LOCUS       sequence-361530-        6616 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  encodes aTc inducible FsRT-Cas1–Cas2 expression cassette and 
            FsCRISPR Array 2 transcribed by BBa_J23106.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6616)
  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 6616)
  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_1064_pET30_pCat-tetR-Term-ptetA-FsRT-Cas1(opt)-Cas2(opt)-3xTerm_
            J23106-Leader-ARRAY2-DR2-FaqI_Term_NotI
FEATURES             Location/Qualifiers
     source          1..6616
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(241..1056)
                     /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(241..1056)
                     /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     964..983
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     1048..1067
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      1178..1766
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1667..1686
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1920..1937
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    1952..2091
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2017..2036)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2177..2199
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2193..2384)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     RBS             2914..2925
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             2932..3555
                     /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(3005..3024)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     terminator      3600..3671
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3687..3714
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     protein_bind    3729..3747
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3754..3772
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     terminator      complement(6062..6105)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(6148..6166)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      6162..6209
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        6234..6268
                     /label=BBa_J23106
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.47"
ORIGIN
        1 gaccggcggt cttaagtttt ttggctgaag cggccgctat tcttttgatt tataagggat
       61 tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
      121 ttttaacaaa atattaacgc ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga
      181 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gaattaattc
      241 ttagaaaaac tcatcgagca tcaaatgaaa ctgcaattta ttcatatcag gattatcaat
      301 accatatttt tgaaaaagcc gtttctgtaa tgaaggagaa aactcaccga ggcagttcca
      361 taggatggca agatcctggt atcggtctgc gattccgact cgtccaacat caatacaacc
      421 tattaatttc ccctcgtcaa aaataaggtt atcaagtgag aaatcaccat gagtgacgac
      481 tgaatccggt gagaatggca aaagtttatg catttctttc cagacttgtt caacaggcca
      541 gccattacgc tcgtcatcaa aatcactcgc atcaaccaaa ccgttattca ttcgtgattg
      601 cgcctgagcg agacgaaata cgcgatcgct gttaaaagga caattacaaa caggaatcga
      661 atgcaaccgg cgcaggaaca ctgccagcgc atcaacaata ttttcacctg aatcaggata
      721 ttcttctaat acctggaatg ctgttttccc ggggatcgca gtggtgagta accatgcatc
      781 atcaggagta cggataaaat gcttgatggt cggaagaggc ataaattccg tcagccagtt
      841 tagtctgacc atctcatctg taacatcatt ggcaacgcta cctttgccat gtttcagaaa
      901 caactctggc gcatcgggct tcccatacaa tcgatagatt gtcgcacctg attgcccgac
