LOCUS       sequence-361529-        6606 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  encodes aTc inducible FsRT-Cas1–Cas2 expression cassette and 
            FsCRISPR Array 1 transcribed by BBa_J23103.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6606)
  AUTHORS   Schmidt F, Zimmermann J, Tanna T, Farouni R, Conway T, Macpherson 
            AJ, Platt RJ
  TITLE     Noninvasive assessment of gut function using transcriptional 
            recording sentinel cells.
  JOURNAL   Science. 2022 May 13;376(6594):eabm6038. doi: 
            10.1126/science.abm6038. Epub 2022 May 13.
  PUBMED    35549411
REFERENCE   2  (bases 1 to 6606)
  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: "Science."; 
            date: "2022-05-13"; volume: "376(6594):eabm6038. doi"; pages: " 
            10.1126/science.abm6038. Epub 2022 May 13"
COMMENT     Sequence Label: 
            pFS_1142_pET30_pCat-tetR-Term-ptetA-FsRT-Cas1(opt)-Cas2(opt)-3xTerm_
            J23103-Leader1-ARRAY1-DR1-FaqI_Term_NotI
FEATURES             Location/Qualifiers
     source          1..6606
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(220..1035)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(220..1035)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     943..962
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     1027..1046
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      1157..1745
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1646..1665
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1899..1916
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    1931..2070
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(1996..2015)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2156..2178
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2172..2363)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     RBS             2893..2904
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             2911..3534
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     complement(2984..3003)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     terminator      3579..3650
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3666..3693
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     protein_bind    3708..3726
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3733..3751
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     terminator      complement(6041..6084)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(6127..6145)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      6141..6188
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        6213..6247
                     /label=BBa_J23112
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0"
     promoter        6213..6247
                     /label=BBa_J23103
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.01"
ORIGIN
        1 tggctgaagc ggccgctatt cttttgattt ataagggatt ttgccgattt cggcctattg
       61 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgct
      121 tacaatttag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc
      181 taaatacatt caaatatgta tccgctcatg aattaattct tagaaaaact catcgagcat
      241 caaatgaaac tgcaatttat tcatatcagg attatcaata ccatattttt gaaaaagccg
      301 tttctgtaat gaaggagaaa actcaccgag gcagttccat aggatggcaa gatcctggta
      361 tcggtctgcg attccgactc gtccaacatc aatacaacct attaatttcc cctcgtcaaa
      421 aataaggtta tcaagtgaga aatcaccatg agtgacgact gaatccggtg agaatggcaa
      481 aagtttatgc atttctttcc agacttgttc aacaggccag ccattacgct cgtcatcaaa
      541 atcactcgca tcaaccaaac cgttattcat tcgtgattgc gcctgagcga gacgaaatac
      601 gcgatcgctg ttaaaaggac aattacaaac aggaatcgaa tgcaaccggc gcaggaacac
