LOCUS       sequence-384623-        5728 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes Cas4/1, leader, repeat and one spacer from A. 
            acidoterrestris (type V-B).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5728)
  AUTHORS   Wu WY, Jackson SA, Almendros C, Haagsma AC, Yilmaz S, Gort G, van 
            der Oost J, Brouns SJJ, Staals RHJ
  TITLE     Adaptation by Type V-A and V-B CRISPR-Cas Systems Demonstrates 
            Conserved Protospacer Selection Mechanisms Between Diverse 
            CRISPR-Cas Types.
  JOURNAL   CRISPR J. 2022 Aug;5(4):536-547. doi: 10.1089/crispr.2021.0150. Epub
            2022 Jul 12.
  PUBMED    35833800
REFERENCE   2  (bases 1 to 5728)
  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"; doi: 
            "10.1089/crispr.2021.0150"; journalName: "CRISPR J"; date: 
            "2022-08"; volume: "5"; issue: "4"; pages: "536-547"
COMMENT     Sequence Label: pCas-4/1-Delta-2_VB
FEATURES             Location/Qualifiers
     source          1..5728
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     526..545
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(565..642)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     688..707
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        688..706
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    711..734
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     RBS             787..809
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     primer_bind     complement(3035..3053)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3049..3096
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(3267..4058)
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     promoter        complement(4059..4150)
                     /gene="bla"
                     /label=AmpR promoter
     CDS             complement(join(5210..5728,1..564))
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=lacI
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
ORIGIN
        1 tgggcccgct aacagcgcga tttgctggtg acccaatgcg accagatgct ccacgcccag
       61 tcgcgtaccg tcttcatggg agaaaataat actgttgatg ggtgtctggt cagagacatc
      121 aagaaataac gccggaacat tagtgcaggc agcttccaca gcaatggcat cctggtcatc
      181 cagcggatag ttaatgatca gcccactgac gcgttgcgcg agaagattgt gcaccgccgc
      241 tttacaggct tcgacgccgc ttcgttctac catcgacacc accacgctgg cacccagttg
      301 atcggcgcga gatttaatcg ccgcgacaat ttgcgacggc gcgtgcaggg ccagactgga
      361 ggtggcaacg ccaatcagca acgactgttt gcccgccagt tgttgtgcca cgcggttggg
      421 aatgtaattc agctccgcca tcgccgcttc cactttttcc cgcgttttcg cagaaacgtg
      481 gctggcctgg ttcaccacgc gggaaacggt ctgataagag acaccggcat actctgcgac
      541 atcgtataac gttactggtt tcacattcac caccctgaat tgactctctt ccgggcgcta
      601 tcatgccata ccgcgaaagg ttttgcgcca ttcgatggtg tccgggatct cgacgctctc
      661 ccttatgcga ctcctgcatt aggaaattaa tacgactcac tatagggaga gaattgtgag
      721 cggataacaa ttcccctgta gattaattaa gcggccgccc tgcaggactc gagttctaga
      781 aataattttg tttaacttta agaaggagat atacatatgg ccgtaccgat tcggatgctg
