LOCUS       sequence-376531-        5569 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Express -Eco1 ADE2 editing ncRNA and gRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5569)
  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 5569)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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.002
FEATURES             Location/Qualifiers
     source          1..5569
                     /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(3103..3122)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3104..3122)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3132..3149)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3132..3148)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3141..3163)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3289..3744
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3376..3395)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3586..3607
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3875..4678)
                     /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(4679..4894)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(4918..4937)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5037..5059
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5097..5115)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    join(5156..5569,1..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 cctcaaaagg
     2281 gtgcgcatct gatgagtccg tgaggacgaa acgagctagc tcgtcatgcg cacccttagc
     2341 gagaggttta tcattaaggt caacctctgg atgttgtttc ggcatcctgc attgaatctg
     2401 agttactgtc tgttttcctt ggaaatgttc tatttagaaa caggggaatt gcttattaac
     2461 gaaattgcct gaaggcctca caactctgga cattatacca ttgatgcttg cgtcacttca
     2521 ggaaacccgt ttcttctgac gtaagggtgc gcaattaacg aaattgcccc agtttcagag
     2581 ctatgctgga aacagcatag caagttgaaa taaggctagt ccgttatcaa cttgaaaaag
     2641 tggcaccgag tcggtgcttt ttgatggccg gcatggtccc agcctcctcg ctggcgccgg
     2701 ctgggcaaca ccttcgggtg gcgaatggga cttaaagaaa gtggaatatt cattcatatc
     2761 atattttttc tattaactgc ctggtttctt ttaaattttt tattggttgt cgacttgaac
     2821 ggagtgacaa tatatatata tatatttaat aatgacatca ttatctgtaa atctgattct
     2881 taatgctatt ctagttatgt aagagtggtc ctttccataa aaaaaaaaaa aaagaaaaaa
     2941 gaattttagg aatacaatgc agcttgtaag taaaatctgg aatattcata tcgccacaac
     3001 ttcttatgct tataaaagca ctaatgcctg aatttatgtt gaaaatatgt gtcacaaata
     3061 aagaaactgt gacatctgac acatttccac gtacccaatt cgccctatag tgagtcgtat
     3121 tacgcgcgct cactggccgt cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc
     3181 caacttaatc gccttgcagc acatccccct ttcgccagct ggcgtaatag cgaagaggcc
     3241 cgcaccgatc gcccttccca acagttgcgc agcctgaatg gcgaatggac gcgccctgta
     3301 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
     3361 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
     3421 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
     3481 acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat
     3541 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
     3601 aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc
     3661 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta
     3721 acaaaatatt aacgcttaca atttcctgat gcggtatttt ctccttacgc atctgtgcgg
     3781 tatttcacac cgcatagggt aataactgat ataattaaat tgaagctcta atttgtgagt
     3841 ttagtataca tgcatttact tataatacag ttttttagtt ttgctggccg catcttctca
     3901 aatatgcttc ccagcctgct tttctgtaac gttcaccctc taccttagca tcccttccct
     3961 ttgcaaatag tcctcttcca acaataataa tgtcagatcc tgtagagacc acatcatcca
     4021 cggttctata ctgttgaccc aatgcgtctc ccttgtcatc taaacccaca ccgggtgtca
     4081 taatcaacca atcgtaacct tcatctcttc cacccatgtc tctttgagca ataaagccga
     4141 taacaaaatc tttgtcgctc ttcgcaatgt caacagtacc cttagtatat tctccagtag
     4201 atagggagcc cttgcatgac aattctgcta acatcaaaag gcctctaggt tcctttgtta
     4261 cttcttctgc cgcctgcttc aaaccgctaa caatacctgg gcccaccaca ccgtgtgcat
     4321 tcgtaatgtc tgcccattct gctattctgt atacacccgc agagtactgc aatttgactg
     4381 tattaccaat gtcagcaaat tttctgtctt cgaagagtaa aaaattgtac ttggcggata
     4441 atgcctttag cggcttaact gtgccctcca tggaaaaatc agtcaagata tccacatgtg
     4501 tttttagtaa acaaattttg ggacctaatg cttcaactaa ctccagtaat tccttggtgg
     4561 tacgaacatc caatgaagca cacaagtttg tttgcttttc gtgcatgata ttaaatagct
     4621 tggcagcaac aggactagga tgagtagcag cacgttcctt atatgtagct ttcgacatga
     4681 tttatcttcg tttcctgcag gtttttgttc tgtgcagttg ggttaagaat actgggcaat
     4741 ttcatgtttc ttcaacacta catatgcgta tatataccaa tctaagtctg tgctccttcc
     4801 ttcgttcttc cttctgttcg gagattaccg aatcaaaaaa atttcaagga aaccgaaatc
     4861 aaaaaaaaga ataaaaaaaa aatgatgaat tgaaaaggtg gtatggtgca ctctcagtac
     4921 aatctgctct gatgccgcat agttaagcca gccccgacac ccgccaacac ccgctgacgc
     4981 gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg
     5041 gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgagac gaaagggcct
     5101 cgtgatacgc ctatttttat aggttaatgt catgataata atggtttctt agacggatcg
     5161 cttgcctgta acttacacgc gcctcgtatc ttttaatgat ggaataattt gggaatttac
     5221 tctgtgttta tttattttta tgttttgtat ttggatttta gaaagtaaat aaagaaggta
     5281 gaagagttac ggaatgaaga aaaaaaaata aacaaaggtt taaaaaattt caacaaaaag
     5341 cgtactttac atatatattt attagacaag aaaagcagat taaatagata tacattcgat
     5401 taacgataag taaaatgtaa aatcacagga ttttcgtgtg tggtcttcta cacagacaag
     5461 atgaaacaat tcggcattaa tacctgagag caggaagagc aagataaaag gtagtatttg
     5521 ttggcgatcc ccctagagtc ttttacatct tcggaaaaca aaaactatt
//
