LOCUS       sequence-399077-        2841 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a guide RNA to silence trxB after phosphate depletion.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2841)
  AUTHORS   Hennigan JN, Menacho-Melgar R, Sarkar P, Golovsky M, Lynch MD
  TITLE     
  JOURNAL   Metab Eng. 2024 Sep;85:116-130. doi: 10.1016/j.ymben.2024.07.012. 
            Epub 2024 Jul 24.
  PUBMED    39059674
REFERENCE   2  (bases 1 to 2841)
  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: "Metab 
            Eng."; date: "2024-09"; volume: "85:116-130. doi"; pages: " 
            10.1016/j.ymben.2024.07.012. Epub 2024 Jul 24"
COMMENT     Sequence Label: pCASCADE-LCAmp-trxB
FEATURES             Location/Qualifiers
     source          1..2841
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(7..633)
                     /codon_start=1
                     /gene="tetR"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MMSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWH
                     VKNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGT
                     RPTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPT
                     TDSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     538..557
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     terminator      642..713
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      729..756
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     primer_bind     complement(1148..1165)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1282..1827)
                     /direction=LEFT
                     /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     complement(1362..1381)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(1941..2035)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(2056..2715)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     2630..2649
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(2716..2818)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
ORIGIN
        1 gacgtcttaa gacccacttt cacatttaag ttgtttttct aatccgcata tgatcaattc
       61 aaggccgaat aagaaggctg gctctgcacc ttggtgatca aataattcga tagcttgtcg
      121 taataatggc ggcatactat cagtagtagg tgtttccctt tcttctttag cgacttgatg
      181 ctcttgatct tccaatacgc aacctaaagt aaaatgcccc acagcgctga gtgcatataa
      241 tgcattctct agtgaaaaac cttgttggca taaaaaggct aattgatttt cgagagtttc
      301 atactgtttt tctgtaggcc gtgtacctaa atgtactttt gctccatcgc gatgacttag
      361 taaagcacat ctaaaacttt tagcgttatt acgtaaaaaa tcttgccagc tttccccttc
      421 taaagggcaa aagtgagtat ggtgcctatc taacatctca atggctaagg cgtcgagcaa
      481 agcccgctta ttttttacat gccaatacaa tgtaggctgc tctacaccta gcttctgggc
      541 gagtttacgg gttgttaaac cttcgattcc gacctcatta agcagctcta atgcgctgtt
      601 aatcacttta cttttatcta atctagacat catccaggca tcaaataaaa cgaaaggctc
      661 agtcgaaaga ctgggccttt cgttttatct gttgtttgtc ggtgaacgct ctctactaga
      721 gtcacactgg ctcaccttcg ggtgggcctt tctgcgttta tacacagcta acaccacgtc
      781 gtccctatct gctgccctag gtctatgagt ggttgctgga taactctttc tgacacctta
      841 ctatcttaca aatgtaacaa aaaagttatt tttctgtaat tcgagcatgt catgttaccc
      901 cgcgagcata aaacgcgtat attcagggag accacaacgg tttccctcta caaataattt
      961 tgtttaactt tgaattcaaa agatctggta ccacctcgag ttccccgcgc cagcggggat
     1021 aaaccgattg tagggaattt acagacgtaa aaaaagtcga gttccccgcg ccagcgggga
     1081 taaaccgaaa aaaaaacccc gcccctgaca gggcggggtt ttttttccta gggatatatt
     1141 ccgcttcctc gctcactgac tcgctacgct cggtcgttcg actgcggcga gcggaaatgg
     1201 cttacgaacg gggcggagat ttcctggaag atgccaggaa gatacttaac agggaagtga
     1261 gagggccgcg gcaaagccgt ttttccatag gctccgcccc cctgacaagc atcacgaaat
     1321 ctgacgctca aatcagtggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     1381 ccctggcggc tccctcgtgc gctctcctgt tcctgccttt cggtttaccg gtgtcattcc
     1441 gctgttatgg ccgcgtttgt ctcattccac gcctgacact cagttccggg taggcagttc
     1501 gctccaagct ggactgtatg cacgaacccc ccgttcagtc cgaccgctgc gccttatccg
     1561 gtaactatcg tcttgagtcc aacccggaaa gacatgcaaa agcaccactg gcagcagcca
     1621 ctggtaattg atttagagga gttagtcttg aagtcatgcg ccggttaagg ctaaactgaa
     1681 aggacaagtt ttggtgactg cgctcctcca agccagttac ctcggttcaa agagttggta
     1741 gctcagagaa ccttcgaaaa accgccctgc aaggcggttt tttcgttttc agagcaagag
     1801 attacgcgca gaccaaaacg atctcaagaa gatcatctta ttaatcagat aaaatatttc
     1861 tagatttcag tgcaatttat ctcttcaaat gtagcacctg aagtcagccc catacgatat
     1921 aagttgttac tagtgcttgg attctcacca ataaaaaacg cccggcggca accgagcgtt
     1981 ctgaacaaat ccagatggag ttctgaggtc attactggat ctatcaacag gagtccaagc
     2041 gagctcgata tcaaattacg ccccgccctg ccactcatcg cagtactgtt gtaattcatt
     2101 aagcattctg ccgacatgga agccatcaca aacggcatga tgaacctgaa tcgccagcgg
     2161 catcagcacc ttgtcgcctt gcgtataata tttgcccatg gtgaaaacgg gggcgaagaa
     2221 gttgtccata ttggccacgt ttaaatcaaa actggtgaaa ctcacccagg gattggctga
     2281 gacgaaaaac atattctcaa taaacccttt agggaaatag gccaggtttt caccgtaaca
     2341 cgccacatct tgcgaatata tgtgtagaaa ctgccggaaa tcgtcgtggt attcactcca
     2401 gagcgatgaa aacgtttcag tttgctcatg gaaaacggtg taacaagggt gaacactatc
     2461 ccatatcacc agctcaccgt ctttcattgc catacgaaat tccggatgag cattcatcag
     2521 gcgggcaaga atgtgaataa aggccggata aaacttgtgc ttatttttct ttacggtctt
     2581 taaaaaggcc gtaatatcca gctgaacggt ctggttatag gtacattgag caactgactg
     2641 aaatgcctca aaatgttctt tacgatgcca ttgggatata tcaacggtgg tatatccagt
     2701 gatttttttc tccattttag cttccttagc tcctgaaaat ctcgataact caaaaaatac
     2761 gcccggtagt gatcttattt cattatggtg aaagttggaa cctcttacgt gccgatcaac
     2821 gtctcatttt cgccagatat c
//
