LOCUS       sequence-275332-        5606 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  T7 RNAP-driven expression of a non-cognate SNIPR trigger RNA with a 
            mutation at position 33.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5606)
  AUTHORS   Hong F, Ma D, Wu K, Mina LA, Luiten RC, Liu Y, Yan H, Green AA
  TITLE     Precise and Programmable Detection of Mutations Using Ultraspecific 
            Riboregulators.
  JOURNAL   Cell. 2020 Feb 25. pii: S0092-8674(20)30155-0. doi: 
            10.1016/j.cell.2020.02.011.
  PUBMED    32109416
REFERENCE   2  (bases 1 to 5606)
  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"; journalName: "Cell. 2020 
            Feb 25. pii: S0092-8674(20)30155-0. doi: 
            10.1016/j.cell.2020.02.011."
COMMENT     Sequence Label: p_SNIPR_target_GUmut33AtoG
FEATURES             Location/Qualifiers
     source          1..5606
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      83..129
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(304..309,310..326,327..332))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(359..377)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        445..549
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(550..618,619..1410)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAQPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(768..787)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1581..2169
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2070..2089
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2323..2340
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2355..2494
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2420..2439)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2580..2602
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2596..2787)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4099..5181)
                     /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
                     EACKAAVHNLLAQRVSGLIINYPLYDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     5143..5162
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5182..5259)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5465..5484
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     join(5590..5606,1..3)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        join(5590..5606,1..2)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 gggtgaatat cctattcgat attcacaata tcgcacttac ctaccatgtc aatcacttcg
       61 tcttattcat ttcattctat cttagcataa ccccttgggg cctctaaacg ggtcttgagg
      121 ggttttttgc tgaaaggagg aactatatcc ggatatcccg caagaggccc ggcagtaccg
      181 gcataaccaa gcctatgcct acagcatcca gggtgacggt gccgaggatg acgatgagcg
      241 cattgttaga tttcatacac ggtgcctgac tgcgttagca atttaactgt gataaactac
      301 cgcattaaag cttatcgatg ataagctgtc aaacatgaga attcttgaag acgaaagggc
      361 ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca
      421 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
      481 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
      541 aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt
      601 tgccttcctg tttttgctca accagaaacg ctggtgaaag taaaagatgc tgaagatcag
      661 ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt
      721 tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg
      781 gtattatccc gtgttgacgc cgggcaagag caactcggtc gccgcataca ctattctcag
      841 aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta
      901 agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg
      961 acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta
     1021 actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac
     1081 accacgatgc ctgcagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt
     1141 actctagctt cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca
     1201 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
     1261 cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta
     1321 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag
     1381 ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt
     1441 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
     1501 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     1561 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     1621 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     1681 tttccgaagg taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag
