LOCUS       pSC-MRscRNA             5265 bp ds-DNA     circular     19-MAY-2025
DEFINITION  sgRNA with 1x MS2 for yeast cells.
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
COMMENT     Imported using the Genbank importer. File name:
            addgene-plasmid-62313-sequence-111865.gbk
FEATURES             Location/Qualifiers
     primer          68..87
                     /label="T7 (universal primer)"
                     /note="sequence: taatacgactcactataggg"
                     /ApEinfo_revcolor="#ff9ccd"
                     /ApEinfo_fwdcolor="#ff9ccd"
     promoter        112..380
                     /label="SNR52 promoter"
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /note="promoter for the S. cerevisiae small nucleolar RNA gene SNR52"
     misc_feature    381..400
                     /label="TetO sgRNA"
                     /ApEinfo_revcolor="#ff9ccd"
                     /ApEinfo_fwdcolor="#ff9ccd"
     misc_feature    401..430
                     /label="dCas9 handle"
                     /ApEinfo_revcolor="#c7b0e3"
                     /ApEinfo_fwdcolor="#c7b0e3"
     misc_RNA        401..461
                     /label="gRNA scaffold"
                     /ApEinfo_revcolor="#faac61"
                     /ApEinfo_fwdcolor="#faac61"
                     /note="guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system"
     misc_feature    447..461
                     /label="b2 terminator"
                     /ApEinfo_revcolor="#ffef86"
                     /ApEinfo_fwdcolor="#ffef86"
     misc_feature    462..546
                     /label="Eterna Theo/MS2 Aptamers (7721394 - A)"
                     /ApEinfo_revcolor="#ff9ccd"
                     /ApEinfo_fwdcolor="#ff9ccd"
     terminator      547..566
                     /label="SUP4 terminator"
                     /ApEinfo_revcolor="#f8d3a9"
                     /ApEinfo_fwdcolor="#f8d3a9"
                     /note="transcription terminator for the S. cerevisiae SUP4 tRNA gene"
     rep_origin      complement(1036..1624)
                     /label="ori"
                     /ApEinfo_revcolor="#faac61"
                     /ApEinfo_fwdcolor="#faac61"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication"
                     /direction="LEFT"
     misc_feature    complement(1795..2655)
                     /label="AmpR"
                     /ApEinfo_revcolor="#ff9ccd"
                     /ApEinfo_fwdcolor="#ff9ccd"
     CDS             complement(1795..2655)
                     /label="AmpR"
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /note="confers resistance to ampicillin, carbenicillin, andrelated antibiotics"
                     /gene="bla"
                     /product="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYIELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFFHNMGDHVTRLDRWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGASLIKHW*"
     promoter        complement(2656..2760)
                     /label="AmpR promoter"
                     /ApEinfo_revcolor="#ffef86"
                     /ApEinfo_fwdcolor="#ffef86"
                     /gene="bla"
     misc_feature    2797..3300
                     /label="CEN/ARS"
                     /ApEinfo_revcolor="#b7e6d7"
                     /ApEinfo_fwdcolor="#b7e6d7"
                     /note="S. cerevisiae CEN6 centromere fused to an autonomously replicating sequence"
     promoter        3564..3779
                     /label="URA3 promoter"
                     /ApEinfo_revcolor="#c6c9d1"
                     /ApEinfo_fwdcolor="#c6c9d1"
                     /gene="S. cerevisiae URA3"
     misc_feature    3780..4583
                     /label="URA3"
                     /ApEinfo_revcolor="#b1ff67"
                     /ApEinfo_fwdcolor="#b1ff67"
     CDS             3780..4583
                     /label="URA3"
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /note="yeast auxotrophic marker, counterselectable with 5-fluoroorotic acid (5-FOA)"
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required for uracil biosynthesis"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELLELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQYSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGEYTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDDVVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN*"
     rep_origin      complement(4714..5169)
                     /label="f1 ori"
                     /ApEinfo_revcolor="#f58a5e"
                     /ApEinfo_fwdcolor="#f58a5e"
                     /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis"
                     /direction="LEFT"
ORIGIN
        1 gcgattaagt tgggtaacgc cagggttttc ccagtcacga cgttgtaaaa cgacggccag
       61 tgaattgtaa tacgactcac tatagggcga attggagctc actcggaatt ctctttgaaa
      121 agataatgta tgattatgct ttcactcata tttatacaga aacttgatgt tttctttcga
      181 gtatatacaa ggtgattaca tgtacgtttg aagtacaact ctagattttg tagtgccctc
      241 ttgggctagc ggtaaaggtg cgcatttttt cacaccctac aatgttctgt tcaaaagatt
      301 ttggtcaaac gctgtagaag tgaaagttgg tgcgcatgtt tcggcgttcg aaacttctcc
      361 gcagtgaaag ataaatgatc acttttctct atcactgata gttttagagc tagaaatagc
      421 aagttaaaat aaggctagtc cgttatcaac ttgaaaaagt gTGTATGTGA TCTGATACCA
      481 GCCTCATAGC TTGAGGATCA CCCAAGTGGA GCGGAAACGC TGGCCCTTGG CAGCAGATCG
      541 CATACTtttt tttgtttttt atgtctctgc agagttcggt accagctttt gttcccttta