      961 attatcgcga gcccatttat acccatataa atcagcatcc atgttggaat ttaatcgcgg
     1021 cctagagcaa gacgtttccc gttgaatatg gctcataaca ccccttgtat tactgtttat
     1081 gtaagcagac agttttattg ttcatgacca aaatccctta acgtgagttt tcgttccact
     1141 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     1201 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     1261 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     1321 ctgtccttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     1381 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     1441 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     1501 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     1561 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     1621 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     1681 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     1741 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     1801 tcttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     1861 accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca
     1921 gcgagtcagt gagcgaggaa gcggaagagc gcctgatgcg gtattttctc cttacgcatc
     1981 tgtgcggtat ttcacaccgc aatggtgcac tctcagtaca atctgctctg atgccgcata
     2041 gttaagccag tatacactcc gctatcgcta cgtgactggg tcatggctgc gccccgacac
     2101 ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga
     2161 caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa
     2221 cgcgcgaggc agctgcggta aagctcatca gcgtggtcgt gaagcgattc acagatgtct
     2281 gcctgttcat ccgcgtccag ctcgttgagt ttctccagaa gcgttaatgt ctggcttctg
     2341 ataaagcggg ccatgttaag ggcggttttt tcctgtttgg tcactgatgc ctccgtgtaa
     2401 gggggatttc tgttcatggg ggtaatgata ccgatgaaac gagagaggat gctcacgata
     2461 cgggttactg atgatgaaca tgcccggtta ctggaacgtt gtgagggtaa acaactggcg
     2521 gtatggatgc ggcgggacca gagaaaaatc actcagggtc aatgccagcg cttcgttaat
     2581 acagatgtag gtgttccaca gggtagccag cagcatcctg cgatgcagat ccggaacata
     2641 atggtgcagg gcgctgactt ccgcgtttcc agactttacg aaacacggaa accgaagatc
     2701 attcatgttg ttgctcaggt cgcagacgtt ttgcagcagc agtcgcttca cgttcgctcg
     2761 cgtatcggtg attcattctg ctaaccagta aggcaacccc gccagcctag ccgggtcctc
     2821 aacgacagga gcacgatcat gcgcacccgt ggggccgcca tgccggcggc acgtaagagg
     2881 ttccaacttt caccataatg aaacatacta gagaaagagg agaaatacta gatgtccaga
     2941 ttagataaaa gtaaagtgat taacagcgca ttagagctgc ttaatgaggt cggaatcgaa
     3001 ggtttaacaa cccgtaaact cgcccagaag ctaggtgtag agcagcctac attgtattgg
     3061 catgtaaaaa ataagcgggc tttgctcgac gccttagcca ttgagatgtt agataggcac
     3121 catactcact tttgcccttt agaaggggaa agctggcaag attttttacg taataacgct
     3181 aaaagtttta gatgtgcttt actaagtcat cgcgatggag caaaagtaca tttaggtaca
     3241 cggcctacag aaaaacagta tgaaactctc gaaaatcaat tagccttttt atgccaacaa
     3301 ggtttttcac tagagaatgc attatatgca ctcagcgctg tggggcattt tactttaggt
     3361 tgcgtattgg aagatcaaga gcatcaagtc gctaaagaag aaagggaaac acctactact
     3421 gatagtatgc cgccattatt acgacaagct atcgaattat ttgatcacca aggtgcagag
     3481 ccagccttct tattcggcct tgaattgatc atatgcggat tagaaaaaca acttaaatgt
     3541 gaaagtgggt cctaataaca ctgatagtgc tagtgtagat cactactaga gccaggcatc
     3601 aaataaaacg aaaggctcag tcgaaagact gggcctttcg ttttatctgt tgtttgtcgg
     3661 tgaacgctct ctactagagt cacactggct caccttcggg tgggcctttc tgcgtttata
     3721 tactagagtc cctatcagtg atagagattg acatccctat cagtgataga gatactgagc
     3781 actactagag aagcttgcgg aggagcatgc atgtttacca tcgacgagat gctgagcaag
