LOCUS       pLBH545_Tet-Cas14a1_Locus                4607 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Expresses Cas14a1 locus (including crRNA and tracrRNA) under control
            of a tetracycline inducible promoter.
ACCESSION   .
VERSION     .
KEYWORDS    pLBH545_Tet-Cas14a1_Locus
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4607)
  AUTHORS   Harrington LB, Burstein D, Chen JS, Paez-Espino D, Ma E, Witte IP, 
            Cofsky JC, Kyrpides NC, Banfield JF, Doudna JA
  TITLE     Programmed DNA destruction by miniature CRISPR-Cas14 enzymes.
  JOURNAL   Science. 2018 Oct 18. pii: science.aav4294. doi: 
            10.1126/science.aav4294.
  PUBMED    30337455
REFERENCE   2  (bases 1 to 4607)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            //gt12365.cn
FEATURES             Location/Qualifiers
     source          1..4607
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        42..97
                     /gene="tetR"
                     /label=tetR/tetA promoters
                     /note="overlapping promoters for bacterial tetR and tetA"
     protein_bind    78..96
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     RBS             115..126
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     terminator      2252..2279
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     primer_bind     complement(2307..2324)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2441..2986)
                     /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(2521..2540)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      3100..3194
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(3215..3874)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     3789..3808
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(3875..3977)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             complement(4007..23)
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /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="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     4538..4557
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
ORIGIN
        1 ttacttttat ctaatctaga catcattaat tcctaatttt tgttgacact ctatcgttga
       61 tagagttatt ttaccactcc ctatcagtga tagagaaaag aattcaaaag atctaaagag
      121 gagaaaggat ctatggccaa aaacaccatt accaaaacac tgaaactgcg tattgtgcgt
      181 ccgtataata gcgcagaagt ggaaaaaatt gttgccgacg aaaaaaacaa ccgcgaaaaa
      241 atcgcactgg aaaagaacaa agacaaagtg aaagaagcct gcagcaaaca tctgaaagtt
      301 gcagcatatt gtaccacaca ggttgaacgt aatgcatgcc tgttttgtaa agcacgtaaa
      361 ctggatgaca aattctacca aaaactgcgt ggtcagtttc cggatgcagt tttttggcaa
      421 gaaatcagcg aaatttttcg ccagctgcag aaacaggcag cagaaatcta taatcagagc
      481 ctgatcgaac tgtactacga gatttttatc aaaggcaaag gtattgcaaa tgccagcagc
      541 gttgaacatt atctgagtga tgtttgttat acccgtgcag cagaactgtt taaaaacgca
      601 gcaattgcaa gcggtctgcg tagcaaaatc aaaagcaatt ttcgtctgaa agaactgaaa
      661 aacatgaaaa gtggtctgcc gaccaccaaa agcgataatt ttccgattcc gctggttaaa
      721 cagaaaggtg gtcagtatac cggttttgaa attagcaatc ataatagcga cttcatcatc
      781 aagattccgt ttggtcgttg gcaggtcaaa aaagagattg ataaatatcg tccgtgggag
      841 aaatttgact ttgaacaggt tcagaaaagc ccgaaaccga ttagcctgct gctgagcacc
      901 cagcgtcgta aacgtaataa aggttggagc aaagatgaag gcaccgaagc cgaaatcaaa
      961 aaagttatga atggcgatta tcagaccagc tacattgaag ttaaacgtgg cagcaaaatc
     1021 ggtgaaaaaa gcgcatggat gctgaatctg agcattgatg ttccgaaaat tgataaaggt
     1081 gtggatccga gcattattgg tggtattgat gttggtgtta aatcaccgct ggtttgcgca
     1141 attaacaatg catttagccg ttatagcatc agcgataacg acctgtttca cttcaacaag
     1201 aaaatgtttg cacgtcgtcg tatcctgctg aaaaaaaacc gtcataaacg tgcaggtcat
     1261 ggtgcaaaaa acaaactgaa accgatcacc attctgaccg aaaaaagtga acgttttcgc
     1321 aaaaagctga ttgaacgttg ggcatgtgaa atcgcggatt tcttcattaa aaacaaagtt
     1381 ggcaccgtgc agatggaaaa tctggaaagc atgaaacgta aagaggacag ctattttaac
