LOCUS       sequence-326023-        7245 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  sMHT-113 fused to GFP and RFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7245)
  AUTHORS   Fulk EM, Huh D, Atkinson JT, Lie M, Masiello CA, Silberg JJ
  TITLE     A Split Methyl Halide Transferase AND Gate That Reports by 
            Synthesizing an Indicator Gas.
  JOURNAL   ACS Synth Biol. 2020 Nov 20;9(11):3104-3113. doi: 
            10.1021/acssynbio.0c00355. Epub 2020 Oct 26.
  PUBMED    33104325
REFERENCE   2  (bases 1 to 7245)
  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: 
            "10.1021/acssynbio.0c00355"; journalName: "ACS Synth Biol"; date: 
            "2020-11-20- 20"; volume: "9"; issue: "11"; pages: "3104-3113"
COMMENT     Sequence Label: pDH013
FEATURES             Location/Qualifiers
     source          1..7245
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(77..171)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(192..851)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     766..785
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     primer_bind     1020..1038
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        join(1059..1077,1078..1083,1084..1100,1101..1106,
                     1107..1132)
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator 
                     sites (Lutz and Bujard, 1997)"
     protein_bind    1059..1077
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1084..1102
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     RBS             1203..1211
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     CDS             1230..1247
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1257..1274
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1278..1310
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             1881..2594
                     /codon_start=1
                     /product="S65T variant of Aequorea victoria green 
                     fluorescent protein (Heim et al., 1995)"
                     /label=GFP (S65T)
                     /note="excitable with blue light"
                     /translation="MRKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(1911..1939)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     2534..2554
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      complement(2653..2684)
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     protein_bind    2773..2789
                     /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             2892..2900
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     CDS             2907..3581
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     CDS             4152..4178
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     terminator      4214..4285
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      4301..4328
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     promoter        4380..4414
                     /label=BBa_J23106
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.47"
     CDS             4449..5075
                     /codon_start=1
                     /gene="tetR"
                     /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="MMSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWH
                     VKNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGT
                     RPTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPT
                     TDSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     complement(4525..4544)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     terminator      5081..5167
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     promoter        5216..5293
                     /gene="lacI"
                     /label=lacI promoter
     CDS             5294..6376
                     /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     complement(5313..5332)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     terminator      6382..6468
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     rep_origin      complement(6646..7234)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6726..6745)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
       61 catgactagt gcttggattc tcaccaataa aaaacgcccg gcggcaaccg agcgttctga
      121 acaaatccag atggagttct gaggtcatta ctggatctat caacaggagt ccaagcgagc