      841 aatgaactct cttattgcga acgtttatat catttgatgc acgtgcaagg gttattcgaa
      901 aaatctgcgg acacagtgga aggcagcggt cagcacgacc gggcggaacg ccgcaaaaga
      961 ccatcagacg tggggcatga tgtgtgggat attgcaccgc agagcctcca tctcggcgac
     1021 gaggatttgg ggattgtggg caaattggat gcgattcgct atcaggacga cgggtattgg
     1081 gagcccgttg aagcgaaaca ttcaagcgca ccaaacggtg gcacttcatt tcgcctcggc
     1141 gagtgggaat tggacggcaa ggcgtggccg aatgaccaga tacagttgtg tgcacaagga
     1201 ttgttactga gagccaacgg atttccgacc attgcgggta agttgttcta ccgaggaaac
     1261 agacagacgg taaggattgt attttcggat gatttgattg ctgcaacggt tgccaccatc
     1321 cggagagcac acgagttgga acaggcttcg atgccctcgc cactgctgaa ttccgacaag
     1381 tgcttccgat gctcgttgaa tggtgtctgt cttcctgacg aaacgattcg cttgacgagt
     1441 ccggacggag agaccgttcc gatacgagat attgtcccca gtcgagatga tttaggaacc
     1501 gtgtacgtgt cggagcctgg gacccgggtc ggaaagcgtg gttacgagtt gaccatcacg
     1561 tcgaaagatg ggatggaatc gaccgttccg ttaaaagatg ttcggcacat ctccctcttc
     1621 gggaacgttc agatttccac ccaattggtg cacgattgta tggagtccgg cgtcaccatc
     1681 agctacttga ctgccgccgg ccgtctcgtc gggatgaacc acaatttgat ttcgaaaaac
     1741 gtcttggtgc ggcgtgaaca gttcaagcgg tttggggacg agcgtgatcg actgacactt
     1801 gcgaggtata tcgttcgcgc gaaaattctc aaccagcgca ccttgcttcg tcgaaatgcg
     1861 cacgggctcg acaaggtcat tttgaaagat ttgctcgatc acgcccgacg ttccgaagaa
     1921 gccacttcca ttgactcgtt gctcggcatt gaagggattg cggcgaaaca gtacatgcag
     1981 gcgtttgtgc tgatgctgaa agtaaaaggc gttcctgcgg gcgaaacgct gatgaacggc
     2041 agaaatcgcc gcccaccgaa ggacccggtt aatgcgttgc tgtcacttgg atattcgttg
     2101 ttgcaacggg atatgtatgc cgccttggct gcagtgggcc tggacccctt gatgggcttt
     2161 taccatcgca ttgaaacggg gcgtccgtca ctggtactcg acatgatgga accgtttcga
     2221 gcgatcatcg cggattcggt ggtcgttcga acattgaaca ccggcgaaat tgcctgggat
     2281 gacttttata tcgggcagga gtcctgttcg ttgaagctgc ctggacggaa acgatttttt
     2341 caggcgtttg agcgacgcat gcacgagacc gtgacgcatc cggtgttcgg ttacaaattg
     2401 agttaccgtc ggatgctgga actagaagcg cgcttcttgt cgcgatactt gatgggcgaa
     2461 ttgccggaat atcgcccaat tgtcacgagg tgatggcgat gcgccgatat gtcgaggatt
     2521 gatgtaaact tcaaatacgc gtaaaatata atctgtctag aggacagaat ttttcaacgg
     2581 gtgtgccaat ggccactttc caggtggcaa agcccgttga gcttctcaaa aagaacgctc
     2641 gctcagtgtt ctgacctttc gagcgcctgt tcagggcgaa aaccctggga ggcgctcgaa
     2701 tcataggtgg gacaagggat tcgcggcgaa aacgacggga gagaacgaga atgaaacgag
     2761 atgaaatcga tgggaaacat caagtgctcg tttatgggct tggatctccg cgtggttcaa
     2821 gggctgcgag cgcccgctgt cggatcactg agcgagcgat ctgagaagtg gcactgtttg
     2881 gtaaaggtaa aaagacgaat gatgcatcct aacattggaa gtggataacg gatccgcgat
     2941 cgcggcgcgc cacctggtgg ccggccggta ccacgcgtgc gcgctgatcc ggctgctaac
     3001 aaagcccgaa aggaagctga gttggctgct gccaccgctg agcaataact agcataaccc
     3061 cttggggcct ctaaacgggt cttgaggggt tttttgctga aacctcaggc atttgagaag
     3121 cacacggtca cactgcttcc ggtagtcaat aaaccggtaa accagcaata gacataagcg
     3181 gctatttaac gaccctgccc tgaaccgacg accgggtcat cgtggccgga tcttgcggcc
     3241 cctcggcttg aacgaattgt tagacattat ttgccgacta ccttggtgat ctcgcctttc
