatatat atatatatat ttaataatga
     3661 catcattatc tgtaaatctg attcttaatg ctattctagt tatgtaagag tggtcctttc
     3721 cataaaaaaa aaaaaaagaa aaaagaattt taggaataca atgcagcttg taagtaaaat
     3781 ctggaatatt catatcgcca caacttctta tgcttataaa agcactaatg cctgaattta
     3841 tgttgaaaat atgtgtcaca aataaagaaa ctgtgacatc tgacacattt ccacgtaccc
     3901 aattcgccct atagtgagtc gtattacgcg cgctcactgg ccgtcgtttt acaacgtcgt
     3961 gactgggaaa accctggcgt tacccaactt aatcgccttg cagcacatcc ccctttcgcc
     4021 agctggcgta atagcgaaga ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg
     4081 aatggcgaat ggacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
     4141 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt
     4201 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag
     4261 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt
     4321 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
     4381 tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt
     4441 cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt
     4501 aacaaaaatt taacgcgaat tttaacaaaa tattaacgct tacaatttcc tgatgcggta
     4561 ttttctcctt acgcatctgt gcggtatttc acaccgcata gggtaataac tgatataatt
     4621 aaattgaagc tctaatttgt gagtttagta tacatgcatt tacttataat acagtttttt
     4681 agttttgctg gccgcatctt ctcaaatatg cttcccagcc tgcttttctg taacgttcac
     4741 cctctacctt agcatccctt ccctttgcaa atagtcctct tccaacaata ataatgtcag
     4801 atcctgtaga gaccacatca tccacggttc tatactgttg acccaatgcg tctcccttgt
     4861 catctaaacc cacaccgggt gtcataatca accaatcgta accttcatct cttccaccca
     4921 tgtctctttg agcaataaag ccgataacaa aatctttgtc gctcttcgca atgtcaacag
     4981 tacccttagt atattctcca gtagataggg agcccttgca tgacaattct gctaacatca
     5041 aaaggcctct aggttccttt gttacttctt ctgccgcctg cttcaaaccg ctaacaatac
     5101 ctgggcccac cacaccgtgt gcattcgtaa tgtctgccca ttctgctatt ctgtatacac
     5161 ccgcagagta ctgcaatttg actgtattac caatgtcagc aaattttctg tcttcgaaga
     5221 gtaaaaaatt gtacttggcg gataatgcct ttagcggctt aactgtgccc tccatggaaa
     5281 aatcagtcaa gatatccaca tgtgttttta gtaaacaaat tttgggacct aatgcttcaa
     5341 ctaactccag taattccttg gtggtacgaa catccaatga agcacacaag tttgtttgct
     5401 tttcgtgcat gatattaaat agcttggcag caacaggact aggatgagta gcagcacgtt
     5461 ccttatatgt agctttcgac atgatttatc ttcgtttcct gcaggttttt gttctgtgca
     5521 gttgggttaa gaatactggg caatttcatg tttcttcaac actacatatg cgtatatata
     5581 ccaatctaag tctgtgctcc ttccttcgtt cttccttctg ttcggagatt accgaatcaa
     5641 aaaaatttca aggaaaccga aatcaaaaaa aagaataaaa aaaaaatgat gaattgaaaa
     5701 ggtggtatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagccccg
     5761 acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta
     5821 cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc
     5881 gaaacgcgcg agacgaaagg gcctcgtgat acgcctattt ttataggtta atgtcatgat
     5941 aataatggtt tcttagacgg atcgcttgcc tgtaacttac acgcgcctcg tatcttttaa
     6001 tgatggaata atttgggaat ttactctgtg tttatttatt tttatgtttt gtatttggat
     6061 tttagaaagt aaataaagaa ggtagaagag ttacggaatg aagaaaaaaa aataaacaaa
     6121 ggtttaaaaa atttcaacaa aaagcgtact ttacatatat atttattaga caagaaaagc
     6181 agattaaata gatatacatt cgattaacga taagtaaaat gtaaaatcac aggattttcg
     6241 tgtgtggtct tctacacaga caagatgaaa caattcggca ttaatacctg agagcaggaa
     6301 gagcaagata aaaggtagta tttgttggcg atccccctag agtcttttac atcttcggaa
     6361 aacaaaaact att
//
