LOCUS       sequence-384634-        5657 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes Cas4, Cas1, Cas2, leader, repeat and one spacer from F. 
            Novicida (type V-A).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5657)
  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 5657)
  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: pCas4_1_2_VA
FEATURES             Location/Qualifiers
     source          1..5657
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(189..1271)
                     /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"
     primer_bind     1233..1252
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1272..1349)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1395..1414
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1395..1413
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1418..1441
                     /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             1494..1516
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     primer_bind     complement(3671..3689)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3685..3732
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(3903..4694)
                     /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(4695..4786)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 atttcagtgc aatttatctc ttcaaatgta gcacctgaag tcagccccat acgatataag
       61 ttgtaattct catgttagtc atgccccgcg cccaccggaa ggagctgact gggttgaagg
      121 ctctcaaggg catcggtcga gatcccggtg cctaatgagt gagctaactt acattaattg
      181 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
      241 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ccagggtggt ttttcttttc
      301 accagtgaga cgggcaacag ctgattgccc ttcaccgcct ggccctgaga gagttgcagc
      361 aagcggtcca cgctggtttg ccccagcagg cgaaaatcct gtttgatggt ggttaacggc
      421 gggatataac atgagctgtc ttcggtatcg tcgtatccca ctaccgagat gtccgcacca
      481 acgcgcagcc cggactcggt aatggcgcgc attgcgccca gcgccatctg atcgttggca
      541 accagcatcg cagtgggaac gatgccctca ttcagcattt gcatggtttg ttgaaaaccg
      601 gacatggcac tccagtcgcc ttcccgttcc gctatcggct gaatttgatt gcgagtgaga
      661 tatttatgcc agccagccag acgcagacgc gccgagacag aacttaatgg gcccgctaac
      721 agcgcgattt gctggtgacc caatgcgacc agatgctcca cgcccagtcg cgtaccgtct
      781 tcatgggaga aaataatact gttgatgggt gtctggtcag agacatcaag aaataacgcc
      841 ggaacattag tgcaggcagc ttccacagca atggcatcct ggtcatccag cggatagtta
      901 atgatcagcc cactgacgcg ttgcgcgaga agattgtgca ccgccgcttt acaggcttcg
      961 acgccgcttc gttctaccat cgacaccacc acgctggcac ccagttgatc ggcgcgagat
     1021 ttaatcgccg cgacaatttg cgacggcgcg tgcagggcca gactggaggt ggcaacgcca
     1081 atcagcaacg actgtttgcc cgccagttgt tgtgccacgc ggttgggaat gtaattcagc
     1141 tccgccatcg ccgcttccac tttttcccgc gttttcgcag aaacgtggct ggcctggttc
     1201 accacgcggg aaacggtctg ataagagaca ccggcatact ctgcgacatc gtataacgtt
     1261 actggtttca cattcaccac cctgaattga ctctcttccg ggcgctatca tgccataccg
     1321 cgaaaggttt tgcgccattc gatggtgtcc gggatctcga cgctctccct tatgcgactc
     1381 ctgcattagg aaattaatac gactcactat agggagagaa ttgtgagcgg ataacaattc
     1441 ccctgtagat taattaagcg gccgccctgc aggactcgag ttctagaaat aattttgttt
     1501 aactttaaga aggagatata catatggaaa attatatttt gataacatgg ttaaatgatt
     1561 ttatattctg tccttactcg atatatttgc ataatatcta tagtaatgcc tcagatacta
     1621 catactattc atctagccaa acaaaagggc gcgatgctca taaaagtatc gataaaggaa
     1681 tctatagtac caaaaaagat gacctgatcg gtatcgatgt tattaaccat aaatatggtt
     1741 tggttggtaa aattgatgtt tttcataaag ataagggctt acttgtggag agaaaaaggc
     1801 aaatcaagac tatctatgat ggctataaat atcagcttta tgcgcaatat ttttgtctcc
     1861 aagagatggg ctatgatgtc aaagccatta aattttattc gatggttgat aataaatcat
     1921 acccaatagc tataccaact tcagctgagt tagaaaagtt tgaaaaacat attcaaacaa
     1981 tcaagcaata taatccaatg gataactcat ttaggcaaaa tattgaaaag tgtaaatttt
     2041 gtatatatgc aaacttatgt gataaaacgg acttgtagat tatgtttagt aaaaatgata
     2101 ttgaatcaaa gaatatagtt tttgttaata tttttgatgg agtgaaactt agtctatcat
     2161 tggggaatat agttataaaa gataaagaaa ctgatgaggt gaaaactaag ctttctgttc
     2221 ataaagttct tgcattgttt atcgtaggta atatgacgat gacctcgcaa cttttagaga
