LOCUS       sequence-162820-        3898 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses CRISPR array containing 3 copies of Desulfovibrio vulgaris
            spacer 2 and flanking repeats in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3898)
  AUTHORS   Hochstrasser ML, Taylor DW, Kornfeld JE, Nogales E, Doudna JA
  TITLE     DNA Targeting by a Minimal CRISPR RNA-Guided Cascade.
  JOURNAL   Mol Cell. 2016 Sep 1;63(5):840-51. doi: 
            10.1016/j.molcel.2016.07.027.
  PUBMED    27588603
REFERENCE   2  (bases 1 to 3898)
  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: 
            "10.1016/j.molcel.2016.07"; journalName: "Mol Cell"; date: 
            "2016-09-1- 1"; volume: "63"; issue: "5"; pages: "840-51"
COMMENT     Sequence Label: D. vulgaris sp2 CRISPR/pACYCDuet-1
FEATURES             Location/Qualifiers
     source          1..3898
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(138..1220)
                     /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     1182..1201
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1221..1298)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1340..1359
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1340..1358
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1614..1658
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(1696..1714)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1710..1757
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(1978..2637)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     2552..2571
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(2638..2740)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(3132..3149)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3266..3811)
                     /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(3346..3365)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgatataagt tgtaattctc atgttagtca tgccccgcgc ccaccggaag gagctgactg
       61 ggttgaaggc tctcaagggc atcggtcgag atcccggtgc ctaatgagtg agctaactta
      121 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
      181 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc cagggtggtt
      241 tttcttttca ccagtgagac gggcaacagc tgattgccct tcaccgcctg gccctgagag
      301 agttgcagca agcggtccac gctggtttgc cccagcaggc gaaaatcctg tttgatggtg
      361 gttaacggcg ggatataaca tgagctgtct tcggtatcgt cgtatcccac taccgagatg
      421 tccgcaccaa cgcgcagccc ggactcggta atggcgcgca ttgcgcccag cgccatctga
      481 tcgttggcaa ccagcatcgc agtgggaacg atgccctcat tcagcatttg catggtttgt
      541 tgaaaaccgg acatggcact ccagtcgcct tcccgttccg ctatcggctg aatttgattg
      601 cgagtgagat atttatgcca gccagccaga cgcagacgcg ccgagacaga acttaatggg
      661 cccgctaaca gcgcgatttg ctggtgaccc aatgcgacca gatgctccac gcccagtcgc
      721 gtaccgtctt catgggagaa aataatactg ttgatgggtg tctggtcaga gacatcaaga
      781 aataacgccg gaacattagt gcaggcagct tccacagcaa tggcatcctg gtcatccagc
      841 ggatagttaa tgatcagccc actgacgcgt tgcgcgagaa gattgtgcac cgccgcttta
      901 caggcttcga cgccgcttcg ttctaccatc gacaccacca cgctggcacc cagttgatcg
      961 gcgcgagatt taatcgccgc gacaatttgc gacggcgcgt gcagggccag actggaggtg
     1021 gcaacgccaa tcagcaacga ctgtttgccc gccagttgtt gtgccacgcg gttgggaatg
     1081 taattcagct ccgccatcgc cgcttccact ttttcccgcg ttttcgcaga aacgtggctg
     1141 gcctggttca ccacgcggga aacggtctga taagagacac cggcatactc tgcgacatcg
