LOCUS       sequence-370319-        7595 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes Cas12a from F. novicida (type V-A).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7595)
  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 7595)
  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: pCas12a
FEATURES             Location/Qualifiers
     source          1..7595
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     950..969
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(989..1066)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1112..1131
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1112..1130
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1131..1155
                     /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)."
     primer_bind     complement(5161..5179)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      5175..5222
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(5443..6102)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     6017..6036
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(6103..6205)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(6597..6614)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6731..7276)
                     /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(6811..6830)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7501..7595,1..988))
                     /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 gggtggtttt tcttttcacc agtgagacgg gcaacagctg attgcccttc accgcctggc
       61 cctgagagag ttgcagcaag cggtccacgc tggtttgccc cagcaggcga aaatcctgtt
      121 tgatggtggt taacggcggg atataacatg agctgtcttc ggtatcgtcg tatcccacta
      181 ccgagatgtc cgcaccaacg cgcagcccgg actcggtaat ggcgcgcatt gcgcccagcg
      241 ccatctgatc gttggcaacc agcatcgcag tgggaacgat gccctcattc agcatttgca
      301 tggtttgttg aaaaccggac atggcactcc agtcgccttc ccgttccgct atcggctgaa
      361 tttgattgcg agtgagatat ttatgccagc cagccagacg cagacgcgcc gagacagaac
      421 ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa tgcgaccaga tgctccacgc
      481 ccagtcgcgt accgtcttca tgggagaaaa taatactgtt gatgggtgtc tggtcagaga
      541 catcaagaaa taacgccgga acattagtgc aggcagcttc cacagcaatg gcatcctggt
      601 catccagcgg atagttaatg atcagcccac tgacgcgttg cgcgagaaga ttgtgcaccg
      661 ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga caccaccacg ctggcaccca
      721 gttgatcggc gcgagattta atcgccgcga caatttgcga cggcgcgtgc agggccagac
      781 tggaggtggc aacgccaatc agcaacgact gtttgcccgc cagttgttgt gccacgcggt
      841 tgggaatgta attcagctcc gccatcgccg cttccacttt ttcccgcgtt ttcgcagaaa
      901 cgtggctggc ctggttcacc acgcgggaaa cggtctgata agagacaccg gcatactctg
      961 cgacatcgta taacgttact ggtttcacat tcaccaccct gaattgactc tcttccgggc
     1021 gctatcatgc cataccgcga aaggttttgc gccattcgat ggtgtccggg atctcgacgc
     1081 tctcccttat gcgactcctg cattaggaaa ttaatacgac tcactatagg ggaattgtga
     1141 gcggataaca attcccctgt agaaataatt ttgtttaact ttaataagga gatataccat
     1201 ggatgtcaat ttatcaagaa tttgttaata aatatagttt aagtaaaact ctaagatttg
     1261 agttaatccc acagggtaaa acacttgaaa acataaaagc aagaggtttg attttagatg
     1321 atgagaaaag agctaaagac tacaaaaagg ctaaacaaat aattgataaa tatcatcagt
     1381 tttttataga ggagatatta agttcggttt gtattagcga agatttatta caaaactatt
     1441 ctgatgttta ttttaaactt aaaaagagtg atgatgataa tctacaaaaa gattttaaaa
     1501 gtgcaaaaga tacgataaag aaacaaatat ctgaatatat aaaggactca gagaaattta
     1561 agaatttgtt taatcaaaac cttatcgatg ctaaaaaagg gcaagagtca gatttaattc
     1621 tatggctaaa gcaatctaag gataatggta tagaactatt taaagccaat agtgatatca
     1681 cagatataga tgaggcgtta gaaataatca aatcttttaa aggttggaca acttatttta
