LOCUS       sequence-222899-        7476 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses TtCsm6 from a SUMO-tagged bacterial expression construct.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7476)
  AUTHORS   Gootenberg JS, Abudayyeh OO, Kellner MJ, Joung J, Collins JJ, Zhang 
            F
  TITLE     Multiplexed and portable nucleic acid detection platform with Cas13,
            Cas12a, and Csm6.
  JOURNAL   Science. 2018 Apr 27;360(6387):439-444. doi: 
            10.1126/science.aaq0179. Epub 2018 Feb 15.
  PUBMED    29449508
REFERENCE   2  (bases 1 to 7476)
  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: 
            "10.1126/science.aaq0179"; journalName: "Science"; date: 
            "2018-04-27- 27"; volume: "360"; issue: "6387"; pages: "439-444"
COMMENT     Sequence Label: pC0075	TtCsm6  His6-TwinStrep-SUMO-BsaI
FEATURES             Location/Qualifiers
     source          1..7476
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1..25
                     /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             40..62
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             81..98
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             108..125
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             join(138..161,162..197,198..221)
                     /codon_start=1
                     /product="two Strep-Tag(R)II moieties connected by a linker
                     for enhanced binding to Strep-Tactin(R), an engineered form
                     of streptavidin (Schmidt et al., 2013)"
                     /label=Twin-Strep-tag
                     /translation="WSHPQFEKGGGSGGGSGGSAWSHPQFEK"
     CDS             222..515
                     /codon_start=1
                     /gene="S. cerevisiae SMT3 (truncated)"
                     /product="cleavable ubiquitin-like protein tag"
                     /label=SUMO
                     /translation="MSDSEVNQEAKPEVKPEVKPETHINLKVSDGSSEIFFKIKKTTPL
                     RRLMEAFAKRQGKEMDSLRFLYDGIRIQADQTPEDLDMEDNDIIEAHREQIGG"
     primer_bind     complement(1958..1976)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1972..2019
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(2194..2199,2200..2216,2217..2222))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(2249..2267)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2335..2439
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2440..2508,2509..3300)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2658..2677)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3471..4059
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3960..3979
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4213..4230
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4245..4384
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4310..4329)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4470..4492
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4486..4677)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5989..7071)
                     /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     7033..7052
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(7072..7149)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     7355..7374
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     join(7458..7476,1)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        7458..7476
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ggaattgtga gcggataaca attcccctct agaaataatt ttgtttaact ttaagaagga
       61 gatataccat gggcagcagc catcatcatc atcatcacag cagcggcctg gtgccgcgcg
      121 gcagccatat ggctagctgg agccatccgc agtttgaaaa aggtggtggt agcggtggtg
      181 gttcaggtgg tagtgcatgg tcacaccctc agtttgagaa aatgtcggac tcagaagtca
      241 atcaagaagc taagccagag gtcaagccag aagtcaagcc tgagactcac atcaatttaa
      301 aggtgtccga tggatcttca gagatcttct tcaagatcaa aaagaccact cctttaagaa
      361 ggctgatgga agcgttcgct aaaagacagg gtaaggaaat ggactcctta agattcttgt
      421 acgacggtat tagaatccaa gctgatcaga cccctgaaga tttggacatg gaggataacg
      481 atattattga ggctcacaga gaacagattg gtggatctat ggaggacctt gatgcgttgt
      541 gggagcgcta tcgtgaagct gttcgcgcgg gtgggaatcc ccaggctctg taccaggaga