      181 tcgatatcaa attacgcccc gccctgccac tcatcgcagt actgttgtaa ttcattaagc
      241 attctgccga catggaagcc atcacaaacg gcatgatgaa cctgaatcgc cagcggcatc
      301 agcaccttgt cgccttgcgt ataatatttg cccatggtga aaacgggggc gaagaagttg
      361 tccatattgg ccacgtttaa atcaaaactg gtgaaactca cccagggatt ggctgagacg
      421 aaaaacatat tctcaataaa ccctttaggg aaataggcca ggttttcacc ataacacgcc
      481 acatcttgcg aatatatgtg tagaaactgc cggaaatcgt cgtggtattc actccagagc
      541 gatgaaaacg tttcagtttg ctcatggaaa acggtgtaac aagggtgaac actatcccat
      601 atcaccagct caccgtcttt cattgccata cgaaattccg gatgagcatt catcaggcgg
      661 gcaagaatgt gaataaaggc cggataaaac ttgtgcttat ttttctttac ggtctttaaa
      721 aaggccgtaa tatccagctg aacggtctgg ttataggtac attgagcaac tgactgaaat
      781 gcctcaaaat gttctttacg atgccattgg gatatatcaa cggtggtata tccagtgatt
      841 tttttctcca ttttagcttc cttagctcct gaaaatctcg ataactcaaa aaatacgacc
      901 ggtagtgatc ttatttcatt atggtgaaag ttggaacctc ttacgggccg gccaacgtct
      961 cattttcgcc agatatcgac gtctaagaaa ccattattat catgacatta acctataaaa
     1021 ataggcgtat cacgaggccc tttcgtcttc acctcgagtc cctatcagtg atagagattg
     1081 acatccctat cagtgataga gatactgagc acatcagcag gacgcactga ccgggcccaa
     1141 gttcacttaa aaaggagatc aacaatgaaa gcaattttcg tactgaaaca tcttaatcat
     1201 gctaaggagg ttttctaatg ggcagcagcc atcatcatca tcatcacagc agcggcctgg
     1261 tgccgcgcgg cagccatatg gctagcatga ctggtggaca gcaaatgggt gcgggatcca
     1321 tgagcaccgt ggcgaacatt gcgccggtgt ttaccggcga ttgcaaaacc attccgaccc
     1381 cggaagaatg cgcgaccttt ctgtataaag ttgtgaacag cggcggctgg gaaaaatgct
     1441 gggtggaaga agtgattccg tgggatctgg gcgtgccgac cccgctggtg ctgcatctgg
     1501 tgaaaaacaa cgcgctgccg aacggcaaag gcctggtgcc gggctgcggc ggcggctatg
     1561 atgttgtggc gatggcgaac ccggaacgct ttatggtggg cctggatatt agcgaaaacg
     1621 cgctgaaaaa agcgcgcgaa acctttagca ccatgccggc ggccgcgggt ggcggtggct
     1681 ctggaggtgg tgggtccatg aacgaaaaag aagaactgaa atctaaaaaa gcggaactgc
     1741 gcaaccgcat cgaacagctg aaacagaaac gcgaacagct gaaacagaaa atcgcgaacc
     1801 tgcgcaaaga aatcgaagcg tataaaggtg gaggaggttc tggaggcggt ggaagtggtg
     1861 gcggaggtag cgcggccgcg atgcgtaaag gagaagaact tttcactgga gttgtcccaa
     1921 ttcttgttga attagatggt gatgttaatg ggcacaaatt ttccgtcagt ggagagggtg
     1981 aaggtgatgc aacatacgga aaacttaccc ttaaatttat ttgcactact ggaaaactac
     2041 ctgttccatg gccaacactt gtcactactt tcggttatgg tgttcaatgc tttgcgagat
     2101 acccagatca tatgaaacag catgactttt tcaagagtgc catgcccgaa ggttatgtac
     2161 aggaaagaac tatatttttc aaagatgacg ggaactacaa gacacgtgct gaagtcaagt
     2221 ttgaaggtga tacccttgtt aatagaatcg agttaaaagg tattgatttt aaagaagatg
     2281 gaaacattct tggacacaaa ttggaataca actataactc acacaatgta tacatcatgg
     2341 cagacaaaca aaagaatgga atcaaagtta acttcaaaat tagacacaac attgaagatg
     2401 gaagcgttca actagcagac cattatcaac aaaatactcc aattggcgat ggccctgtcc
     2461 ttttaccaga caaccattac ctgtccacac aatctgccct ttcgaaagat cccaacgaaa
     2521 agagggacca catggtcctt cttgagtttg taacagctgc tgggattaca catggcatgg
     2581 atgaactata caaataataa ggatcctgat tcgaacaatt gaaaaaacct cgcgccttac
     2641 ctgttgagta atagtcaaaa gcctccggtc ggaggctttt gactttctgc ttactgaatt
     2701 tcggtggtgc cgttaattaa ccggtgggcc tcgtcttcac ctcgagataa atgtgagcgg
     2761 ataacattga cattgtgagc ggataacaag atactgagca catcagcagg acgcactgac
     2821 cgggcccaag ttcacttaaa aaggagatca acaatgaaag caattttcgt actgaaacat
     2881 cttaatcatg ctaaggaggt tttctaatgg cgagtagcga agacgttatc aaagagttca
     2941 tgcgtttcaa agttcgtatg gaaggttccg ttaacggtca cgagttcgaa atcgaaggtg
     3001 aaggtgaagg tcgtccgtac gaaggtaccc agaccgctaa actgaaagtt accaaaggtg
     3061 gtccgctgcc gttcgcttgg gacatcctgt ccccgcagtt ccagtacggt tccaaagctt
     3121 acgttaaaca cccggctgac atcccggact acctgaaact gtccttcccg gaaggtttca
     3181 aatgggaacg tgttatgaac ttcgaagacg gtggtgttgt taccgttacc caggactcct
     3241 ccctgcaaga cggtgagttc atctacaaag ttaaactgcg tggtaccaac ttcccgtccg
     3301 acggtccggt tatgcagaaa aaaaccatgg gttgggaagc ttccaccgaa cgtatgtacc
     3361 cggaagacgg tgctctgaaa ggtgaaatca aaatgcgtct gaaactgaaa gacggtggtc
     3421 actacgacgc tgaagttaaa accacctaca tggctaaaaa accggttcag ctgccgggtg
     3481 cttacaaaac cgacatcaaa ctggacatca cctcccacaa cgaagactac accatcgttg
     3541 aacagtacga acgtgctgaa ggtcgtcact ccaccggtgc tggtggagga ggttctggag
     3601 gcggtggaag tggtggcgga ggtagcgcgg ccgcaatgtc tatcgcggcc accctggaaa