     2281 cctgtaagaa aaatgctata cagctagttt ttatgaaaaa tagctttaga ccatatctat
     2341 gttttggtga tattgctgag gctaattttt tagctagata taagcaatat agtgtagttg
     2401 agcaagatat aagtttagca aggattttta taacatcaaa gatacgcaat caacataact
     2461 tagtcaaaag cctaagagat aaaactccag agcagcaaga gatagtcaaa aagaataaac
     2521 agctaatagc agagttagaa aatacaacaa gcctagcgga gctaatgggt atagagggca
     2581 atgttgccaa aaatttcttc aaaggattct atggacattt agatagttgg caagggcgca
     2641 aacctagaat aaaacaggat ccatataatg ttgttttaga cttgggctat agtatgttgt
     2701 ttaattttgt agagtgtttt ttgcgacttt ttggctttga tttatacaag ggcttttgtc
     2761 atcagacttg gtataagcgt aaatccctag tttgtgactt tgttgagcca tttagatgta
     2821 tagtggataa ccaagttaga aaatcatgga atctcgggca attttctgta gaggattttg
     2881 gttgcaaaaa tgagcagttt tatataaaaa aagataaaac aaaagactac tcaaaaatac
     2941 tttttgccga gattatcagc tacaagctag agatatttga atatgtaaga gaattttatc
     3001 gtgcctttat gcgaggcaaa gaaattgcag agtatccaat attttgttat gaaactagga
     3061 gggtgtatgt tgatagtcag ttatgatttt agtaataata aagtacgtgc aaagtttgcc
     3121 aaatttctag aaagttatgg tgtacgttta caatattcgg tatttgagct caaatatagc
     3181 aagagaatgt tagacttgat tttagctgag atagaaaata actatgtacc actatttaca
     3241 aatgctgata gtgttttaat ctttaatgct ccagataaag atgtgataaa atatggttat
     3301 gcgattcata gagaacaaga ggttgttttt atagactaaa aattgcaaac cttagtcttt
     3361 atgttaaaat aactactaag ttcttagaga tatttaaaaa tatgactgtt gttatatatc
     3421 aaaatgctaa aaaaatcata gattttaggt ctttttttgc tgatttaggc aaaaacgggt
     3481 ctaagaactt taaataattt ctactgttgt agatgagaag tcatttaata aggccactgt
     3541 taaaataaca ttggaagtgg ataacggatc cgcgatcgcg gcgcgccacc tggtggccgg
     3601 ccggtaccac gcgtgcgcgc tgatccggct gctaacaaag cccgaaagga agctgagttg
     3661 gctgctgcca ccgctgagca ataactagca taaccccttg gggcctctaa acgggtcttg
     3721 aggggttttt tgctgaaacc tcaggcattt gagaagcaca cggtcacact gcttccggta
     3781 gtcaataaac cggtaaacca gcaatagaca taagcggcta tttaacgacc ctgccctgaa
     3841 ccgacgaccg ggtcatcgtg gccggatctt gcggcccctc ggcttgaacg aattgttaga
     3901 cattatttgc cgactacctt ggtgatctcg cctttcacgt aatggacaaa ttcttccaac
     3961 tgatctgcgc gcgaggccaa gcgatcttct tcttgtccaa gataagcctg tctagcttca
     4021 agtatgacgg gctgatactg ggccggcagg cgctccattg cccagtcggc agcgacatcc
     4081 ttcggcgcga ttttgccggt tactgcgctg taccaaatgc gggacaacgt aagcactaca
     4141 tttcgctcat cgccagccca gtcgggcggc gagttccata gcgttaaggt ttcatttagc
     4201 gcctcaaata gatcctgttc aggaaccgga tcaaagagtt cctccgccgc tggacctacc
     4261 aaggcaacgc tatgttctct tgcttttgtc agcaagatag ccagatcaat gtcgatcgtg
     4321 gctggctcga agatacctgc aagaatgtca ttgcgctgcc attctccaaa ttgcagttcg
     4381 cgcttagctg gataacgcca cggaatgatg tcgtcgtgca caacaatggt gacttctaca
     4441 gcgcggagaa tctcgctctc tccaggggaa gccgaagttt ccaaaaggtc gttgatcaaa
     4501 gctcgccgcg ttgtttcatc aagccttacg gtcaccgtaa ccagcaaatc aatatcactg
     4561 tgcggcttca ggccgccatc cactgcggag ccgtacaaat gtacggccag caacgtcggt
     4621 tcgagatggc gctcgatgac gccaactacc tctgatagtt gagtcgatac ttcggcgatc
     4681 accgcttccc tcatactctt cctttttcat tattattgaa gcatttatca gggttattgt
     4741 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagc tagctcactc
     4801 ggtcgctacg ctccgggcgt gagactgcgg cgggcgctgc ggacacatac aaagttaccc
     4861 acagattccg tggataagca ggggactaac atgtgaggca aaacagcagg gccgcgccgg
     4921 tggcgttttt ccataggctc cgccctcctg ccagagttca cataaacaga cgcttttccg
     4981 gtgcatctgt gggagccgtg aggctcaacc atgaatctga cagtacgggc gaaacccgac
     5041 aggacttaaa gatccccacc gtttccggcg ggtcgctccc tcttgcgctc tcctgttccg
     5101 accctgccgt ttaccggata cctgttccgc ctttctccct tacgggaagt gtggcgcttt
     5161 ctcatagctc acacactggt atctcggctc ggtgtaggtc gttcgctcca agctgggctg
     5221 taagcaagaa ctccccgttc agcccgactg ctgcgcctta tccggtaact gttcacttga
     5281 gtccaacccg gaaaagcacg gtaaaacgcc actggcagca gccattggta actgggagtt
     5341 cgcagaggat ttgtttagct aaacacgcgg ttgctcttga agtgtgcgcc aaagtccggc
     5401 tacactggaa ggacagattt ggttgctgtg ctctgcgaaa gccagttacc acggttaagc
     5461 agttccccaa ctgacttaac cttcgatcaa accacctccc caggtggttt tttcgaaggt
     5521 taagtcagtt ggggaactgc ttaaccttcg atcaaaccac ctccccaggt ggttttttcg
     5581 tttacagggc aaaagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     5641 tctactgaac cgctctt
//