     3301 acgtaatgga caaattcttc caactgatct gcgcgcgagg ccaagcgatc ttcttcttgt
     3361 ccaagataag cctgtctagc ttcaagtatg acgggctgat actgggccgg caggcgctcc
     3421 attgcccagt cggcagcgac atccttcggc gcgattttgc cggttactgc gctgtaccaa
     3481 atgcgggaca acgtaagcac tacatttcgc tcatcgccag cccagtcggg cggcgagttc
     3541 catagcgtta aggtttcatt tagcgcctca aatagatcct gttcaggaac cggatcaaag
     3601 agttcctccg ccgctggacc taccaaggca acgctatgtt ctcttgcttt tgtcagcaag
     3661 atagccagat caatgtcgat cgtggctggc tcgaagatac ctgcaagaat gtcattgcgc
     3721 tgccattctc caaattgcag ttcgcgctta gctggataac gccacggaat gatgtcgtcg
     3781 tgcacaacaa tggtgacttc tacagcgcgg agaatctcgc tctctccagg ggaagccgaa
     3841 gtttccaaaa ggtcgttgat caaagctcgc cgcgttgttt catcaagcct tacggtcacc
     3901 gtaaccagca aatcaatatc actgtgcggc ttcaggccgc catccactgc ggagccgtac
     3961 aaatgtacgg ccagcaacgt cggttcgaga tggcgctcga tgacgccaac tacctctgat
     4021 agttgagtcg atacttcggc gatcaccgct tccctcatac tcttcctttt tcattattat
     4081 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
     4141 aataaacaaa tagctagctc actcggtcgc tacgctccgg gcgtgagact gcggcgggcg
     4201 ctgcggacac atacaaagtt acccacagat tccgtggata agcaggggac taacatgtga
     4261 ggcaaaacag cagggccgcg ccggtggcgt ttttccatag gctccgccct cctgccagag
     4321 ttcacataaa cagacgcttt tccggtgcat ctgtgggagc cgtgaggctc aaccatgaat
     4381 ctgacagtac gggcgaaacc cgacaggact taaagatccc caccgtttcc ggcgggtcgc
     4441 tccctcttgc gctctcctgt tccgaccctg ccgtttaccg gatacctgtt ccgcctttct
     4501 cccttacggg aagtgtggcg ctttctcata gctcacacac tggtatctcg gctcggtgta
     4561 ggtcgttcgc tccaagctgg gctgtaagca agaactcccc gttcagcccg actgctgcgc
     4621 cttatccggt aactgttcac ttgagtccaa cccggaaaag cacggtaaaa cgccactggc
     4681 agcagccatt ggtaactggg agttcgcaga ggatttgttt agctaaacac gcggttgctc
     4741 ttgaagtgtg cgccaaagtc cggctacact ggaaggacag atttggttgc tgtgctctgc
     4801 gaaagccagt taccacggtt aagcagttcc ccaactgact taaccttcga tcaaaccacc
     4861 tccccaggtg gttttttcga aggttaagtc agttggggaa ctgcttaacc ttcgatcaaa
     4921 ccacctcccc aggtggtttt ttcgtttaca gggcaaaaga ttacgcgcag aaaaaaagga
     4981 tctcaagaag atcctttgat cttttctact gaaccgctct tatttcagtg caatttatct
     5041 cttcaaatgt agcacctgaa gtcagcccca tacgatataa gttgtaattc tcatgttagt
     5101 catgccccgc gcccaccgga aggagctgac tgggttgaag gctctcaagg gcatcggtcg
     5161 agatcccggt gcctaatgag tgagctaact tacattaatt gcgttgcgct cactgcccgc
     5221 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     5281 aggcggtttg cgtattgggc gccagggtgg tttttctttt caccagtgag acgggcaaca
     5341 gctgattgcc cttcaccgcc tggccctgag agagttgcag caagcggtcc acgctggttt
     5401 gccccagcag gcgaaaatcc tgtttgatgg tggttaacgg cgggatataa catgagctgt
     5461 cttcggtatc gtcgtatccc actaccgaga tgtccgcacc aacgcgcagc ccggactcgg
     5521 taatggcgcg cattgcgccc agcgccatct gatcgttggc aaccagcatc gcagtgggaa
     5581 cgatgccctc attcagcatt tgcatggttt gttgaaaacc ggacatggca ctccagtcgc
     5641 cttcccgttc cgctatcggc tgaatttgat tgcgagtgag atatttatgc cagccagcca
     5701 gacgcagacg cgccgagaca gaacttaa
//