      661 tgccagcgca tcaacaatat tttcacctga atcaggatat tcttctaata cctggaatgc
      721 tgttttcccg gggatcgcag tggtgagtaa ccatgcatca tcaggagtac ggataaaatg
      781 cttgatggtc ggaagaggca taaattccgt cagccagttt agtctgacca tctcatctgt
      841 aacatcattg gcaacgctac ctttgccatg tttcagaaac aactctggcg catcgggctt
      901 cccatacaat cgatagattg tcgcacctga ttgcccgaca ttatcgcgag cccatttata
      961 cccatataaa tcagcatcca tgttggaatt taatcgcggc ctagagcaag acgtttcccg
     1021 ttgaatatgg ctcataacac cccttgtatt actgtttatg taagcagaca gttttattgt
     1081 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
     1141 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
     1201 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
     1261 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgtccttcta gtgtagccgt
     1321 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     1381 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     1441 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     1501 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     1561 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     1621 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
     1681 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
     1741 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggt cttttgctgg ccttttgctc
     1801 acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt
     1861 gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag
     1921 cggaagagcg cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca
     1981 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt atacactccg
     2041 ctatcgctac gtgactgggt catggctgcg ccccgacacc cgccaacacc cgctgacgcg
     2101 ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg
     2161 agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgaggca gctgcggtaa
     2221 agctcatcag cgtggtcgtg aagcgattca cagatgtctg cctgttcatc cgcgtccagc
     2281 tcgttgagtt tctccagaag cgttaatgtc tggcttctga taaagcgggc catgttaagg
     2341 gcggtttttt cctgtttggt cactgatgcc tccgtgtaag ggggatttct gttcatgggg
     2401 gtaatgatac cgatgaaacg agagaggatg ctcacgatac gggttactga tgatgaacat
     2461 gcccggttac tggaacgttg tgagggtaaa caactggcgg tatggatgcg gcgggaccag
     2521 agaaaaatca ctcagggtca atgccagcgc ttcgttaata cagatgtagg tgttccacag
     2581 ggtagccagc agcatcctgc gatgcagatc cggaacataa tggtgcaggg cgctgacttc
     2641 cgcgtttcca gactttacga aacacggaaa ccgaagatca ttcatgttgt tgctcaggtc
     2701 gcagacgttt tgcagcagca gtcgcttcac gttcgctcgc gtatcggtga ttcattctgc
     2761 taaccagtaa ggcaaccccg ccagcctagc cgggtcctca acgacaggag cacgatcatg
     2821 cgcacccgtg gggccgccat gccggcggca cgtaagaggt tccaactttc accataatga
     2881 aacatactag agaaagagga gaaatactag atgtccagat tagataaaag taaagtgatt
     2941 aacagcgcat tagagctgct taatgaggtc ggaatcgaag gtttaacaac ccgtaaactc
     3001 gcccagaagc taggtgtaga gcagcctaca ttgtattggc atgtaaaaaa taagcgggct
     3061 ttgctcgacg ccttagccat tgagatgtta gataggcacc atactcactt ttgcccttta
     3121 gaaggggaaa gctggcaaga ttttttacgt aataacgcta aaagttttag atgtgcttta
     3181 ctaagtcatc gcgatggagc aaaagtacat ttaggtacac ggcctacaga aaaacagtat
     3241 gaaactctcg aaaatcaatt agccttttta tgccaacaag gtttttcact agagaatgca
     3301 ttatatgcac tcagcgctgt ggggcatttt actttaggtt gcgtattgga agatcaagag
     3361 catcaagtcg ctaaagaaga aagggaaaca cctactactg atagtatgcc gccattatta
     3421 cgacaagcta tcgaattatt tgatcaccaa ggtgcagagc cagccttctt attcggcctt
     3481 gaattgatca tatgcggatt agaaaaacaa cttaaatgtg aaagtgggtc ctaataacac
     3541 tgatagtgct agtgtagatc actactagag ccaggcatca aataaaacga aaggctcagt
     3601 cgaaagactg ggcctttcgt tttatctgtt gtttgtcggt gaacgctctc tactagagtc
