LOCUS       sequence-325307-        4360 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Constitutive expression of single-spacer CRISPR array with spacer #2
            targeting deGFP mRNA for BzCas13b in bacteria..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4360)
  AUTHORS   Wandera KG, Beisel CL
  TITLE     Rapidly Characterizing CRISPR-Cas13 Nucleases Using Cell-Free 
            Transcription-Translation Systems.
  JOURNAL   Methods Mol Biol. 2022;2404:135-153. doi: 
            10.1007/978-1-0716-1851-6_7.
  PUBMED    34694607
REFERENCE   2  (bases 1 to 4360)
  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: "Methods Mol
            Biol."; date: "2022"; volume: "2404:135-153. doi"; pages: " 
            10.1007/978-1-0716-1851-6_7"
COMMENT     Sequence Label: pBzCas13b-gRNA-2
FEATURES             Location/Qualifiers
     source          1..4360
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(1..6,7..23,24..29)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     promoter        152..186
                     /label=J23119 promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119)"
     primer_bind     complement(544..563)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     CDS             complement(2416..3075)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     2990..3009
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(3076..3178)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(3570..3587)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3704..4249)
                     /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(3784..3803)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttgacagctt atcatcgata agctttaatg cggtagttta tcacagttaa attgctaacg
       61 cagtcaggca ccgtgtatga aatctaacaa tgcgctcatc gtcatcctcg gcaccgtcac
      121 cctggatgct gtaggcatag gcttggttta attgacagct agctcagtcc taggtataat
      181 gctagcgctg agttggaact gctctcattt tggagggtaa tcacaacggt cggggtagcg
      241 gctgaagcac tgcacgcgtt ggaactgctc tcattttgga gggtaatcac aacatcgtcc
      301 attccgacag catcgccagt cactatggcg tgctgctagc gctatatgcg ttgatgcaat
      361 ttctatgcgc acccgttctc ggagcactgt ccgaccgctt tggccgccgc ccagtcctgc
      421 tcgcttcgct acttggagcc actatcgact acgcgatcat ggcgaccaca cccgtcctgt
      481 ggatcctcta cgccggacgc atcgtggccg gcatcaccgg cgccacaggt gcggttgctg
      541 gcgcctatat cgccgacatc accgatgggg aagatcgggc tcgccacttc gggctcatga
      601 gcgcttgttt cggcgtgggt atggtggcag gccccgtggc cgggggactg ttgggcgcca
      661 tctccttgca tgcaccattc cttgcggcgg cggtgctcaa cggcctcaac ctactactgg
      721 gctgcttcct aatgcaggag tcgcataagg gagagcgtcg accgatgccc ttgagagcct
      781 tcaacccagt cagctccttc cggtgggcgc ggggcatgac tatcgtcgcc gcacttatga
      841 ctgtcttctt tatcatgcaa ctcgtaggac aggtgccggc agcgctctgg gtcattttcg
      901 gcgaggaccg ctttcgctgg agcgcgacga tgatcggcct gtcgcttgcg gtattcggaa
      961 tcttgcacgc cctcgctcaa gccttcgtca ctggtcccgc caccaaacgt ttcggcgaga
     1021 agcaggccat tatcgccggc atggcggccg acgcgctggg ctacgtcttg ctggcgttcg
     1081 cgacgcgagg ctggatggcc ttccccatta tgattcttct cgcttccggc ggcatcggga
     1141 tgcccgcgtt gcaggccatg ctgtccaggc aggtagatga cgaccatcag ggacagcttc
     1201 aaggatcgct cgcggctctt accagcctaa cttcgatcat tggaccgctg atcgtcacgg
     1261 cgatttatgc cgcctcggcg agcacatgga acgggttggc atggattgta ggcgccgccc
     1321 tataccttgt ctgcctcccc gcgttgcgtc gcggtgcatg gagccgggcc acctcgacct
     1381 gaatggaagc cggcggcacc tcgctaacgg attcaccact ccaagaattg gagccaatca
     1441 attcttgcgg agaactgtga atgcgcaaac caacccttgg cagaacatat ccatcgcgtc
