LOCUS       sequence-197166-        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-G20-R0
FEATURES             Location/Qualifiers
     source          1..2936
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(42..60)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     CDS             239..1033
                     /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(293..312)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     903..922
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     terminator      1064..1158
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     rep_origin      1246..1834
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1735..1754
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(2219..2305)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     misc_feature    2335..2466
                     /label=crRNA leader
                     /note="crRNA leader sequence for the Streptococcus pyogenes
                     CRISPR/Cas system"
     repeat_region   2467..2502
                     /label=DR
                     /note="direct repeat for the Streptococcus pyogenes 
                     CRISPR/Cas system"
     repeat_region   2533..2568
                     /label=DR
                     /note="direct repeat for the Streptococcus pyogenes 
                     CRISPR/Cas system"
     repeat_region   2599..2634
                     /label=DR
                     /note="direct repeat for the Streptococcus pyogenes 
                     CRISPR/Cas system"
     misc_RNA        2755..2833
                     /label=tracrRNA
                     /note="trans-activating CRISPR RNA for the Streptococcus 
                     pyogenes CRISPR/Cas9 system"
ORIGIN
        1 tatcattcta catttaggcg ctgccatctt aagacgaaag ggcctcgtga tacgcctatt
       61 tttataggtt aatgtcatga taataatggt ttcttagacg tcggaattgc cagctggggc
      121 gccctctggt aaggttggga agccctgcaa agtaaactgg atggctttct tgccgccaag
      181 gatctgatgg cgcaggggat caagatctga tcaagagaca ggatgaggat cgtttcgcat
      241 gattgaacaa gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg
      301 ctatgactgg gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc
      361 gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca
      421 ggacgaggca gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct
      481 cgacgttgtc actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga
      541 tctcctgtca tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg
      601 gcggctgcat acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat
      661 cgagcgagca cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga
      721 gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg
      781 cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg
      841 ccgcttttct ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat
      901 agcgttggct acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct
      961 cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga
     1021 cgagttcttc tgagcgggac tctggggttc gagagctcgc ttggactcct gttgatagat
     1081 ccagtaatga cctcagaact ccatctggat ttgttcagaa cgctcggttg ccgccgggcg
     1141 ttttttattg gtgagaatcc aagcactagt catgaccaaa atcccttaac gtgagttttc
     1201 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     1261 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     1321 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     1381 accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     1441 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     1501 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     1561 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     1621 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     1681 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     1741 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     1801 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     1861 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc
     1921 tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgcc
     1981 ccaaaaagcc tcgctttcag cacctgtcgt ttcctttctt ttcagagggt attttaaata
     2041 aaaacattaa gttatgacga agaagaacgg aaacgcctta aaccggaaaa ttttcataaa
     2101 tagcgaaaac ccgcgaggtc gccgccccgt aacctgtcgg atcaccggaa aggacccgta
     2161 aagtgataat gattatcatc tacatatcac aacgtgcgtg gagggtctag ggcggcggat
     2221 ttgtcctact caggagagcg ttcaccgaca aacaacagat aaaacgaaag gcccagtctt
     2281 tcgactgagc ctttcgtttt atttgatgcc tctagcacgc gtaccatggg atcctatttc
     2341 ttaataacta aaaatatggt ataatactct taataaatgc agtaatacag gggcttttca
     2401 agactgaagt ctagctgaga caaatagtgc gattacgaaa ttttttagac aaaaatagtc
     2461 tacgaggttt tagagctatg ctgttttgaa tggtcccaaa acccttcacc ttcaccacga
     2521 acagagaatt tggttttaga gctatgctgt tttgaatggt cccaaaacat cgcacatcct
     2581 ggtcgcgaca ttaagagtgt tttagagcta tgctgttttg aatggtccca aaacttcagc
     2641 acactgagac ttgttgagtt gaattcatga gtattaagta ttgttttatg gctgataaat
     2701 ttctttgaat ttctccttga ttatttgtta taaaagttat aaaataatct tgttggaacc
     2761 attcaaaaca gcatagcaag ttaaaataag gctagtccgt tatcaacttg aaaaagtggc
     2821 accgagtcgg tgcttttttt gatacttcta ttctactctg actgcaaacc aaaaaaacaa
     2881 gcgctttcaa aacgcttgtt ttatcatttt tagggaaatt aatctcttaa tccttt
//