     3841 aacaaccaac gtctggcgtt cgaacacttt gcgaccaaaa acgacggttg cggcccggac
     3901 ggtatgcacg ttagcgaact ggagaagtac tggcgtatga atcacgacca gatcattagc
     3961 gatctgaaga accaggaata ccagccgggt attatcctga ttcgtgagca catgaataaa
     4021 accggcaaac gccgtaatat cgcgagcctg aacgttattg atcgtttcat tacccgtctg
     4081 ctgagccaga aactgaaccg ttatctggcg ccgattttct gcgagaacag ctatgcgtac
     4141 caggacagca aaggcgttat gccggcggtt ctgaaagcga aggagtacgt tgagctgggt
     4201 atgcgtcacg tgattgaaat cgatctgaaa aactactttg acaccattcc gctggaaaac
     4261 ctgatcccgg agatcgaacg ctacatcacc gatgaggcgg ttctgcatct gatcaagcaa
     4321 tacctgtttt gcgatattag cttcgagggc aaaatcagcc gtaaaaccca aggtatcgtt
     4381 caaggtaacg cgatcagccc gatcctgagc aacctgtacc tgaacgactt cgacaaggaa
     4441 ctggacgaga gcaagctgtg ctggattcgt tacgcggaca atatctacat ctatatggat
     4501 agctatgaga aggcgctgct ggtgtatagc gagctgaccg agcgtctgga gcgtcgcaag
     4561 ctgaccgtta acaaggagaa aagcggcgtt tttgatgtga gcacccgtag cattctgggc
     4621 tacgacatcc tgattcgtaa caagaaagtg gacgttcgta agcatattta caaaagcgtt
     4681 aatcagtaca gcaactggca cgatagccgc ctggagttta tcaacggtcg ttaccacatc
     4741 accagcgatg gcatcctgaa tcgccaagac ttcggcctgc tgttcgagaa tgagcagaaa
     4801 aaacactaca tcccggttga ggttagcgat cagctgaaca tctacggtaa tgttaccctg
     4861 gcgagcaatg tgctgcaaag tttcagcaac cgcgaaatca aagtgagctt cttcgacaag
     4921 tatggtcgtc tgattggtag cttcctgccg gaaaaaacca agaaaagcgc ggagatcatt
     4981 ctggtgcaga gcaagaacta tctgaacgaa gatgtgcgta tggataccgc gcgccgtatg
     5041 gaaattgcgg gtctgcacaa catccgtgcg aatctgcgct actacgataa gaaacacaag
     5101 ggcgacttta aagagaaggt tgacgcgatt agcggctaca ttgacgcgct gaaccgtgcg
     5161 ccgagcgtta atgatatgat gctgctggaa gcgaaagcgc gccaactgta ttatacctgt
     5221 tttaaccaaa tcctggaaac cagcgacttc cagttcgaaa agcgtaccaa gcgtccgccg
     5281 aaagatgcga tcaatgcgtg cattagcttc ggcaataccc tgctgtataa tctatttgtg
     5341 aatattatct ggaagaaggg tctggacccg cgctttggcg ttgttcacgc gagcaacaaa
     5401 cgcaatcaaa gcctgaacct ggacttcgcg gacatcttta aaccgatcgt gattgaccgc
     5461 attatcttca ccatgatcaa taagaaaatg ctgaccctgc tgaccgattt tgaaaccagc
     5521 aaccagggtg tgtatctgag ccgtgagggc aagaacatct tcctgcaaat gtacgaggag
     5581 aagctgaaaa gccgcattac catcaagggt aaagagatga gctattacca gctgctggag
     5641 agcgaagtgc agaattacaa gaatttcatt ctgaccggcg aaacctataa accgtataag
     5701 tattattaac gtacgttaac tttaagaagg agaacttaag atgtatgtta tcctggtgta
     5761 cgacatccac cagaagcgtg ttggtaaagc gctgaaaatc tgccgtaagt acctgatcca
     5821 catccaaaag agcgtgttcg aaggcaacat caccgagagc aaactgaagg cgctgaaaga
     5881 ggaactgggt cacctgattg acacccagat ggatagcgtg attatctacc acctggacag
     5941 cgttaagtac accaaaaagg agcagatcgg tattgtgcag agcaccagca atgtgatcta
     6001 actcgagtag cctagcataa ccccgcgggg cctcttcggg ggtctcgcgg ggttttttgc
     6061 tataaaacga aaggctcagt cgaaagactg ggcctttcgt tttatctgct aacaaagccc
     6121 gaaaggaagc tgagttggct gctgccaccg ctgagcaata actagcataa ccccttgggg
     6181 cctctaaacg ggtcttgagg ggttttttgc tgaaaggagg aactatatcc ggatttacgg
     6241 ctagctcagt cctaggtata gtgctagctc tagtagtggc ggggaagctt atgttccata
     6301 gcaaaaagtc ggtcagtctc gtggctgaaa tcatgagttc cacaaaatgg ctgaaattca
     6361 aggaaaatca ggaatctcag aaaaacgatc gaccgacttt ttcgataaaa tggttgcaaa
     6421 aatgagaaaa atctgattta atagaatctg aaaacagcgg aaatgctgtt gtcgtacttt
     6481 acctaaaagg aattgaaacg tccccgccag gttgaatccg atatttggag gtacgatgga
     6541 acagtctggg tgggattgag aagagaaaag aaaaccgccg atcctgtcca ccgcattact
     6601 gcaaggtagt ggacaa
//