     1741 agggttttca tgaaaataga aaaaatgttt atagtagcaa tgatattcct acatctatta
     1801 tttataggat agtagatgat aatttgccta aatttctaga aaataaagct aagtatgaga
     1861 gtttaaaaga caaagctcca gaagctataa actatgaaca aattaaaaaa gatttggcag
     1921 aagagctaac ctttgatatt gactacaaaa catctgaagt taatcaaaga gttttttcac
     1981 ttgatgaagt ttttgagata gcaaacttta ataattatct aaatcaaagt ggtattacta
     2041 aatttaatac tattattggt ggtaaatttg taaatggtga aaatacaaag agaaaaggta
     2101 taaatgaata tataaatcta tactcacagc aaataaatga taaaacactc aaaaaatata
     2161 aaatgagtgt tttatttaag caaattttaa gtgatacaga atctaaatct tttgtaattg
     2221 ataagttaga agatgatagt gatgtagtta caacgatgca aagtttttat gagcaaatag
     2281 cagcttttaa aacagtagaa gaaaaatcta ttaaagaaac actatcttta ttatttgatg
     2341 atttaaaagc tcaaaaactt gatttgagta aaatttattt taaaaatgat aaatctctta
     2401 ctgatctatc acaacaagtt tttgatgatt atagtgttat tggtacagcg gtactagaat
     2461 atataactca acaaatagca cctaaaaatc ttgataaccc tagtaagaaa gagcaagaat
     2521 taatagccaa aaaaactgaa aaagcaaaat acttatctct agaaactata aagcttgcct
     2581 tagaagaatt taataagcat agagatatag ataaacagtg taggtttgaa gaaatacttg
     2641 caaactttgc ggctattccg atgatatttg atgaaatagc tcaaaacaaa gacaatttgg
     2701 cacagatatc tatcaaatat caaaatcaag gtaaaaaaga cctacttcaa gctagtgcgg
     2761 aagatgatgt taaagctatc aaggatcttt tagatcaaac taataatctc ttacataaac
     2821 taaaaatatt tcatattagt cagtcagaag ataaggcaaa tattttagac aaggatgagc
     2881 atttttatct agtatttgag gagtgctact ttgagctagc gaatatagtg cctctttata
     2941 acaaaattag aaactatata actcaaaagc catatagtga tgagaaattt aagctcaatt
     3001 ttgagaactc gactttggct aatggttggg ataaaaataa agagcctgac aatacggcaa
     3061 ttttatttat caaagatgat aaatattatc tgggtgtgat gaataagaaa aataacaaaa
     3121 tatttgatga taaagctatc aaagaaaata aaggcgaggg ttataaaaaa attgtttata
     3181 aacttttacc tggcgcaaat aaaatgttac ctaaggtttt cttttctgct aaatctataa
     3241 aattttataa tcctagtgaa gatatactta gaataagaaa tcattccaca catacaaaaa
     3301 atggtagtcc tcaaaaagga tatgaaaaat ttgagtttaa tattgaagat tgccgaaaat
     3361 ttatagattt ttataaacag tctataagta agcatccgga gtggaaagat tttggattta
     3421 gattttctga tactcaaaga tataattcta tagatgaatt ttatagagaa gttgaaaatc
     3481 aaggctacaa actaactttt gaaaatatat cagagagcta tattgatagc gtagttaatc
     3541 agggtaaatt gtacctattc caaatctata ataaagattt ttcagcttat agcaaagggc
     3601 gaccaaatct acatacttta tattggaaag cgctgtttga tgagagaaat cttcaagatg
     3661 tggtttataa gctaaatggt gaggcagagc ttttttatcg taaacaatca atacctaaaa
     3721 aaatcactca cccagctaaa gaggcaatag ctaataaaaa caaagataat cctaaaaaag
     3781 agagtgtttt tgaatatgat ttaatcaaag ataaacgctt tactgaagat aagtttttct
     3841 ttcactgtcc tattacaatc aattttaaat ctagtggagc taataagttt aatgatgaaa
     3901 tcaatttatt gctaaaagaa aaagcaaatg atgttcatat attaagtata gatagaggtg
     3961 aaagacattt agcttactat actttggtag atggtaaagg caatatcatc aaacaagata
     4021 ctttcaacat cattggtaat gatagaatga aaacaaacta ccatgataag cttgctgcaa
     4081 tagagaaaga tagggattca gctaggaaag actggaaaaa gataaataac atcaaagaga
     4141 tgaaagaggg ctatctatct caggtagttc atgaaatagc taagctagtt atagagtata
     4201 atgctattgt ggtttttgag gatttaaatt ttggatttaa aagagggcgt ttcaaggtag
     4261 agaagcaggt ctatcaaaag ttagaaaaaa tgctaattga gaaactaaac tatctagttt
     4321 tcaaagataa tgagtttgat aaaactgggg gagtgcttag agcttatcag ctaacagcac
     4381 cttttgagac ttttaaaaag atgggtaaac aaacaggtat tatctactat gtaccagctg
     4441 gttttacttc aaaaatttgt cctgtaactg gttttgtaaa tcagttatat cctaagtatg
     4501 aaagtgtcag caaatctcaa gagttcttta gtaagtttga caagatttgt tataaccttg
     4561 ataagggcta ttttgagttt agttttgatt ataaaaactt tggtgacaag gctgccaaag
     4621 gcaagtggac tatagctagc tttgggagta gattgattaa ctttagaaat tcagataaaa
