LOCUS       sequence-192582-        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-G10-R20
FEATURES             Location/Qualifiers
     source          1..2936
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(12..30)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     CDS             209..1003
                     /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(263..282)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     873..892
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     terminator      1034..1128
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     rep_origin      1216..1804
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1705..1724
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(2189..2275)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     misc_feature    2305..2436
                     /label=crRNA leader
                     /note="crRNA leader sequence for the Streptococcus pyogenes
                     CRISPR/Cas system"
     repeat_region   2437..2472
                     /label=DR
                     /note="direct repeat for the Streptococcus pyogenes 
                     CRISPR/Cas system"
     repeat_region   2503..2538
                     /label=DR
                     /note="direct repeat for the Streptococcus pyogenes 
                     CRISPR/Cas system"
     repeat_region   2569..2604
                     /label=DR
                     /note="direct repeat for the Streptococcus pyogenes 
                     CRISPR/Cas system"
     misc_RNA        2725..2803
                     /label=tracrRNA
                     /note="trans-activating CRISPR RNA for the Streptococcus 
                     pyogenes CRISPR/Cas9 system"
ORIGIN
        1 aagacgaaag ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt
       61 ttcttagacg tcggaattgc cagctggggc gccctctggt aaggttggga agccctgcaa
      121 agtaaactgg atggctttct tgccgccaag gatctgatgg cgcaggggat caagatctga
      181 tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc
      241 tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg
      301 ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac
      361 cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat cgtggctggc
      421 cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg
      481 gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga
      541 gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg
      601 cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg
      661 tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt
      721 cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc
      781 ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg
      841 gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga
      901 gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc
      961 gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc
     1021 gagagctcgc ttggactcct gttgatagat ccagtaatga cctcagaact ccatctggat
     1081 ttgttcagaa cgctcggttg ccgccgggcg ttttttattg gtgagaatcc aagcactagt
     1141 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
     1201 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     1261 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     1321 gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta
     1381 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     1441 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     1501 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     1561 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagcatt gagaaagcgc
     1621 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     1681 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     1741 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     1801 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     1861 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg
     1921 agctgatacc gctcgccgca gccgaacgcc ccaaaaagcc tcgctttcag cacctgtcgt
     1981 ttcctttctt ttcagagggt attttaaata aaaacattaa gttatgacga agaagaacgg
     2041 aaacgcctta aaccggaaaa ttttcataaa tagcgaaaac ccgcgaggtc gccgccccgt
     2101 aacctgtcgg atcaccggaa aggacccgta aagtgataat gattatcatc tacatatcac
     2161 aacgtgcgtg gagggtctag ggcggcggat ttgtcctact caggagagcg ttcaccgaca
     2221 aacaacagat aaaacgaaag gcccagtctt tcgactgagc ctttcgtttt atttgatgcc
     2281 tctagcacgc gtaccatggg atcctatttc ttaataacta aaaatatggt ataatactct
     2341 taataaatgc agtaatacag gggcttttca agactgaagt ctagctgaga caaatagtgc
     2401 gattacgaaa ttttttagac aaaaatagtc tacgaggttt tagagctatg ctgttttgaa
     2461 tggtcccaaa acccttcacc ttgtggtgct tgagagaatt tggttttaga gctatgctgt
     2521 tttgaatggt cccaaaacag tttggcggtc tgggtgcctt catacggagt tttagagcta
     2581 tgctgttttg aatggtccca aaacttcagc acactgagac ttgttgagtt gaattcatga
     2641 gtattaagta ttgttttatg gctgataaat ttctttgaat ttctccttga ttatttgtta
     2701 taaaagttat aaaataatct tgttggaacc attcaaaaca gcatagcaag ttaaaataag
     2761 gctagtccgt tatcaacttg aaaaagtggc accgagtcgg tgcttttttt gatacttcta
     2821 ttctactctg actgcaaacc aaaaaaacaa gcgctttcaa aacgcttgtt ttatcatttt
     2881 tagggaaatt aatctcttaa tccttttatc attctacatt taggcgctgc catctt
//