     1501 cgccatctcc agcagccgca cgcggcgcat ctcgggcagc gttgggtcct ggccacgggt
     1561 gcgcatgatc gtgctcctgt cgttgaggac ccggctaggc tggcggggtt gccttactgg
     1621 ttagcagaat gaatcaccga tacgcgagcg aacgtgaagc gactgctgct gcaaaacgtc
     1681 tgcgacctga gcaacaacat gaatggtctt cggtttccgt gtttcgtaaa gtctggaaac
     1741 gcggaagtcc cctacgtgct gctgaagttg cccgcaacag agagtggaac caaccggtga
     1801 taccacgata ctatgactga gagtcaacgc catgagcggc ctcatttctt attctgagtt
     1861 acaacagtcc gcaccgctgc cggtagctcc ttccggtggg cgcggggcat gactatcgtc
     1921 gccgcactta tgactgtctt ctttatcatg caactcgtag gacaggtgcc ggcagcgccc
     1981 aacagtcccc cggccacggg gcctgccacc atacccacgc cgaaacaagc gccctgcacc
     2041 attatgttcc ggatctgcat cgcaggatgc tgctggctac cctgtggaac acctacatct
     2101 gtattaacga agcgctaacc gtttttatca ggctctggga ggcagaataa atgatcatat
     2161 cgtcaattat tacctccacg gggagagcct gagcaaactg gcctcaggca tttgagaagc
     2221 acacggtcac actgcttccg gtagtcaata aaccggtaaa ccagcaatag acataagcgg
     2281 ctatttaacg accctgccct gaaccgacga ccgggtcgaa tttgctttcg aatttctgcc
     2341 attcatccgc ttattatcac ttattcaggc gtagcaacca ggcgtttaag ggcaccaata
     2401 actgccttaa aaaaattacg ccccgccctg ccactcatcg cagtactgtt gtaattcatt
     2461 aagcattctg ccgacatgga agccatcaca aacggcatga tgaacctgaa tcgccagcgg
     2521 catcagcacc ttgtcgcctt gcgtataata tttgcccatg gtgaaaacgg gggcgaagaa
     2581 gttgtccata ttggccacgt ttaaatcaaa actggtgaaa ctcacccagg gattggctga
     2641 gacgaaaaac atattctcaa taaacccttt agggaaatag gccaggtttt caccgtaaca
     2701 cgccacatct tgcgaatata tgtgtagaaa ctgccggaaa tcgtcgtggt attcactcca
     2761 gagcgatgaa aacgtttcag tttgctcatg gaaaacggtg taacaagggt gaacactatc
     2821 ccatatcacc agctcaccgt ctttcattgc catacggaat tccggatgag cattcatcag
     2881 gcgggcaaga atgtgaataa aggccggata aaacttgtgc ttatttttct ttacggtctt
     2941 taaaaaggcc gtaatatcca gctgaacggt ctggttatag gtacattgag caactgactg
     3001 aaatgcctca aaatgttctt tacgatgcca ttgggatata tcaacggtgg tatatccagt
     3061 gatttttttc tccattttag cttccttagc tcctgaaaat ctcgataact caaaaaatac
     3121 gcccggtagt gatcttattt cattatggtg aaagttggaa cctcttacgt gccgatcaac
     3181 gtctcatttt cgccaaaagt tggcccaggg cttcccggta tcaacaggga caccaggatt
     3241 tatttattct gcgaagtgat cttccgtcac aggtatttat tcggcgcaaa gtgcgtcggg
     3301 tgatgctgcc aacttactga tttagtgtat gatggtgttt ttgaggtgct ccagtggctt
     3361 ctgtttctat cagctgtccc tcctgttcag ctactgacgg ggtggtgcgt aacggcaaaa
     3421 gcaccgccgg acatcagcgc tagcggagtg tatactggct tactatgttg gcactgatga
     3481 gggtgtcagt gaagtgcttc atgtggcagg agaaaaaagg ctgcaccggt gcgtcagcag
     3541 aatatgtgat acaggatata ttccgcttcc tcgctcactg actcgctacg ctcggtcgtt
     3601 cgactgcggc gagcggaaat ggcttacgaa cggggcggag atttcctgga agatgccagg
     3661 aagatactta acagggaagt gagagggccg cggcaaagcc gtttttccat aggctccgcc
     3721 cccctgacaa gcatcacgaa atctgacgct caaatcagtg gtggcgaaac ccgacaggac
     3781 tataaagata ccaggcgttt ccccctggcg gctccctcgt gcgctctcct gttcctgcct
     3841 ttcggtttac cggtgtcatt ccgctgttat ggccgcgttt gtctcattcc acgcctgaca
     3901 ctcagttccg ggtaggcagt tcgctccaag ctggactgta tgcacgaacc ccccgttcag
     3961 tccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccgga aagacatgca
     4021 aaagcaccac tggcagcagc cactggtaat tgatttagag gagttagtct tgaagtcatg
     4081 cgccggttaa ggctaaactg aaaggacaag ttttggtgac tgcgctcctc caagccagtt
     4141 acctcggttc aaagagttgg tagctcagag aaccttcgaa aaaccgccct gcaaggcggt
     4201 tttttcgttt tcagagcaag agattacgcg cagaccaaaa cgatctcaag aagatcatct
     4261 tattaatcag ataaaatatt tctagatttc agtgcaattt atctcttcaa atgtagcacc
     4321 tgaagtcagc cccatacgat ataagttgta attctcatgt
//