      601 tggtatggcc ggcgttgtta gcactgtggc gcgagaaacc tcgcgtttac cccttccctc
      661 aagcatttgc ggtgtcagtg catacccttg gaacgagccc ggaggcaact gctcttgcga
      721 tcttaggtgc aggcgccgag cgcgtttatg tgttacacac tccggagtca gcgcgcttct
      781 tgccgcgctt acgtcaagac accggaaagg acttgtaccc agttgagatt ggaaagagtg
      841 acgtagaggc gatctatcgt gaagtgaagc gcttattgga aaagcacccc gaagtccctg
      901 tcgcattgga cctgacttcg gggaccaagg caatgtcggc agggttggcc gccgcaggat
      961 tttttttcca acgtttttat cccaaagtcc gtgtggtgta tgtagataac gaagactatg
     1021 acccagaatt gcgccgcccg cgcgccggga cggagaaact tcgcattttg cccaatccac
     1081 acgaggcttt ggctgaagtc gacgcattgt ttgccaaaga gctttatggc aaaggagaat
     1141 ttgggcaggc ggcggcttac tttcgcggaa tggttggtcg cactggtaat caagcatacg
     1201 cattatatgc attgctggcg gagatgtacc gcgcctggcg cgccttggat ttcggtgaag
     1261 cattaaaggc ggggcgtaag ctgctgggac aactgtctca aaatgtgtgg ttgaatcatc
     1321 cactgaacgc ccgtcgcgaa gcgcttgagg cgcaggtcgc attattagag gccgtcgacc
     1381 gctttttgaa ggcgcgtgac ttcgcattaa aggaaggcgt gtatggtctg gcccgcactc
     1441 ttcttcatct ggcccaagag gcgaaagaag aagccgcggt acttgcagca ttgtatgcat
     1501 accgcgcatt ggagcttctt ttacaggagc gcttagctct gcttggtcgt cgcgcggagg
     1561 cgcctggtct gagcccagaa gaagccgagg cattgcgcaa ggctttagcc gagctgttag
     1621 gagttcttcc tgaagaggtt cgtcttccag ctaaattagg tttattagat ttattggcct
     1681 ttttgcgctt gaaaggagat gaggcgttgg gccgcctgtc tctggctgaa ctgcgtggct
     1741 tagccggtgc tcttaaaggc cgtaattcag cccttcttgt acatgggttt gatgtgccca
     1801 gtcctaaggc cgtcgagggc atcgcccgtc tggcgcaagg tcttctgcag gatcttgagg
     1861 cacgtacggc tctgggcccg ctgtctccgg aacccgtacc cttggggttc taaggccgca
     1921 ctcgaggccc gaaaggaagc tgagttggct gctgccaccg ctgagcaata actagcataa
     1981 ccccttgggg cctctaaacg ggtcttgagg ggttttttgc tgaaaggagg aactatatcc
     2041 ggatatcccg caagaggccc ggcagtaccg gcataaccaa gcctatgcct acagcatcca
     2101 gggtgacggt gccgaggatg acgatgagcg cattgttaga tttcatacac ggtgcctgac
     2161 tgcgttagca atttaactgt gataaactac cgcattaaag cttatcgatg ataagctgtc
     2221 aaacatgaga attcttgaag acgaaagggc ctcgtgatac gcctattttt ataggttaat
     2281 gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga
     2341 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     2401 ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt
     2461 gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg
     2521 ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg
     2581 gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg
     2641 agcactttta aagttctgct atgtggcgcg gtattatccc gtgttgacgc cgggcaagag
     2701 caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca
     2761 gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg
     2821 agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc
     2881 gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg
     2941 aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgcagcaat ggcaacaacg
     3001 ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca attaatagac
     3061 tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg
     3121 tttattgctg ataaatctgg agccggtgag cgtgggtcac gcggtatcat tgcagcactg
     3181 gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact
     3241 atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa
     3301 ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca tttttaattt
     3361 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     3421 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     3481 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     3541 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     3601 cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt caagaactct
     3661 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     3721 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     3781 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     3841 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     3901 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     3961 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     4021 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     4081 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     4141 gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga
