LOCUS       sequence-296087-        3333 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For CRISPR-Cas12a genome editing in E. coli. Contains a 
            constituitively expressed CRISPR array targeting the ahpC gene in E.
            coli and donor DNA to insert an rfp translational fusion in the ahpC
            gene..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3333)
  AUTHORS   Jervis AJ, Hanko EKR, Dunstan MS, Robinson CJ, Takano E, Scrutton NS
  TITLE     A plasmid toolset for CRISPR-mediated genome editing and CRISPRi 
            gene regulation in Escherichia coli.
  JOURNAL   Microb Biotechnol. 2021 Mar 12. doi: 10.1111/1751-7915.13780.
  PUBMED    33710766
REFERENCE   2  (bases 1 to 3333)
  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: "Microb 
            Biotechnol. 2021 Mar 12. doi: 10.1111/1751-7915.13780."
COMMENT     Sequence Label: pTF-ahpC-rfp
FEATURES             Location/Qualifiers
     source          1..3333
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             124..915
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      1090..1678
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1579..1598
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1832..1849
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        1926..1960
                     /label=J23119(SpeI) promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119) modified to end with an SpeI site"
     terminator      2067..2138
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      2154..2181
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             2400..3077
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
ORIGIN
        1 tcgagttcat gtgcagctcc atcagcaaaa ggggatgata agtttatcac caccgactat
       61 ttgcaacagt gccgttgatc gtgctatgat cgactgatgt catcagcggt ggagtgcaat
      121 gtcatgaggg aagcggtgat cgccgaagta tcgactcaac tatcagaggt agttggcgtc
      181 atcgagcgcc atctcgaacc gacgttgctg gccgtacatt tgtacggctc cgcagtggat
      241 ggcggcctga agccacacag tgatattgat ttgctggtta cggtgaccgt aaggcttgat
      301 gaaacaacgc ggcgagcttt gatcaacgac cttttggaaa cttcggcttc ccctggagag
      361 agcgagattc tccgcgctgt agaagtcacc attgttgtgc acgacgacat cattccgtgg
      421 cgttatccag ctaagcgcga actgcaattt ggagaatggc agcgcaatga cattcttgca
      481 ggtatcttcg agccagccac gatcgacatt gatctggcta tcttgctgac aaaagcaaga
      541 gaacatagcg ttgccttggt aggtccagcg gcggaggaac tctttgatcc ggttcctgaa
      601 caggatctat ttgaggcgct aaatgaaacc ttaacgctat ggaactcgcc gcccgactgg
      661 gctggcgatg agcgaaatgt agtgcttacg ttgtcccgca tttggtacag cgcagtaacc
      721 ggcaaaatcg cgccgaagga tgtcgctgcc gactgggcaa tggagcgcct gccggcccag
      781 tatcagcccg tcatacttga agctagacag gcttatcttg gacaagaaga agatcgcttg
      841 gcctcgcgcg cagatcagtt ggaagaattt gtccactacg tgaaaggcga gatcaccaag
      901 gtagtcggca aataagatgc cgctcgccag tcgattggct gagctcatga agttcctatt
      961 ccgaagttcc gcgaacgcgt aaaggatcta ggtgaagatc ctttttgata atctcatgac
     1021 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa
     1081 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc
     1141 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt
     1201 aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg
     1261 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc
     1321 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt
     1381 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga
     1441 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct
     1501 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
     1561 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
     1621 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
     1681 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt
     1741 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
     1801 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
     1861 gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc gcatatgctg
     1921 gatccttgac agctagctca gtcctaggta taatactagt aatttctact cttgtagata
     1981 ccaaccaggt ccagagacgg agtttttttg aagcttgggc ccgaacaaaa actcggatcc
     2041 aaactcgagt aaggatctcc aggcatcaaa taaaacgaaa ggctcagtcg aaagactggg
     2101 cctttcgttt tatctgttgt ttgtcggtga acgctctcta ctagagtcac actggctcac
     2161 cttcgggtgg gcctttctgc gtttatacct agggtacgat gtccttgatt aacaccaaaa
     2221 ttaaaccttt taaaaaccag gcattcaaaa acggcgaatt catcgaaatc accgaaaaag
     2281 ataccgaagg ccgctggagc gtcttcttct tctacccggc tgactttact ttcgtatgcc
     2341 cgaccgaact gggtgacgtt gctgaccact acgaagaact gcagaaactg ggcgtagaca
     2401 tggcgagtag cgaagacgtt atcaaagagt tcatgcgttt caaagttcgt atggaaggtt
     2461 ccgttaacgg tcacgagttc gaaatcgaag gtgaaggtga aggtcgtccg tacgaaggta
     2521 cccagaccgc taaactgaaa gttaccaaag gtggtccgct gccgttcgct tgggacatcc
     2581 tgtccccgca gttccagtac ggttccaaag cttacgttaa acacccggct gacatcccgg
     2641 actacctgaa actgtccttc ccggaaggtt tcaaatggga acgtgttatg aacttcgaag
     2701 acggtggtgt tgttaccgtt acccaggact cctccctgca agacggtgag ttcatctaca
     2761 aagttaaact gcgtggtacc aacttcccgt ccgacggtcc ggttatgcag aaaaaaacca
     2821 tgggttggga agcttccacc gaacgtatgt acccggaaga cggtgctctg aaaggtgaaa
     2881 tcaaaatgcg tctgaaactg aaagacggtg gtcactacga cgctgaagtt aaaaccacct
     2941 acatggctaa aaaaccggtt cagctgccgg gtgcttacaa aaccgacatc aaactggaca
     3001 tcacctccca caacgaagac tacaccatcg ttgaacagta cgaacgtgct gaaggtcgtc
     3061 actccaccgg tgcttaaatt tccttcgtct ttcacgccat agcggcgttg gcgtcgcccg
     3121 ctcaccccgg tcacttactt gtgtaagctc ccggggattc acaggctagc cgccttgctc
     3181 tgacgcgaaa tacttcggaa attcacctaa ttcttcgggt gctgcggcac ccgattttct
     3241 tccccgcacc atgatgcaag ctgcatccag gtagccgtga attctctaga gtcgacctgc
     3301 agaagcttag atctattacc ctgttatccc tac
//
