LOCUS       sequence-311798-        2904 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Low phosphate inducible gRNA array to silence the xylA and gltA2 
            promoters.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2904)
  AUTHORS   Li S, Ye Z, Moreb EA, Hennigan JN, Castellanos DB, Yang T, Lynch MD
  TITLE     Dynamic control over feedback regulatory mechanisms improves NADPH 
            flux and xylitol biosynthesis in engineered E. coli.
  JOURNAL   Metab Eng. 2021 Jan 15. pii: S1096-7176(21)00013-6. doi: 
            10.1016/j.ymben.2021.01.005.
  PUBMED    33460820
REFERENCE   2  (bases 1 to 2904)
  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. 
            2021 Jan 15. pii: S1096-7176(21)00013-6. doi: 
            10.1016/j.ymben.2021.01.005."
COMMENT     Sequence Label: pCASCADE-gltA2-xylA
FEATURES             Location/Qualifiers
     source          1..2904
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     129..148
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(215..317)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             complement(347..973)
                     /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     878..897
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     terminator      982..1053
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1069..1096
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     primer_bind     complement(1551..1568)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1685..2230)
                     /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(1765..1784)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(2344..2438)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(join(2459..2904,1..214))
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
ORIGIN
        1 ccggatgagc attcatcagg cgggcaagaa tgtgaataaa ggccggataa aacttgtgct
       61 tatttttctt tacggtcttt aaaaaggccg taatatccag ctgaacggtc tggttatagg
      121 tacattgagc aactgactga aatgcctcaa aatgttcttt acgatgccat tgggatatat
      181 caacggtggt atatccagtg atttttttct ccattttagc ttccttagct cctgaaaatc
      241 tcgataactc aaaaaatacg cccggtagtg atcttatttc attatggtga aagttggaac
      301 ctcttacgtg ccgatcaacg tctcattttc gccagatatc gacgtcttaa gacccacttt
      361 cacatttaag ttgtttttct aatccgcata tgatcaattc aaggccgaat aagaaggctg
      421 gctctgcacc ttggtgatca aataattcga tagcttgtcg taataatggc ggcatactat
      481 cagtagtagg tgtttccctt tcttctttag cgacttgatg ctcttgatct tccaatacgc
      541 aacctaaagt aaaatgcccc acagcgctga gtgcatataa tgcattctct agtgaaaaac
      601 cttgttggca taaaaaggct aattgatttt cgagagtttc atactgtttt tctgtaggcc
      661 gtgtacctaa atgtactttt gctccatcgc gatgacttag taaagcacat ctaaaacttt
      721 tagcgttatt acgtaaaaaa tcttgccagc tttccccttc taaagggcaa aagtgagtat
      781 ggtgcctatc taacatctca atggctaagg cgtcgagcaa agcccgctta ttttttacat
      841 gccaatacaa tgtaggctgc tctacaccta gcttctgggc gagtttacgg gttgttaaac
      901 cttcgattcc gacctcatta agcagctcta atgcgctgtt aatcacttta cttttatcta
      961 atctagacat catccaggca tcaaataaaa cgaaaggctc agtcgaaaga ctgggccttt
     1021 cgttttatct gttgtttgtc ggtgaacgct ctctactaga gtcacactgg ctcaccttcg
     1081 ggtgggcctt tctgcgttta tacacagcta acaccacgtc gtccctatct gctgccctag
     1141 gtctatgagt ggttgctgga taactctttc tgacacctta ctatcttaca aatgtaacaa
     1201 aaaagttatt tttctgtaat tcgagcatgt catgttaccc cgcgagcata aaacgcgtat
     1261 attcagggag accacaacgg tttccctcta caaataattt tgtttaactt tgaattcaaa
     1321 agatctggta ccacctcgag ttccccgcgc cagcggggat aaaccgtatt gaccaattca
     1381 ttcgggacag ttattagttc gagttccccg cgccagcggg gataaaccgg gagtgcccaa
     1441 tattacgaca tcatccatct cgagttcccc gcgccagcgg ggataaaccg aaaaaaaaac
     1501 cccgcccctg acagggcggg gttttttttc ctagggatat attccgcttc ctcgctcact
     1561 gactcgctac gctcggtcgt tcgactgcgg cgagcggaaa tggcttacga acggggcgga
     1621 gatttcctgg aagatgccag gaagatactt aacagggaag tgagagggcc gcggcaaagc
     1681 cgtttttcca taggctccgc ccccctgaca agcatcacga aatctgacgc tcaaatcagt
     1741 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctggc ggctccctcg
     1801 tgcgctctcc tgttcctgcc tttcggttta ccggtgtcat tccgctgtta tggccgcgtt
     1861 tgtctcattc cacgcctgac actcagttcc gggtaggcag ttcgctccaa gctggactgt
     1921 atgcacgaac cccccgttca gtccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     1981 tccaacccgg aaagacatgc aaaagcacca ctggcagcag ccactggtaa ttgatttaga
     2041 ggagttagtc ttgaagtcat gcgccggtta aggctaaact gaaaggacaa gttttggtga
     2101 ctgcgctcct ccaagccagt tacctcggtt caaagagttg gtagctcaga gaaccttcga
     2161 aaaaccgccc tgcaaggcgg ttttttcgtt ttcagagcaa gagattacgc gcagaccaaa
     2221 acgatctcaa gaagatcatc ttattaatca gataaaatat ttctagattt cagtgcaatt
     2281 tatctcttca aatgtagcac ctgaagtcag ccccatacga tataagttgt tactagtgct
     2341 tggattctca ccaataaaaa acgcccggcg gcaaccgagc gttctgaaca aatccagatg
     2401 gagttctgag gtcattactg gatctatcaa caggagtcca agcgagctcg atatcaaatt
     2461 acgccccgcc ctgccactca tcgcagtact gttgtaattc attaagcatt ctgccgacat
     2521 ggaagccatc acaaacggca tgatgaacct gaatcgccag cggcatcagc accttgtcgc
     2581 cttgcgtata atatttgccc atggtgaaaa cgggggcgaa gaagttgtcc atattggcca
     2641 cgtttaaatc aaaactggtg aaactcaccc agggattggc tgagacgaaa aacatattct
     2701 caataaaccc tttagggaaa taggccaggt tttcaccgta acacgccaca tcttgcgaat
     2761 atatgtgtag aaactgccgg aaatcgtcgt ggtattcact ccagagcgat gaaaacgttt
     2821 cagtttgctc atggaaaacg gtgtaacaag ggtgaacact atcccatatc accagctcac
     2881 cgtctttcat tgccatacga aatt
//