     1441 attcgcctgc gtggcttttg gccgtatgca gaaatgcaga acaaaatcga attcaaactg
     1501 aagcagtatg gcatcgaaat tcgtaaagtt gcaccgaata ataccagcaa aacctgtagc
     1561 aaatgtggcc atctgaacaa ctatttcaac ttcgagtacc gcaagaaaaa caaattcccg
     1621 cactttaaat gcgaaaaatg caacttcaaa gaaaacgccg attataatgc agccctgaat
     1681 atttcaaacc cgaaactgaa aagcaccaaa gaggaaccgt aaatatttat actttattat
     1741 ccttcattga caaaaatgag aatgttatcc cagataacat ttgatgtaca cagattcaca
     1801 cttcactgat aaagtggaga accgcttcac caaaagctgt cccttagggg attagaactt
     1861 gagtgaaggt gggctgcttg catcagccta atgtcgagaa gtgctttctt cggaaagtaa
     1921 ccctcgaaac aaattcattt ttcctctcca attctgcaca aaaaaaggtg agtccttata
     1981 aaccggcgtg cagaacgccg gctcaccttt tttcttcatt cgattttatg cttaaaagcc
     2041 gtaaaaacgc ggaattcggc gccgttgcag aacccgaata gacgaatgaa ggaatgcaac
     2101 taccttacac cgcttgcgaa agttgtcaga agataatctt gcagaacccg aatagacgaa
     2161 tgaaggaatg caatcttgac agagcccgat tgcgttatct ccaggagaaa catataaaag
     2221 catcaaccgc tgatcggact agagtcacac tggctcacct tcgggtgggc ctttctgcgt
     2281 ttatacctag ggatatattc cgcttcctcg ctcactgact cgctacgctc ggtcgttcga
     2341 ctgcggcgag cggaaatggc ttacgaacgg ggcggagatt tcctggaaga tgccaggaag
     2401 atacttaaca gggaagtgag agggccgcgg caaagccgtt tttccatagg ctccgccccc
     2461 ctgacaagca tcacgaaatc tgacgctcaa atcagtggtg gcgaaacccg acaggactat
     2521 aaagatacca ggcgtttccc cctggcggct ccctcgtgcg ctctcctgtt cctgcctttc
     2581 ggtttaccgg tgtcattccg ctgttatggc cgcgtttgtc tcattccacg cctgacactc
     2641 agttccgggt aggcagttcg ctccaagctg gactgtatgc acgaaccccc cgttcagtcc
     2701 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggaaag acatgcaaaa
     2761 gcaccactgg cagcagccac tggtaattga tttagaggag ttagtcttga agtcatgcgc
     2821 cggttaaggc taaactgaaa ggacaagttt tggtgactgc gctcctccaa gccagttacc
     2881 tcggttcaaa gagttggtag ctcagagaac cttcgaaaaa ccgccctgca aggcggtttt
     2941 ttcgttttca gagcaagaga ttacgcgcag accaaaacga tctcaagaag atcatcttat
     3001 taatcagata aaatatttct agatttcagt gcaatttatc tcttcaaatg tagcacctga
     3061 agtcagcccc atacgatata agttgttact agtgcttgga ttctcaccaa taaaaaacgc
     3121 ccggcggcaa ccgagcgttc tgaacaaatc cagatggagt tctgaggtca ttactggatc
     3181 tatcaacagg agtccaagcg agctcgatat caaattacgc cccgccctgc cactcatcgc
     3241 agtactgttg taattcatta agcattctgc cgacatggaa gccatcacaa acggcatgat
     3301 gaacctgaat cgccagcggc atcagcacct tgtcgccttg cgtataatat ttgcccatgg
     3361 tgaaaacggg ggcgaagaag ttgtccatat tggccacgtt taaatcaaaa ctggtgaaac
     3421 tcacccaggg attggctgag acgaaaaaca tattctcaat aaacccttta gggaaatagg
     3481 ccaggttttc accgtaacac gccacatctt gcgaatatat gtgtagaaac tgccggaaat
     3541 cgtcgtggta ttcactccag agcgatgaaa acgtttcagt ttgctcatgg aaaacggtgt
     3601 aacaagggtg aacactatcc catatcacca gctcaccgtc tttcattgcc atacgaaatt
     3661 ccggatgagc attcatcagg cgggcaagaa tgtgaataaa ggccggataa aacttgtgct
     3721 tatttttctt tacggtcttt aaaaaggccg taatatccag ctgaacggtc tggttatagg
     3781 tacattgagc aactgactga aatgcctcaa aatgttcttt acgatgccat tgggatatat
     3841 caacggtggt atatccagtg atttttttct ccattttagc ttccttagct cctgaaaatc
     3901 tcgataactc aaaaaatacg cccggtagtg atcttatttc attatggtga aagttggaac
     3961 ctcttacgtg ccgatcaacg tctcattttc gccagatatc gacgtcttaa gacccacttt
     4021 cacatttaag ttgtttttct aatccgcata tgatcaattc aaggccgaat aagaaggctg
     4081 gctctgcacc ttggtgatca aataattcga tagcttgtcg taataatggc ggcatactat
     4141 cagtagtagg tgtttccctt tcttctttag cgacttgatg ctcttgatct tccaatacgc
     4201 aacctaaagt aaaatgcccc acagcgctga gtgcatataa tgcattctct agtgaaaaac
     4261 cttgttggca taaaaaggct aattgatttt cgagagtttc atactgtttt tctgtaggcc
     4321 gtgtacctaa atgtactttt gctccatcgc gatgacttag taaagcacat ctaaaacttt
     4381 tagcgttatt acgtaaaaaa tcttgccagc tttccccttc taaagggcaa aagtgagtat
     4441 ggtgcctatc taacatctca atggctaagg cgtcgagcaa agcccgctta ttttttacat
     4501 gccaatacaa tgtaggctgc tctacaccta gcttctgggc gagtttacgg gttgttaaac
     4561 cttcgattcc gacctcatta agcagctcta atgcgctgtt aatcact
//