     3661 acactggctc accttcgggt gggcctttct gcgtttatat actagagtcc ctatcagtga
     3721 tagagattga catccctatc agtgatagag atactgagca ctactagaga agcttgcgga
     3781 ggagcatgca tgtttaccat cgacgagatg ctgagcaaga acaaccaacg tctggcgttc
     3841 gaacactttg cgaccaaaaa cgacggttgc ggcccggacg gtatgcacgt tagcgaactg
     3901 gagaagtact ggcgtatgaa tcacgaccag atcattagcg atctgaagaa ccaggaatac
     3961 cagccgggta ttatcctgat tcgtgagcac atgaataaaa ccggcaaacg ccgtaatatc
     4021 gcgagcctga acgttattga tcgtttcatt acccgtctgc tgagccagaa actgaaccgt
     4081 tatctggcgc cgattttctg cgagaacagc tatgcgtacc aggacagcaa aggcgttatg
     4141 ccggcggttc tgaaagcgaa ggagtacgtt gagctgggta tgcgtcacgt gattgaaatc
     4201 gatctgaaaa actactttga caccattccg ctggaaaacc tgatcccgga gatcgaacgc
     4261 tacatcaccg atgaggcggt tctgcatctg atcaagcaat acctgttttg cgatattagc
     4321 ttcgagggca aaatcagccg taaaacccaa ggtatcgttc aaggtaacgc gatcagcccg
     4381 atcctgagca acctgtacct gaacgacttc gacaaggaac tggacgagag caagctgtgc
     4441 tggattcgtt acgcggacaa tatctacatc tatatggata gctatgagaa ggcgctgctg
     4501 gtgtatagcg agctgaccga gcgtctggag cgtcgcaagc tgaccgttaa caaggagaaa
     4561 agcggcgttt ttgatgtgag cacccgtagc attctgggct acgacatcct gattcgtaac
     4621 aagaaagtgg acgttcgtaa gcatatttac aaaagcgtta atcagtacag caactggcac
     4681 gatagccgcc tggagtttat caacggtcgt taccacatca ccagcgatgg catcctgaat
     4741 cgccaagact tcggcctgct gttcgagaat gagcagaaaa aacactacat cccggttgag
     4801 gttagcgatc agctgaacat ctacggtaat gttaccctgg cgagcaatgt gctgcaaagt
     4861 ttcagcaacc gcgaaatcaa agtgagcttc ttcgacaagt atggtcgtct gattggtagc
     4921 ttcctgccgg aaaaaaccaa gaaaagcgcg gagatcattc tggtgcagag caagaactat
     4981 ctgaacgaag atgtgcgtat ggataccgcg cgccgtatgg aaattgcggg tctgcacaac
     5041 atccgtgcga atctgcgcta ctacgataag aaacacaagg gcgactttaa agagaaggtt
     5101 gacgcgatta gcggctacat tgacgcgctg aaccgtgcgc cgagcgttaa tgatatgatg
     5161 ctgctggaag cgaaagcgcg ccaactgtat tatacctgtt ttaaccaaat cctggaaacc
     5221 agcgacttcc agttcgaaaa gcgtaccaag cgtccgccga aagatgcgat caatgcgtgc
     5281 attagcttcg gcaataccct gctgtataat ctatttgtga atattatctg gaagaagggt
     5341 ctggacccgc gctttggcgt tgttcacgcg agcaacaaac gcaatcaaag cctgaacctg
     5401 gacttcgcgg acatctttaa accgatcgtg attgaccgca ttatcttcac catgatcaat
     5461 aagaaaatgc tgaccctgct gaccgatttt gaaaccagca accagggtgt gtatctgagc
     5521 cgtgagggca agaacatctt cctgcaaatg tacgaggaga agctgaaaag ccgcattacc
     5581 atcaagggta aagagatgag ctattaccag ctgctggaga gcgaagtgca gaattacaag
     5641 aatttcattc tgaccggcga aacctataaa ccgtataagt attattaacg tacgttaact
     5701 ttaagaagga gaacttaaga tgtatgttat cctggtgtac gacatccacc agaagcgtgt
     5761 tggtaaagcg ctgaaaatct gccgtaagta cctgatccac atccaaaaga gcgtgttcga
     5821 aggcaacatc accgagagca aactgaaggc gctgaaagag gaactgggtc acctgattga
     5881 cacccagatg gatagcgtga ttatctacca cctggacagc gttaagtaca ccaaaaagga
     5941 gcagatcggt attgtgcaga gcaccagcaa tgtgatctaa ctcgagtagc ctagcataac
     6001 cccgcggggc ctcttcgggg gtctcgcggg gttttttgct ataaaacgaa aggctcagtc
     6061 gaaagactgg gcctttcgtt ttatctgcta acaaagcccg aaaggaagct gagttggctg
     6121 ctgccaccgc tgagcaataa ctagcataac cccttggggc ctctaaacgg gtcttgaggg
     6181 gttttttgct gaaaggagga actatatccg gactgatagc tagctcagtc ctagggatta
     6241 tgctagctct agtagtggag aattaaattg gaaaaagtcg gtcgatctca tgcctgaaat
     6301 catgaattcc gcaaaatggc ggaaatttaa ggaaaatcag gaatctcaga aaaacgatcg
     6361 accgactttt gtgataaaat ggttgcaaaa aagagaaaaa tttgatttaa tagaatgtga
     6421 aaatagcgga aatgctgatg ttgtacctta cctatgagga attgaaacgt ccccgccagg
     6481 ttgaatccga tatttggagg tacgatggaa cagtctgggt gggattgaga agagaaaaga
     6541 aaaccgccga tcctgtccac cgcattactg caaggtagtg gacaagaccg gcggtcttaa
     6601 gttttt
//