     3661 acgacttagc acgccttgag aatgaaaatg ctcgtctgga gaaagatatc gcgaatctcg
     3721 agcgtgacct ggcaaaatta gagcgtgagg aagcttattt cggtggcggt ggctctggag
     3781 gtggtgggtc cgcggccgca aacagcagct gctttagctt tgtgaaagaa gatgtgttta
     3841 cctggcgccc ggaacagccg tttgatttta tttttgatta tgtgtttttt tgcgcgattg
     3901 atccgaaaat gcgcccggcg tggggcaaag cgatgtatga actgctgaaa ccggatggcg
     3961 aactgattac cctgatgtat ccgattacca accatgaagg cggcccgccg tttagcgtga
     4021 gcgaaagcga atatgaaaaa gtgctggtgc cgctgggctt taaacagctg agcctggaag
     4081 attatagcga tctggcggtg gaaccgcgca aaggcaaaga aaaactggcg cgctggaaaa
     4141 aaatgaacaa ctacccttac gatgtgcctg actacgcgta atgactgaga tccggctaat
     4201 ctagaccagg catcaaataa aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat
     4261 ctgttgtttg tcggtgaacg ctctctacta gagtcacact ggctcacctt cgggtgggcc
     4321 tttctgcgtt tataggtacc cgtggatcct ctagagtcga cctgcaggca tgcaagcttt
     4381 ttacggctag ctcagtccta ggtatagtgc tagcccagcc agagaaacac tctttaacag
     4441 ggggtagtat gatgtctcgt ttagataaaa gtaaagtgat taacagcgca ttagagctgc
     4501 ttaatgaggt cggaatcgaa ggtttaacaa cccgtaaact cgcccagaag ctaggtgtag
     4561 agcagcctac attgtattgg catgtaaaaa ataagcgggc tttgctcgac gccttagcca
     4621 ttgagatgtt agataggcac catactcact tttgcccttt agaaggggaa agctggcaag
     4681 attttttacg taataacgct aaaagtttta gatgtgcttt actaagtcat cgcgatggag
     4741 caaaagtaca tttaggtaca cggcctacag aaaaacagta tgaaactctc gaaaatcaat
     4801 tagccttttt atgccaacaa ggtttttcac tagagaatgc attatatgca ctcagcgcag
     4861 tggggcattt tactttaggt tgcgtattgg aagatcaaga gcatcaagtc gctaaagaag
     4921 aaagggaaac acctactact gatagtatgc cgccattatt acgacaagct atcgaattat
     4981 ttgatcacca aggtgcagag ccagccttct tattcggcct tgaattgatc atatgcggat
     5041 tagaaaaaca acttaaatgt gaaagtgggt cttaaggcat caaataaaac gaaaggctca
     5101 gtcgaaagac tgggcctttc gttttatctg ttgtttgtcg gtgaacgctc tcctgagtag
     5161 gacaaatccg ccgccctaga cctagggcgt tcggctgccg gcatgcattt acgttgacac
     5221 catcgaatgg cgcaaaacct ttcgcggtat ggcatgatag cgcccggaag agagtcaatt
     5281 cagggtggtg aatgtgaaac cagtaacgtt atacgatgtc gcagagtatg ccggtgtctc
     5341 ttatcagacc gtttcccgcg tggtgaacca ggccagccac gtttctgcga aaacgcggga
     5401 aaaagtggaa gcggcgatgg cggagctgaa ttacattccc aaccgcgtgg cacaacaact
     5461 ggcgggcaaa cagtcgttgc tgattggcgt tgccacctcc agtctggccc tgcacgcgcc
     5521 gtcgcaaatt gtcgcggcga ttaaatctcg cgccgatcaa ctgggtgcca gcgtggtggt
     5581 gtcgatggta gaacgaagcg gcgtcgaagc ctgtaaagcg gcggtgcaca atcttctcgc
     5641 gcaacgcgtc agtgggctga tcattaacta tccgctggat gaccaggatg ccattgctgt
     5701 ggaagctgcc tgcactaatg ttccggcgtt atttcttgat gtctctgacc agacacccat
     5761 caacagtatt attttctccc atgaagacgg tacgcgactg ggcgtggagc atctggtcgc
     5821 attgggtcac cagcaaatcg cgctgttagc gggcccatta agttctgtct cggcgcgtct
     5881 gcgtctggct ggctggcata aatatctcac tcgcaatcaa attcagccga tagcggaacg
     5941 ggaaggcgac tggagtgcca tgtccggttt tcaacaaacc atgcaaatgc tgaatgaggg
     6001 catcgttccc actgcgatgc tggttgccaa cgatcagatg gcgctgggcg caatgcgcgc
     6061 cattaccgag tccgggctgc gcgttggtgc ggatatctcg gtagtgggat acgacgatac
     6121 cgaagacagc tcatgttata tcccgccgtt aaccaccatc aaacaggatt ttcgcctgct
     6181 ggggcaaacc agcgtggacc gcttgctgca actctctcag ggccaggcgg tgaagggcaa
     6241 tcagctgttg cccgtctcac tggtgaaaag aaaaaccacc ctggcgccca atacgcaaac
     6301 cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg tttcccgact
     6361 ggaaagcggg cagtgaggca tcaaataaaa cgaaaggctc agtcgaaaga ctgggccttt
     6421 cgttttatct gttgtttgtc ggtgaacgct ctcctgagta ggacaaatcc gccgccctag
     6481 acctagggcg ttcggctgcg gcgagcggta tcagcgagcg gtatcagctc actcaaaggc
     6541 ggtaatacgg ttatccacag aatcagggga taacgcatgc aagaacatgt gagcaaaagg
     6601 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     6661 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     6721 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     6781 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     6841 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     6901 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     6961 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     7021 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     7081 tagaaggaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     7141 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
     7201 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat ccttt
//
