LOCUS       sequence-369670-        5005 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  B-gTEMP for bacterial expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5005)
  AUTHORS   Lu K, Wazawa T, Sakamoto J, Vu CQ, Nakano M, Kamei Y, Nagai T
  TITLE     Intracellular Heat Transfer and Thermal Property Revealed by 
            Kilohertz Temperature Imaging with a Genetically Encoded 
            Nanothermometer.
  JOURNAL   Nano Lett. 2022 Jul 27;22(14):5698-5707. doi: 
            10.1021/acs.nanolett.2c00608. Epub 2022 Jul 6.
  PUBMED    35792763
REFERENCE   2  (bases 1 to 5005)
  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/acs.nanolett.2c00608"; journalName: "Nano Lett"; date: 
            "2022-07-27- 27"; volume: "22"; issue: "14"; pages: "5698-5707"
COMMENT     Sequence Label: B-gTEMP/pRSET-b
FEATURES             Location/Qualifiers
     source          1..5005
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     2..21
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2..20
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             52..74
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             94..111
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             115..147
                     /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             join(151..159,160..174)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             895..2325
                     /codon_start=1
                     /product="tandem dimeric (pseudo-monomeric) derivative of 
                     DsRed (Shaner et al., 2004)"
                     /label=tdTomato
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFLGHGTGSTGSGSSGTASSEDNNMAVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPY
                     EGTQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVM
                     NFEDGGLVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGV
                     LKGEIHQALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYE
                     RSEGRHHLFLYGMDELYK"
     primer_bind     complement(2388..2406)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2402..2449
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2546..3001
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2633..2652)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2843..2864
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3027..3131
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3132..3200,3201..3992)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3350..3369)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4163..4751
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4652..4671
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4905..4922
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 ttaatacgac tcactatagg gagaccacaa cggtttccct ctagaaataa ttttgtttaa
       61 ctttaagaag gagatataca tatgcggggt tctcatcatc atcatcatca tggtatggct
      121 agcatgactg gtggacagca aatgggtcgg gatctgtacg acgatgacga taaggatccg
      181 atggtgtcca agggcgaaga ggacaacatg gccagcctgc ctgccaccca cgagctgcac
      241 atcttcggca gcatcaacgg cgtggacttc gacatggtgg gacagggcac cggcaacccc
      301 aacgacggct acgaggaact gaacctgaag tccacaaagg gcgacctgca gttcagcccc
      361 tggattctgg tgccccacat cggctacggc ttccaccagt acctgcccta ccccgacggc
      421 atgagccctt tccaggccgc tatggtggat ggcagcggct accaggtgca ccggaccatg
      481 cagtttgagg acggcgccag cctgaccgtg aactaccggt acacatacga gggcagccac
      541 atcaagggcg aggcccaagt gaagggcaca ggctttccag ccgacggccc cgtgatgacc
      601 aatagcctga cagccgccga ctggtgcaga agcaagaaaa cctaccccaa tgacaagacc
      661 atcatcagca ccttcaagtg gtcctacacc accggcaatg gcaagcggta cagaagcacc
      721 gcccggacca cctacacctt cgccaaacct atggccgcca actacctgaa gaaccagcct
      781 atgtacgtgt tccgcaagac cgagctgaag cactccaaga cagaactgaa cttcaaagag
      841 tggcagaaag ccttcaccga cgtgatgggc atggacgagc tgtacaaggg taccatggtg
      901 agcaagggcg aggaggtcat caaagagttc atgcgcttca aggtgcgcat ggagggctcc
      961 atgaacggcc acgagttcga gatcgagggc gagggcgagg gccgccccta cgagggcacc
     1021 cagaccgcca agctgaaggt gaccaagggc ggccccctgc ccttcgcctg ggacatcctg
     1081 tccccccagt tcatgtacgg ctccaaggcg tacgtgaagc accccgccga catccccgat
     1141 tacaagaagc tgtccttccc cgagggcttc aagtgggagc gcgtgatgaa cttcgaggac
     1201 ggcggtctgg tgaccgtgac ccaggactcc tccctgcagg acggcacgct gatctacaag
     1261 gtgaagatgc gcggcaccaa cttccccccc gacggccccg taatgcagaa gaagaccatg
     1321 ggctgggagg cctccaccga gcgcctgtac ccccgcgacg gcgtgctgaa gggcgagatc
     1381 caccaggccc tgaagctgaa ggacggcggc cactacctgg tggagttcaa gaccatctac
     1441 atggccaaga agcccgtgca actgcccggc tactactacg tggacaccaa gctggacatc
     1501 acctcccaca acgaggacta caccatcgtg gaacagtacg agcgctccga gggccgccac
     1561 cacctgttcc tggggcatgg caccggcagc accggcagcg gcagctccgg caccgcctcc
     1621 tccgaggaca acaacatggc cgtcatcaaa gagttcatgc gcttcaaggt gcgcatggag
     1681 ggctccatga acggccacga gttcgagatc gagggcgagg gcgagggccg cccctacgag
     1741 ggcacccaga ccgccaagct gaaggtgacc aagggcggcc ccctgccctt cgcctgggac
     1801 atcctgtccc cccagttcat gtacggctcc aaggcgtacg tgaagcaccc cgccgacatc
     1861 cccgattaca agaagctgtc cttccccgag ggcttcaagt gggagcgcgt gatgaacttc
     1921 gaggacggcg gtctggtgac cgtgacccag gactcctccc tgcaggacgg cacgctgatc
     1981 tacaaggtga agatgcgcgg caccaacttc ccccccgacg gccccgtaat gcagaagaag
     2041 accatgggct gggaggcctc caccgagcgc ctgtaccccc gcgacggcgt gctgaagggc
     2101 gagatccacc aggccctgaa gctgaaggac ggcggccact acctggtgga gttcaagacc
     2161 atctacatgg ccaagaagcc cgtgcaactg cccggctact actacgtgga caccaagctg
     2221 gacatcacct cccacaacga ggactacacc atcgtggaac agtacgagcg ctccgagggc
     2281 cgccaccacc tgttcctgta cggcatggac gagctgtaca agtaagaatt cgaagcttga
     2341 tccggctgct aacaaagccc gaaaggaagc tgagttggct gctgccaccg ctgagcaata
     2401 actagcataa ccccttgggg cctctaaacg ggtcttgagg ggttttttgc tgaaaggagg
     2461 aactatatcc ggatctggcg taatagcgaa gaggcccgca ccgatcgccc ttcccaacag
     2521 ttgcgcagcc tgaatggcga atgggacgcg ccctgtagcg gcgcattaag cgcggcgggt
     2581 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc
     2641 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
     2701 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat
     2761 tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg
     2821 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct
     2881 atctcggtct attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa
     2941 aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac gcttacaatt
     3001 taggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     3061 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     3121 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
     3181 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
     3241 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
     3301 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
     3361 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc
     3421 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
     3481 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
     3541 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
     3601 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
     3661 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
     3721 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
     3781 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
     3841 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
     3901 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
     3961 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     4021 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     4081 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     4141 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     4201 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     4261 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt
     4321 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     4381 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     4441 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     4501 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     4561 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     4621 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     4681 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     4741 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     4801 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
     4861 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg
     4921 aggaagcgga agagcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt
     4981 aatgcaggat ctcgatcccg cgaaa
//