     4681 atcataattg ggatactcga gaagtttatc caactaaaga gttggagaaa ttgctaaaag
     4741 attattctat cgaatatggg catggcgaat gtatcaaagc agctatttgc ggtgagagcg
     4801 acaaaaagtt ttttgctaag ctaactagtg tcctaaatac tatcttacaa atgcgtaact
     4861 caaaaacagg tactgagtta gattatctaa tttcaccagt agcagatgta aatggcaatt
     4921 tctttgattc gcgacaggcg ccaaaaaata tgcctcaaga tgctgatgcc aatggtgctt
     4981 atcatattgg gctaaaaggt ctgatgctac taggtaggat caaaaataat caagagggca
     5041 aaaaactcaa tttggttatc aaaaatgaag agtattttga gttcgtgcag aataggaata
     5101 actaacccag agaattcgag ctcggcgcgc ctgcaggtcg acaagcttgc gctgctgcca
     5161 ccgctgagca ataactagca taaccccttg gggcctctaa acgggtcttg aggggttttt
     5221 tgctgaaacc tcaggcattt gagaagcaca cggtcacact gcttccggta gtcaataaac
     5281 cggtaaacca gcaatagaca taagcggcta tttaacgacc ctgccctgaa ccgacgaccg
     5341 ggtcgaattt gctttcgaat ttctgccatt catccgctta ttatcactta ttcaggcgta
     5401 gcaaccaggc gtttaagggc accaataact gccttaaaaa aattacgccc cgccctgcca
     5461 ctcatcgcag tactgttgta attcattaag cattctgccg acatggaagc catcacaaac
     5521 ggcatgatga acctgaatcg ccagcggcat cagcaccttg tcgccttgcg tataatattt
     5581 gcccatagtg aaaacggggg cgaagaagtt gtccatattg gccacgttta aatcaaaact
     5641 ggtgaaactc acccagggat tggctgagac gaaaaacata ttctcaataa accctttagg
     5701 gaaataggcc aggttttcac cgtaacacgc cacatcttgc gaatatatgt gtagaaactg
     5761 ccggaaatcg tcgtggtatt cactccagag cgatgaaaac gtttcagttt gctcatggaa
     5821 aacggtgtaa caagggtgaa cactatccca tatcaccagc tcaccgtctt tcattgccat
     5881 acggaactcc ggatgagcat tcatcaggcg ggcaagaatg tgaataaagg ccggataaaa
     5941 cttgtgctta tttttcttta cggtctttaa aaaggccgta atatccagct gaacggtctg
     6001 gttataggta cattgagcaa ctgactgaaa tgcctcaaaa tgttctttac gatgccattg
     6061 ggatatatca acggtggtat atccagtgat ttttttctcc attttagctt ccttagctcc
     6121 tgaaaatctc gataactcaa aaaatacgcc cggtagtgat cttatttcat tatggtgaaa
     6181 gttggaacct cttacgtgcc gatcaacgtc tcattttcgc caaaagttgg cccagggctt
     6241 cccggtatca acagggacac caggatttat ttattctgcg aagtgatctt ccgtcacagg
     6301 tatttattcg gcgcaaagtg cgtcgggtga tgctgccaac ttactgattt agtgtatgat
     6361 ggtgtttttg aggtgctcca gtggcttctg tttctatcag ctgtccctcc tgttcagcta
     6421 ctgacggggt ggtgcgtaac ggcaaaagca ccgccggaca tcagcgctag cggagtgtat
     6481 actggcttac tatgttggca ctgatgaggg tgtcagtgaa gtgcttcatg tggcaggaga
     6541 aaaaaggctg caccggtgcg tcagcagaat atgtgataca ggatatattc cgcttcctcg
     6601 ctcactgact cgctacgctc ggtcgttcga ctgcggcgag cggaaatggc ttacgaacgg
     6661 ggcggagatt tcctggaaga tgccaggaag atacttaaca gggaagtgag agggccgcgg
     6721 caaagccgtt tttccatagg ctccgccccc ctgacaagca tcacgaaatc tgacgctcaa
     6781 atcagtggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggcggct
     6841 ccctcgtgcg ctctcctgtt cctgcctttc ggtttaccgg tgtcattccg ctgttatggc
     6901 cgcgtttgtc tcattccacg cctgacactc agttccgggt aggcagttcg ctccaagctg
     6961 gactgtatgc acgaaccccc cgttcagtcc gaccgctgcg ccttatccgg taactatcgt
     7021 cttgagtcca acccggaaag acatgcaaaa gcaccactgg cagcagccac tggtaattga
     7081 tttagaggag ttagtcttga agtcatgcgc cggttaaggc taaactgaaa ggacaagttt
     7141 tggtgactgc gctcctccaa gccagttacc tcggttcaaa gagttggtag ctcagagaac
     7201 cttcgaaaaa ccgccctgca aggcggtttt ttcgttttca gagcaagaga ttacgcgcag
     7261 accaaaacga tctcaagaag atcatcttat taatcagata aaatatttct agatttcagt
     7321 gcaatttatc tcttcaaatg tagcacctga agtcagcccc atacgatata agttgtaatt
     7381 ctcatgttag tcatgccccg cgcccaccgg aaggagctga ctgggttgaa ggctctcaag
     7441 ggcatcggtc gagatcccgg tgcctaatga gtgagctaac ttacattaat tgcgttgcgc
     7501 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     7561 cgcgcgggga gaggcggttt gcgtattggg cgcca
//