     1741 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     1801 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     1861 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     1921 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     1981 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     2041 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     2101 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     2161 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     2221 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt
     2281 gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag
     2341 gaagcggaag agcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac
     2401 cgcaatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatacac
     2461 tccgctatcg ctacgtgact gggtcatggc tgcgccccga cacccgccaa cacccgctga
     2521 cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc
     2581 cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga ggcagctgcg
     2641 gtaaagctca tcagcgtggt cgtgaagcga ttcacagatg tctgcctgtt catccgcgtc
     2701 cagctcgttg agtttctcca gaagcgttaa tgtctggctt ctgataaagc gggccatgtt
     2761 aagggcggtt ttttcctgtt tggtcactga tgcctccgtg taagggggat ttctgttcat
     2821 gggggtaatg ataccgatga aacgagagag gatgctcacg atacgggtta ctgatgatga
     2881 acatgcccgg ttactggaac gttgtgaggg taaacaactg gcggtatgga tgcggcggga
     2941 ccagagaaaa atcactcagg gtcaatgcca gcgcttcgtt aatacagatg taggtgttcc
     3001 acagggtagc cagcagcatc ctgcgatgca gatccggaac ataatggtgc agggcgctga
     3061 cttccgcgtt tccagacttt acgaaacacg gaaaccgaag accattcatg ttgttgctca
     3121 ggtcgcagac gttttgcagc agcagtcgct tcacgttcgc tcgcgtatcg gtgattcatt
     3181 ctgctaacca gtaaggcaac cccgccagcc tagccgggtc ctcaacgaca ggagcacgat
     3241 catgcgcacc cgtggccagg acccaacgct gcccgagatg cgccgcgtgc ggctgctgga
     3301 gatggcggac gcgatggata tgttctgcca agggttggtt tgcgcattca cagttctccg
     3361 caagaattga ttggctccaa ttcttggagt ggtgaatccg ttagcgaggt gccgccggct
     3421 tccattcagg tcgaggtggc ccggctccat gcaccgcgac gcaacgcggg gaggcagaca
     3481 aggtataggg cggcgcctac aatccatgcc aacccgttcc atgtgctcgc cgaggcggca
     3541 taaatcgccg tgacgatcag cggtccaatg atcgaagtta ggctggtaag agccgcgagc
     3601 gatccttgaa gctgtccctg atggtcgtca tctacctgcc tggacagcat ggcctgcaac
     3661 gcgggcatcc cgatgccgcc ggaagcgaga agaatcataa tggggaaggc catccagcct
     3721 cgcgtcgcga acgccagcaa gacgtagccc agcgcgtcgg ccgccatgcc ggcgataatg
     3781 gcctgcttct cgccgaaacg tttggtggcg ggaccagtga cgaaggcttg agcgagggcg
     3841 tgcaagattc cgaataccgc aagcgacagg ccgatcatcg tcgcgctcca gcgaaagcgg
     3901 tcctcgccga aaatgaccca gagcgctgcc ggcacctgtc ctacgagttg catgataaag
     3961 aagacagtca taagtgcggc gacgatagtc atgccccgcg cccaccggaa ggagctgact
     4021 gggttgaagg ctctcaaggg catcggtcga gatcccggtg cctaatgagt gagctaactt
     4081 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     4141 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ccagggtggt
     4201 ttttcttttc accagtgaga cgggcaacag ctgattgccc ttcaccgcct ggccctgaga
     4261 gagttgcagc aagcggtcca cgctggtttg ccccagcagg cgaaaatcct gtttgatggt
     4321 ggttaacggc gggatataac atgagctgtc ttcggtatcg tcgtatccca ctaccgagat
     4381 atccgcacca acgcgcagcc cggactcggt aatggcgcgc attgcgccca gcgccatctg
     4441 atcgttggca accagcatcg cagtgggaac gatgccctca ttcagcattt gcatggtttg
     4501 ttgaaaaccg gacatggcac tccagtcgcc ttcccgttcc gctatcggct gaatttgatt
     4561 gcgagtgaga tatttatgcc agccagccag acgcagacgc gccgagacag aacttaatgg
     4621 gcccgctaac agcgcgattt gctggtgacc caatgcgacc agatgctcca cgcccagtcg
     4681 cgtaccgtct tcatgggaga aaataatact gttgatgggt gtctggtcag agacatcaag
     4741 aaataacgcc ggaacattag tgcaggcagc ttccacagca atggcatcct ggtcatacag
     4801 cggatagtta atgatcagcc cactgacgcg ttgcgcgaga agattgtgca ccgccgcttt
     4861 acaggcttcg acgccgcttc gttctaccat cgacaccacc acgctggcac ccagttgatc
     4921 ggcgcgagat ttaatcgccg cgacaatttg cgacggcgcg tgcagggcca gactggaggt
     4981 ggcaacgcca atcagcaacg actgtttgcc cgccagttgt tgtgccacgc ggttgggaat
     5041 gtaattcagc tccgccatcg ccgcttccac tttttcccgc gttttcgcag aaacgtggct
     5101 ggcctggttc accacgcggg aaacggtctg ataagagaca ccggcatact ctgcgacatc
     5161 gtataacgtt actggtttca cattcaccac cctgaattga ctctcttccg ggcgctatca
     5221 tgccataccg cgaaaggttt tgcgccattc gatggtgtcc gggatctcga cgctctccct
     5281 tatgcgactc ctgcattagg aagcagccca gtagtaggtt gaggccgttg agcaccgccg
     5341 ccgcaaggaa tggtgcatgc aaggagatgg cgcccaacag tcccccggcc acggggcctg
     5401 ccaccatacc cacgccgaaa caagcgctca tgagcccgaa gtggcgagcc cgatcttccc
     5461 catcggtgat gtcggcgata taggcgccag caaccgcacc tgtggcgccg gtgatgccgg
     5521 ccacgatgcg tccggcgtag aggatcgaga tctcgggcga gctactcgtt tgcgtagaag
     5581 tagttgcgct aatacgactc actata
//