     4201 acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcctgat gcggtatttt
     4261 ctccttacgc atctgtgcgg tatttcacac cgcaatggtg cactctcagt acaatctgct
     4321 ctgatgccgc atagttaagc cagtatacac tccgctatcg ctacgtgact gggtcatggc
     4381 tgcgccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc
     4441 atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc
     4501 gtcatcaccg aaacgcgcga ggcagctgcg gtaaagctca tcagcgtggt cgtgaagcga
     4561 ttcacagatg tctgcctgtt catccgcgtc cagctcgttg agtttctcca gaagcgttaa
     4621 tgtctggctt ctgataaagc gggccatgtt aagggcggtt ttttcctgtt tggtcactga
     4681 tgcctccgtg taagggggat ttctgttcat gggggtaatg ataccgatga aacgagagag
     4741 gatgctcacg atacgggtta ctgatgatga acatgcccgg ttactggaac gttgtgaggg
     4801 taaacaactg gcggtatgga tgcggcggga ccagagaaaa atcactcagg gtcaatgcca
     4861 gcgcttcgtt aatacagatg taggtgttcc acagggtagc cagcagcatc ctgcgatgca
     4921 gatccggaac ataatggtgc agggcgctga cttccgcgtt tccagacttt acgaaacacg
     4981 gaaaccgaag accattcatg ttgttgctca ggtcgcagac gttttgcagc agcagtcgct
     5041 tcacgttcgc tcgcgtatcg gtgattcatt ctgctaacca gtaaggcaac cccgccagcc
     5101 tagccgggtc ctcaacgaca ggagcacgat catgcgcacc cgtggccagg acccaacgct
     5161 gcccgagatg cgccgcgtgc ggctgctgga gatggcggac gcgatggata tgttctgcca
     5221 agggttggtt tgcgcattca cagttctccg caagaattga ttggctccaa ttcttggagt
     5281 ggtgaatccg ttagcgaggt gccgccggct tccattcagg tcgaggtggc ccggctccat
     5341 gcaccgcgac gcaacgcggg gaggcagaca aggtataggg cggcgcctac aatccatgcc
     5401 aacccgttcc atgtgctcgc cgaggcggca taaatcgccg tgacgatcag cggtccaatg
     5461 atcgaagtta ggctggtaag agccgcgagc gatccttgaa gctgtccctg atggtcgtca
     5521 tctacctgcc tggacagcat ggcctgcaac gcgggcatcc cgatgccgcc ggaagcgaga
     5581 agaatcataa tggggaaggc catccagcct cgcgtcgcga acgccagcaa gacgtagccc
     5641 agcgcgtcgg ccgccatgcc ggcgataatg gcctgcttct cgccgaaacg tttggtggcg
     5701 ggaccagtga cgaaggcttg agcgagggcg tgcaagattc cgaataccgc aagcgacagg
     5761 ccgatcatcg tcgcgctcca gcgaaagcgg tcctcgccga aaatgaccca gagcgctgcc
     5821 ggcacctgtc ctacgagttg catgataaag aagacagtca taagtgcggc gacgatagtc
     5881 atgccccgcg cccaccggaa ggagctgact gggttgaagg ctctcaaggg catcggtcga
     5941 gatcccggtg cctaatgagt gagctaactt acattaattg cgttgcgctc actgcccgct
     6001 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     6061 ggcggtttgc gtattgggcg ccagggtggt ttttcttttc accagtgaga cgggcaacag
     6121 ctgattgccc ttcaccgcct ggccctgaga gagttgcagc aagcggtcca cgctggtttg
     6181 ccccagcagg cgaaaatcct gtttgatggt ggttaacggc gggatataac atgagctgtc
     6241 ttcggtatcg tcgtatccca ctaccgagat atccgcacca acgcgcagcc cggactcggt
     6301 aatggcgcgc attgcgccca gcgccatctg atcgttggca accagcatcg cagtgggaac
     6361 gatgccctca ttcagcattt gcatggtttg ttgaaaaccg gacatggcac tccagtcgcc
     6421 ttcccgttcc gctatcggct gaatttgatt gcgagtgaga tatttatgcc agccagccag
     6481 acgcagacgc gccgagacag aacttaatgg gcccgctaac agcgcgattt gctggtgacc
     6541 caatgcgacc agatgctcca cgcccagtcg cgtaccgtct tcatgggaga aaataatact
     6601 gttgatgggt gtctggtcag agacatcaag aaataacgcc ggaacattag tgcaggcagc
     6661 ttccacagca atggcatcct ggtcatccag cggatagtta atgatcagcc cactgacgcg
     6721 ttgcgcgaga agattgtgca ccgccgcttt acaggcttcg acgccgcttc gttctaccat
     6781 cgacaccacc acgctggcac ccagttgatc ggcgcgagat ttaatcgccg cgacaatttg
     6841 cgacggcgcg tgcagggcca gactggaggt ggcaacgcca atcagcaacg actgtttgcc
     6901 cgccagttgt tgtgccacgc ggttgggaat gtaattcagc tccgccatcg ccgcttccac
     6961 tttttcccgc gttttcgcag aaacgtggct ggcctggttc accacgcggg aaacggtctg
     7021 ataagagaca ccggcatact ctgcgacatc gtataacgtt actggtttca cattcaccac
     7081 cctgaattga ctctcttccg ggcgctatca tgccataccg cgaaaggttt tgcgccattc
     7141 gatggtgtcc gggatctcga cgctctccct tatgcgactc ctgcattagg aagcagccca
     7201 gtagtaggtt gaggccgttg agcaccgccg ccgcaaggaa tggtgcatgc aaggagatgg
     7261 cgcccaacag tcccccggcc acggggcctg ccaccatacc cacgccgaaa caagcgctca
     7321 tgagcccgaa gtggcgagcc cgatcttccc catcggtgat gtcggcgata taggcgccag
     7381 caaccgcacc tgtggcgccg gtgatgccgg ccacgatgcg tccggcgtag aggatcgaga
     7441 tctcgatccc gcgaaattaa tacgactcac tatagg
//
