LOCUS       sequence-197178-        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-R18
FEATURES             Location/Qualifiers
     source          1..2936
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   57..92
                     /label=DR
                     /note="direct repeat for the Streptococcus pyogenes 
                     CRISPR/Cas system"
     misc_RNA        213..291
                     /label=tracrRNA
                     /note="trans-activating CRISPR RNA for the Streptococcus 
                     pyogenes CRISPR/Cas9 system"
     primer_bind     complement(436..454)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     CDS             633..1427
                     /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(687..706)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1297..1316
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     terminator      1458..1552
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     rep_origin      1640..2228
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2129..2148
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(2613..2699)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     misc_feature    2729..2860
                     /label=crRNA leader
                     /note="crRNA leader sequence for the Streptococcus pyogenes
                     CRISPR/Cas system"
     repeat_region   2861..2896
                     /label=DR
                     /note="direct repeat for the Streptococcus pyogenes 
                     CRISPR/Cas system"
     repeat_region   join(2927..2936,1..26)
                     /label=DR
                     /note="direct repeat for the Streptococcus pyogenes 
                     CRISPR/Cas system"
ORIGIN
        1 tatgctgttt tgaatggtcc caaaacagtt tggcggagtg ggtgccttca tacggagttt
       61 tagagctatg ctgttttgaa tggtcccaaa acttcagcac actgagactt gttgagttga
      121 attcatgagt attaagtatt gttttatggc tgataaattt ctttgaattt ctccttgatt
      181 atttgttata aaagttataa aataatcttg ttggaaccat tcaaaacagc atagcaagtt
      241 aaaataaggc tagtccgtta tcaacttgaa aaagtggcac cgagtcggtg ctttttttga
      301 tacttctatt ctactctgac tgcaaaccaa aaaaacaagc gctttcaaaa cgcttgtttt
      361 atcattttta gggaaattaa tctcttaatc cttttatcat tctacattta ggcgctgcca
      421 tcttaagacg aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa
      481 tggtttctta gacgtcggaa ttgccagctg gggcgccctc tggtaaggtt gggaagccct
      541 gcaaagtaaa ctggatggct ttcttgccgc caaggatctg atggcgcagg ggatcaagat
      601 ctgatcaaga gacaggatga ggatcgtttc gcatgattga acaagatgga ttgcacgcag
      661 gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa cagacaatcg
      721 gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca
      781 agaccgacct gtccggtgcc ctgaatgaac tgcaggacga ggcagcgcgg ctatcgtggc
      841 tggccacgac gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg
      901 actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac cttgctcctg
      961 ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt gatccggcta
     1021 cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact cggatggaag
     1081 ccggtcttgt cgatcaggat gatctggacg aagagcatca ggggctcgcg ccagccgaac
     1141 tgttcgccag gctcaaggcg cgcatgcccg acggcgagga tctcgtcgtg acccatggcg
     1201 atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc atcgactgtg
     1261 gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt gatattgctg
     1321 aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc gccgctcccg
     1381 attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgagcg ggactctggg
     1441 gttcgagagc tcgcttggac tcctgttgat agatccagta atgacctcag aactccatct
     1501 ggatttgttc agaacgctcg gttgccgccg ggcgtttttt attggtgaga atccaagcac
     1561 tagtcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     1621 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     1681 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     1741 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc
     1801 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     1861 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     1921 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     1981 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     2041 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     2101 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     2161 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     2221 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     2281 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg
     2341 agtgagctga taccgctcgc cgcagccgaa cgccccaaaa agcctcgctt tcagcacctg
     2401 tcgtttcctt tcttttcaga gggtatttta aataaaaaca ttaagttatg acgaagaaga
     2461 acggaaacgc cttaaaccgg aaaattttca taaatagcga aaacccgcga ggtcgccgcc
     2521 ccgtaacctg tcggatcacc ggaaaggacc cgtaaagtga taatgattat catctacata
     2581 tcacaacgtg cgtggagggt ctagggcggc ggatttgtcc tactcaggag agcgttcacc
     2641 gacaaacaac agataaaacg aaaggcccag tctttcgact gagcctttcg ttttatttga
     2701 tgcctctagc acgcgtacca tgggatccta tttcttaata actaaaaata tggtataata
     2761 ctcttaataa atgcagtaat acaggggctt ttcaagactg aagtctagct gagacaaata
     2821 gtgcgattac gaaatttttt agacaaaaat agtctacgag gttttagagc tatgctgttt
     2881 tgaatggtcc caaaaccctt caccttgtgg tgcttgagag aatttggttt tagagc
//
