LOCUS       sequence-197175-        2936 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Repress mCherry and GFP independently, with different strengths.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2936)
  AUTHORS   Vigouroux A, Oldewurtel E, Cui L, Bikard D, van Teeffelen S
  TITLE     Tuning dCas9's ability to block transcription enables robust, 
            noiseless knockdown of bacterial genes.
  JOURNAL   Mol Syst Biol. 2018 Mar 8;14(3):e7899.
  PUBMED    29519933
REFERENCE   2  (bases 1 to 2936)
  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"; journalName: "Mol Syst 
            Biol."; date: "2018-03-8"; volume: "14(3)"; pages: "e7899"
COMMENT     Sequence Label: pCRRNA-G11-R11
FEATURES             Location/Qualifiers
     source          1..2936
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   50..85
                     /label=DR
                     /note="direct repeat for the Streptococcus pyogenes 
                     CRISPR/Cas system"
     misc_RNA        206..284
                     /label=tracrRNA
                     /note="trans-activating CRISPR RNA for the Streptococcus 
                     pyogenes CRISPR/Cas9 system"
     primer_bind     complement(429..447)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     CDS             626..1420
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(680..699)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1290..1309
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     terminator      1451..1545
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     rep_origin      1633..2221
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2122..2141
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(2606..2692)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     misc_feature    2722..2853
                     /label=crRNA leader
                     /note="crRNA leader sequence for the Streptococcus pyogenes
                     CRISPR/Cas system"
     repeat_region   2854..2889
                     /label=DR
                     /note="direct repeat for the Streptococcus pyogenes 
                     CRISPR/Cas system"
     repeat_region   join(2920..2936,1..19)
                     /label=DR
                     /note="direct repeat for the Streptococcus pyogenes 
                     CRISPR/Cas system"
ORIGIN
        1 ttttgaatgg tcccaaaaca gtttggcgga gacccacgct tcatacggag ttttagagct
       61 atgctgtttt gaatggtccc aaaacttcag cacactgaga cttgttgagt tgaattcatg
      121 agtattaagt attgttttat ggctgataaa tttctttgaa tttctccttg attatttgtt
      181 ataaaagtta taaaataatc ttgttggaac cattcaaaac agcatagcaa gttaaaataa
      241 ggctagtccg ttatcaactt gaaaaagtgg caccgagtcg gtgctttttt tgatacttct
      301 attctactct gactgcaaac caaaaaaaca agcgctttca aaacgcttgt tttatcattt
      361 ttagggaaat taatctctta atccttttat cattctacat ttaggcgctg ccatcttaag
      421 acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc
      481 ttagacgtcg gaattgccag ctggggcgcc ctctggtaag gttgggaagc cctgcaaagt
      541 aaactggatg gctttcttgc cgccaaggat ctgatggcgc aggggatcaa gatctgatca
      601 agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc
      661 ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc
      721 tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga
      781 cctgtccggt gccctgaatg aactgcagga cgaggcagcg cggctatcgt ggctggccac
      841 gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct
      901 gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa
      961 agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc
     1021 attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct
     1081 tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc
     1141 caggctcaag gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg
     1201 cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct
     1261 gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct
     1321 tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca
     1381 gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gcgggactct ggggttcgag
     1441 agctcgcttg gactcctgtt gatagatcca gtaatgacct cagaactcca tctggatttg
     1501 ttcagaacgc tcggttgccg ccgggcgttt tttattggtg agaatccaag cactagtcat
     1561 gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat
     1621 caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa
     1681 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa
     1741 ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt agccgtagtt
     1801 aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt
     1861 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata
     1921 gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt
     1981 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
     2041 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
     2101 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
     2161 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
     2221 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
     2281 gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc
     2341 tgataccgct cgccgcagcc gaacgcccca aaaagcctcg ctttcagcac ctgtcgtttc
     2401 ctttcttttc agagggtatt ttaaataaaa acattaagtt atgacgaaga agaacggaaa
     2461 cgccttaaac cggaaaattt tcataaatag cgaaaacccg cgaggtcgcc gccccgtaac
     2521 ctgtcggatc accggaaggg acccgtaaag tgataatgat tatcatctac atatcacaac
     2581 gtgcgtggag ggtctagggc ggcggatttg tcctactcag gagagcgttc accgacaaac
     2641 aacagataaa acgaaaggcc cagtctttcg actgagcctt tcgttttatt tgatgcctct
     2701 agcacgcgta ccatgggatc ctatttctta ataactaaaa atatggtata atactcttaa
     2761 taaatgcagt aatacagggg cttttcaaga ctgaagtcta gctgagacaa atagtgcgat
     2821 tacgaaattt tttagacaaa aatagtctac gaggttttag agctatgctg ttttgaatgg
     2881 tcccaaaacc cttcaccttg tggtgcttca gagaatttgg ttttagagct atgctg
//