      601 gtgagggtta atttcgagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg
      661 ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg
      721 tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc
      781 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
      841 tcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
      901 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
      961 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     1021 cgcgttgctg gcgtttttcc ataggctcgg cccccctgac gagcatcaca aaaatcgacg
     1081 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt tcccccctgg
     1141 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     1201 tctcccttcg ggaagcgtgg cgctttctca atgctcacgc tgtaggtatc tcagttcggt
     1261 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     1321 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     1381 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     1441 cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta tctgcgctct
     1501 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     1561 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     1621 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     1681 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     1741 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     1801 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     1861 ctgactgccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     1921 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     1981 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     2041 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     2101 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     2161 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgaaaaa aagcggttag
     2221 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     2281 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     2341 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     2401 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     2461 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     2521 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     2581 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     2641 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     2701 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     2761 cacatttccc cgaaaagtgc cacctgggtc cttttcatca cgtgctataa aaataattat
     2821 aatttaaatt ttttaatata aatatataaa ttaaaaatag aaagtaaaaa aagaaattaa
     2881 agaaaaaata gtttttgttt tccgaagatg taaaagactc tagggggatc gccaacaaat
     2941 actacctttt atcttgctct tcctgctctc aggtattaat gccgaattgt ttcatcttgt
     3001 ctgtgtagaa gaccacacac gaaaatcctg tgattttaca ttttacttat cgttaatcga
     3061 atgtatatct atttaatctg cttttcttgt ctaataaata tatatgtaaa gtacgctttt
     3121 tgttgaaatt ttttaaacct ttgtttattt ttttttcttc attccgtaac tcttctacct
     3181 tctttattta ctttctaaaa tccaaataca aaacataaaa ataaataaac acagagtaaa
     3241 ttcccaaatt attccatcat taaaagatac gaggcgcgtg taagttacag gcaagcgatc
     3301 cgtcctaaga aaccattatt atcatgacat taacctataa aaataggcgt atcacgaggc
     3361 cctttcgtct cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg
     3421 agacggtcac agcttgtctg taagcggatg ccgggagcag acaagcccgt cagggcgcgt
     3481 cagcgggtgt tggcgggtgt cggggctggc ttaactatgc ggcatcagag cagattgtac
     3541 tgagagtgca ccacgctttt caattcaatt catcattttt tttttattct tttttttgat
     3601 ttcggtttcc ttgaaatttt tttgattcgg taatctccga acagaaggaa gaacgaagga
     3661 aggagcacag acttagattg gtatatatac gcatatgtag tgttgaagaa acatgaaatt
     3721 gcccagtatt cttaacccaa ctgcacagaa caaaaacgtg caggaaacga agataaatca
     3781 tgtcgaaagc tacatataag gaacgtgctg ctactcatcc tagtcctgtt gctgccaagc
     3841 tatttaatat catgcacgaa aagcaaacaa acttgtgtgc ttcattggat gttcgtacca
     3901 ccaaggaatt actggagtta gttgaagcat taggtcccaa aatttgttta ctaaaaacac
     3961 atgtggatat cttgactgat ttttccatgg agggcacagt taagccgcta aaggcattat
     4021 ccgccaagta caatttttta ctcttcgaag acagaaaatt tgctgacatt ggtaatacag
     4081 tcaaattgca gtactctgcg ggtgtataca gaatagcaga atgggcagac attacgaatg
     4141 cacacggtgt ggtgggccca ggtattgtta gcggtttgaa gcaggcggca gaagaagtaa
     4201 caaaggaacc tagaggcctt ttgatgttag cagaattgtc atgcaagggc tccctatcta
     4261 ctggagaata tactaagggt actgttgaca ttgcgaagag cgacaaagat tttgttatcg
     4321 gctttattgc tcaaagagac atgggtggaa gagatgaagg ttacgattgg ttgattatga
     4381 cacccggtgt gggtttagat gacaagggag acgcattggg tcaacagtat agaaccgtgg
     4441 atgatgtggt ctctacagga tctgacatta ttattgttgg aagaggacta tttgcaaagg
     4501 gaagggatgc taaggtagag ggtgaacgtt acagaaaagc aggctgggaa gcatatttga
     4561 gaagatgcgg ccagcaaaac taaaaaactg tattataagt aaatgcatgt atactaaact
     4621 cacaaattag agcttcaatt taattatatc agttattacc ctatgcggtg tgaaataccg
     4681 cacagatgcg taaggagaaa ataccgcatc aggaaattgt aaacgttaat attttgttaa
     4741 aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca
     4801 aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga
     4861 acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc
     4921 agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc
     4981 gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc
     5041 cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg
     5101 caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac
     5161 agggcgcgtc cattcgccat tcaggctgcg caactgttgg gaagggcgat cggtgcgggc
     5221 ctcttcgcta ttacgccagc tggcgaaagg gggatgtgct gcaag
//