     1201 tataacgtta ctggtttcac attcaccacc ctgaattgac tctcttccgg gcgctatcat
     1261 gccataccgc gaaaggtttt gcgccattcg atggtgtccg ggatctcgac gctctccctt
     1321 atgcgactcc tgcattaggt aatacgactc actataggga gagtcgcccc ccacgcgggg
     1381 gcgtggattg aaacgccatg ctcaggctgg cgagtgcgcc actcatcaag tcgcccccca
     1441 cgcgggggcg tggattgaaa cgccatgctc aggctggcga gtgcgccact catcaagtcg
     1501 ccccccacgc gggggcgtgg attgaaacgc catgctcagg ctggcgagtg cgccactcat
     1561 caagtcgccc cccacgcggg ggcgtggatt gaaacggtac cctcgagtct ggtaaagaaa
     1621 ccgctgctgc gaaatttgaa cgccagcaca tggactcgtc tactagcgca gcttaattaa
     1681 cctaggctgc tgccaccgct gagcaataac tagcataacc ccttggggcc tctaaacggg
     1741 tcttgagggg ttttttgctg aaacctcagg catttgagaa gcacacggtc acactgcttc
     1801 cggtagtcaa taaaccggta aaccagcaat agacataagc ggctatttaa cgaccctgcc
     1861 ctgaaccgac gaccgggtcg aatttgcttt cgaatttctg ccattcatcc gcttattatc
     1921 acttattcag gcgtagcaac caggcgttta agggcaccaa taactgcctt aaaaaaatta
     1981 cgccccgccc tgccactcat cgcagtactg ttgtaattca ttaagcattc tgccgacatg
     2041 gaagccatca caaacggcat gatgaacctg aatcgccagc ggcatcagca ccttgtcgcc
     2101 ttgcgtataa tatttgccca tagtgaaaac gggggcgaag aagttgtcca tattggccac
     2161 gtttaaatca aaactggtga aactcaccca gggattggct gagacgaaaa acatattctc
     2221 aataaaccct ttagggaaat aggccaggtt ttcaccgtaa cacgccacat cttgcgaata
     2281 tatgtgtaga aactgccgga aatcgtcgtg gtattcactc cagagcgatg aaaacgtttc
     2341 agtttgctca tggaaaacgg tgtaacaagg gtgaacacta tcccatatca ccagctcacc
     2401 gtctttcatt gccatacgga actccggatg agcattcatc aggcgggcaa gaatgtgaat
     2461 aaaggccgga taaaacttgt gcttattttt ctttacggtc tttaaaaagg ccgtaatatc
     2521 cagctgaacg gtctggttat aggtacattg agcaactgac tgaaatgcct caaaatgttc
     2581 tttacgatgc cattgggata tatcaacggt ggtatatcca gtgatttttt tctccatttt
     2641 agcttcctta gctcctgaaa atctcgataa ctcaaaaaat acgcccggta gtgatcttat
     2701 ttcattatgg tgaaagttgg aacctcttac gtgccgatca acgtctcatt ttcgccaaaa
     2761 gttggcccag ggcttcccgg tatcaacagg gacaccagga tttatttatt ctgcgaagtg
     2821 atcttccgtc acaggtattt attcggcgca aagtgcgtcg ggtgatgctg ccaacttact
     2881 gatttagtgt atgatggtgt ttttgaggtg ctccagtggc ttctgtttct atcagctgtc
     2941 cctcctgttc agctactgac ggggtggtgc gtaacggcaa aagcaccgcc ggacatcagc
     3001 gctagcggag tgtatactgg cttactatgt tggcactgat gagggtgtca gtgaagtgct
     3061 tcatgtggca ggagaaaaaa ggctgcaccg gtgcgtcagc agaatatgtg atacaggata
     3121 tattccgctt cctcgctcac tgactcgcta cgctcggtcg ttcgactgcg gcgagcggaa
     3181 atggcttacg aacggggcgg agatttcctg gaagatgcca ggaagatact taacagggaa
     3241 gtgagagggc cgcggcaaag ccgtttttcc ataggctccg cccccctgac aagcatcacg
     3301 aaatctgacg ctcaaatcag tggtggcgaa acccgacagg actataaaga taccaggcgt
     3361 ttccccctgg cggctccctc gtgcgctctc ctgttcctgc ctttcggttt accggtgtca
     3421 ttccgctgtt atggccgcgt ttgtctcatt ccacgcctga cactcagttc cgggtaggca
     3481 gttcgctcca agctggactg tatgcacgaa ccccccgttc agtccgaccg ctgcgcctta
     3541 tccggtaact atcgtcttga gtccaacccg gaaagacatg caaaagcacc actggcagca
     3601 gccactggta attgatttag aggagttagt cttgaagtca tgcgccggtt aaggctaaac
     3661 tgaaaggaca agttttggtg actgcgctcc tccaagccag ttacctcggt tcaaagagtt
     3721 ggtagctcag agaaccttcg aaaaaccgcc ctgcaaggcg gttttttcgt tttcagagca
     3781 agagattacg cgcagaccaa aacgatctca agaagatcat cttattaatc agataaaata
     3841 tttctagatt tcagtgcaat ttatctcttc aaatgtagca cctgaagtca gccccata
//
