LOCUS       sequence-258321-        3277 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Engineered sgRNA-G6-2MS2 (2 MS2 aptamers) generator circuit 
            including promoter Pbad.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3277)
  AUTHORS   Liu Y, Wan X, Wang B
  TITLE     Engineered CRISPRa enables programmable eukaryote-like gene 
            activation in bacteria.
  JOURNAL   Nat Commun. 2019 Aug 26;10(1):3693. doi: 10.1038/s41467-019-11479-0.
  PUBMED    31451697
REFERENCE   2  (bases 1 to 3277)
  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: "Nat 
            Commun."; date: "2019-08-26"; volume: "10(1):3693. doi"; pages: " 
            10.1038/s41467-019-11479-0"
COMMENT     Sequence Label: pLY124
FEATURES             Location/Qualifiers
     source          1..3277
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             4..882
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     promoter        1034..1136
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             1137..1796
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1203..1222)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      1817..1911
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     rep_origin      2025..2570
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     2471..2490
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2687..2704
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_RNA        complement(2838..2856)
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     misc_RNA        complement(2908..2926)
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     promoter        complement(2970..3254)
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     complement(3028..3047)
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
ORIGIN
        1 agtatggctg aagcgcaaaa tgatcccctg ctgccgggat actcgtttaa tgcccatctg
       61 gtggcgggtt taacgccgat tgaggccaac ggttatctcg atttttttat cgaccgaccg
      121 ctgggaatga aaggttatat tctcaatctc accattcgcg gtcagggggt ggtgaaaaat
      181 cagggacgag aatttgtttg ccgaccgggt gatattttgc tgttcccgcc aggagagatt
      241 catcactacg gtcgtcatcc ggaggctcgc gaatggtatc accagtgggt ttactttcgt
      301 ccgcgcgcct actggcatga atggcttaac tggccgtcaa tatttgccaa tacggggttc
      361 tttcgcccgg atgaagcgca ccagccgcat ttcagcgacc tgtttgggca aatcattaac
      421 gccgggcaag gggaagggcg ctattcggag ctgctggcga taaatctgct tgagcaattg
      481 ttactgcggc gcatggaagc gattaacgag tcgctccatc caccgatgga taatcgggta
      541 cgcgaggctt gtcagtacat cagcgatcac ctggcagaca gcaattttga tatcgccagc
      601 gtcgcacagc atgtttgctt gtcgccgtcg cgtctgtcac atcttttccg ccagcagtta
      661 gggattagcg tcttaagctg gcgcgaggac caacgtatca gccaggcgaa gctgcttttg
      721 agcaccaccc ggatgcctat cgccaccgtc ggtcgcaatg ttggttttga cgatcaactc
      781 tatttctcgc gggtatttaa aaaatgcacc ggggccagcc cgagcgagtt ccgtgccggt
      841 tgtgaagaaa aagtgaatga tgtagccgtc aagttgtcat aatactagag ctcggtacca
      901 aattccagaa aagagacgct ttcgagcgtc ttttttcgtt ttggtccctc tagtattaat
      961 tttgtcttca tgcatgaaga caactaggct ctagaagcgg ccgcgaatta gatatctggc
     1021 gaaaatgaga cgttgatcgg cacgtaagag gttccaactt tcaccataat gaaataagat
     1081 cactaccggg cgtatttttt gagttatcga gattttcagg agctaaggaa gctaaaatgg
     1141 agaaaaaaat cactggatat accaccgttg atatatccca atggcatcgt aaagaacatt
     1201 ttgaggcatt tcagtcagtt gctcaatgta cctataacca gaccgttcag ctggatatta
     1261 cggccttttt aaagaccgta aagaaaaata agcacaagtt ttatccggcc tttattcaca
     1321 ttcttgcccg cctgatgaat gctcatccgg aatttcgtat ggcaatgaaa gacggtgagc
     1381 tggtgatatg ggatagtgtt cacccttgtt acaccgtttt ccatgagcaa actgaaacgt
     1441 tttcatcgct ctggagtgaa taccacgacg atttccggca gtttctacac atatattcgc
     1501 aagatgtggc gtgttacggt gaaaacctgg cctatttccc taaagggttt attgagaata
     1561 tgtttttcgt ctcagccaat ccctgggtga gtttcaccag ttttgattta aacgtggcca
     1621 atatggacaa cttcttcgcc cccgttttca ccatgggcaa atattatacg caaggcgaca
     1681 aggtgctgat gccgctggcg attcaggttc atcatgccgt ttgtgatggc ttccatgtcg
     1741 gcagaatgct taatgaatta caacagtact gcgatgagtg gcagggcggg gcgtaatttg
     1801 atatcgagct cgcttggact cctgttgata gatccagtaa tgacctcaga actccatctg
     1861 gatttgttca gaacgctcgg ttgccgccgg gcgtttttta ttggtgagaa tccaagcact
     1921 agtaacaact tatatcgtat ggggctgact tcaggtgcta catttgaaga gataaattgc
     1981 actgaaatct agaaatattt tatctgatta ataagatgat cttcttgaga tcgttttggt
     2041 ctgcgcgtaa tctcttgctc tgaaaacgaa aaaaccgcct tgcagggcgg tttttcgaag
     2101 gttctctgag ctaccaactc tttgaaccga ggtaactggc ttggaggagc gcagtcacca
     2161 aaacttgtcc tttcagttta gccttaaccg gcgcatgact tcaagactaa ctcctctaaa
     2221 tcaattacca gtggctgctg ccagtggtgc ttttgcatgt ctttccgggt tggactcaag
     2281 acgatagtta ccggataagg cgcagcggtc ggactgaacg gggggttcgt gcatacagtc
     2341 cagcttggag cgaactgcct acccggaact gagtgtcagg cgtggaatga gacaaacgcg
     2401 gccataacag cggaatgaca ccggtaaacc gaaaggcagg aacaggagag cgcacgaggg
     2461 agccgccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc caccactgat
     2521 ttgagcgtca gatttcgtga tgcttgtcag gggggcggag cctatggaaa aacggctttg
     2581 ccgcggccct ctcacttccc tgttaagtat cttcctggca tcttccagga aatctccgcc
     2641 ccgttcgtaa gccatttccg ctcgccgcag tcgaacgacc gagcgtagcg agtcagtgag
     2701 cgaggaagcg gaatatatcc ctaggcctag taggaccaaa acgaaaaaag gccccccttt
     2761 cgggaggcct cttttctgga atttggtacc gagaagcttc aaaaaaagca ccgactcggt
     2821 gccacttggc cctgcagaca tgggtgatcc tcatgttggc caagttgata acggactagc
     2881 cttatttgaa cttgctaggc cctgcagaca tgggtgatcc tcatgttggc ctagctctca
     2941 aacttttttg ccacaacagc ttttctagta tggagaaaca gtagagagtt gcgataaaaa
     3001 gcgtcaggta ggatccgcta atcttatgga taaaaatgct atggcatagc aaagtgtgac
     3061 gccgtgcaaa taatcaatgt ggacttttct gccgtgatta tagacacttt tgttacgcgt
     3121 ttttgtcatg gctttggtcc cgctttgtta cagaatgctt ttaataagcg gggttaccgg
     3181 tttggttagc gagaagagcc agtaaaagac gcagtgacgg caatgtctga tgcaatatgg
     3241 acaattggtt tcttctctga atggcgggag tatgaaa
//
