LOCUS       sequence-387293-        5407 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes Cas4, Cas1, 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 5407)
  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 5407)
  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_Delta-2_VA
FEATURES             Location/Qualifiers
     source          1..5407
                     /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(3421..3439)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3435..3482
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(3653..4444)
                     /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(4445..4536)
                     /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 ttatgacgaa aattgcaaac cttagtcttt atgttaaaat
     3121 aactactaag ttcttagaga tatttaaaaa tatgactgtt gttatatatc aaaatgctaa
     3181 aaaaatcata gattttaggt ctttttttgc tgatttaggc aaaaacgggt ctaagaactt
     3241 taaataattt ctactgttgt agatgagaag tcatttaata aggccactgt taaaataaca
     3301 ttggaagtgg ataacggatc cgcgatcgcg gcgcgccacc tggtggccgg ccggtaccac
     3361 gcgtgcgcgc tgatccggct gctaacaaag cccgaaagga agctgagttg gctgctgcca
     3421 ccgctgagca ataactagca taaccccttg gggcctctaa acgggtcttg aggggttttt
     3481 tgctgaaacc tcaggcattt gagaagcaca cggtcacact gcttccggta gtcaataaac
     3541 cggtaaacca gcaatagaca taagcggcta tttaacgacc ctgccctgaa ccgacgaccg
     3601 ggtcatcgtg gccggatctt gcggcccctc ggcttgaacg aattgttaga cattatttgc
     3661 cgactacctt ggtgatctcg cctttcacgt aatggacaaa ttcttccaac tgatctgcgc
     3721 gcgaggccaa gcgatcttct tcttgtccaa gataagcctg tctagcttca agtatgacgg
     3781 gctgatactg ggccggcagg cgctccattg cccagtcggc agcgacatcc ttcggcgcga
     3841 ttttgccggt tactgcgctg taccaaatgc gggacaacgt aagcactaca tttcgctcat
     3901 cgccagccca gtcgggcggc gagttccata gcgttaaggt ttcatttagc gcctcaaata
     3961 gatcctgttc aggaaccgga tcaaagagtt cctccgccgc tggacctacc aaggcaacgc
     4021 tatgttctct tgcttttgtc agcaagatag ccagatcaat gtcgatcgtg gctggctcga
     4081 agatacctgc aagaatgtca ttgcgctgcc attctccaaa ttgcagttcg cgcttagctg
     4141 gataacgcca cggaatgatg tcgtcgtgca caacaatggt gacttctaca gcgcggagaa
     4201 tctcgctctc tccaggggaa gccgaagttt ccaaaaggtc gttgatcaaa gctcgccgcg
     4261 ttgtttcatc aagccttacg gtcaccgtaa ccagcaaatc aatatcactg tgcggcttca
     4321 ggccgccatc cactgcggag ccgtacaaat gtacggccag caacgtcggt tcgagatggc
     4381 gctcgatgac gccaactacc tctgatagtt gagtcgatac ttcggcgatc accgcttccc
     4441 tcatactctt cctttttcat tattattgaa gcatttatca gggttattgt ctcatgagcg
     4501 gatacatatt tgaatgtatt tagaaaaata aacaaatagc tagctcactc ggtcgctacg
     4561 ctccgggcgt gagactgcgg cgggcgctgc ggacacatac aaagttaccc acagattccg
     4621 tggataagca ggggactaac atgtgaggca aaacagcagg gccgcgccgg tggcgttttt
     4681 ccataggctc cgccctcctg ccagagttca cataaacaga cgcttttccg gtgcatctgt
     4741 gggagccgtg aggctcaacc atgaatctga cagtacgggc gaaacccgac aggacttaaa
     4801 gatccccacc gtttccggcg ggtcgctccc tcttgcgctc tcctgttccg accctgccgt
     4861 ttaccggata cctgttccgc ctttctccct tacgggaagt gtggcgcttt ctcatagctc
     4921 acacactggt atctcggctc ggtgtaggtc gttcgctcca agctgggctg taagcaagaa
     4981 ctccccgttc agcccgactg ctgcgcctta tccggtaact gttcacttga gtccaacccg
     5041 gaaaagcacg gtaaaacgcc actggcagca gccattggta actgggagtt cgcagaggat
     5101 ttgtttagct aaacacgcgg ttgctcttga agtgtgcgcc aaagtccggc tacactggaa
     5161 ggacagattt ggttgctgtg ctctgcgaaa gccagttacc acggttaagc agttccccaa
     5221 ctgacttaac cttcgatcaa accacctccc caggtggttt tttcgaaggt taagtcagtt
     5281 ggggaactgc ttaaccttcg atcaaaccac ctccccaggt ggttttttcg tttacagggc
     5341 aaaagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctactgaac
     5401 cgctctt
//
