LOCUS       sequence-446183-        4091 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of N-terminal His-tagged qmTQ2-cAMP, a cyan 
            fluorescent FLIM biosensor for detecting cAMP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4091)
  AUTHORS   Zhong C, Arai S, Okada Y
  TITLE     Development of fluorescence lifetime biosensors for ATP, cAMP, 
            citrate, and glucose using the mTurquoise2-based platform.
  JOURNAL   Cell Rep Methods. 2024 Nov 18;4(11):100902. doi: 
            10.1016/j.crmeth.2024.100902.
  PUBMED    39561716
REFERENCE   2  (bases 1 to 4091)
  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 Rep 
            Methods."; date: "2024-11-18"; volume: "4(11):100902. doi"; pages: "
            10.1016/j.crmeth.2024.100902"
COMMENT     Sequence Label: pRSETA-qmTQ2-cAMP
FEATURES             Location/Qualifiers
     source          1..4091
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     20..39
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        20..38
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             70..92
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             112..129
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             133..165
                     /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(169..177,178..192)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     primer_bind     complement(262..283)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(523..542)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     CDS             1180..1431
                     /codon_start=1
                     /product="C-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VC155
                     /translation="DKQKNGIKANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNH
                     YLSTQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYK"
     primer_bind     1368..1389
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1492..1510)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1506..1553
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1650..2105
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1737..1756)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1947..1968
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2131..2235
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2236..2304,2305..3096)
                     /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(2454..2473)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3267..3855
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3756..3775
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4009..4026
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gatctcgatc ccgcgaaatt aatacgactc actataggga gaccacaacg gtttccctct
       61 agaaataatt ttgtttaact ttaagaagga gatatacata tgcggggttc tcatcatcat
      121 catcatcatg gtatggctag catgactggt ggacagcaaa tgggtcggga tctgtacgac
      181 gatgacgata aggatcgatg gggatccgag ctcgagatgg tgagcaaggg cgaggagctg
      241 ttcaccgggg tggtgcccat cctggtcgag ctggacggcg acgtaaacgg ccacaagttc
      301 agcgtgtccg gcgagggcga gggcgatgcc acctacggca agctgaccct gaagttcatc
      361 tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct gtcctggggc
      421 gtgcagtgct tcgcccgcta ccccgaccac atgaagcagc acgacttctt caagtccgcc
      481 atgcccgaag gctacgtcca ggagcgcacc atcttcttca aggacgacgg caactacaag
      541 acccgcgccg aggtgaagtt cgagggcgac accctggtga accgcatcga gctgaagggc
      601 atcgacttca aggaggacgg caacatcctg gggcacaagc tggagtacaa ccgtggtacc
      661 gaggagcttg ttgaggctat ggccctcctg tcccagcgag ggcctgatgc cctactcacc
      721 gtggcgctcc ggaagccccc aggtcagcgt acggatgaag aactggacct gatctttgaa
      781 gagctgctgc atatcaaggc ggtggcacac ctttctaact cggtgaagcg ggaactagct
      841 gctgttctgc tctttgaacc acacagcaag gcaggaactg tgttgttcag ccagggggac
      901 aagggtacct cgtggtacat tatctggaag ggatctgtca atgtggtgac ccatggcaag
      961 gggctggtga ccacgttgca cgagggagat gactttggac agctggctct ggtgaacgac
     1021 gcacctcggg cagccaccat catccttcga gaaaataact gtcactttct gcgtgtggac
     1081 aagcaggact tcaaccgcat catcaaggat gtggaagcaa aaaccatgag actggaagaa
     1141 gcactggaat tcttcagcga caacgtctat atcaccgccg acaagcagaa gaacggcatc
     1201 aaggccaact tcaagatccg ccacaacatc gaggacggcg gcgtgcagct cgccgaccac
     1261 taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg
     1321 agcacccagt ccaagctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg
     1381 gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa gtaacgaagc
     1441 ttgatccggc tgctaacaaa gcccgaaagg aagctgagtt ggctgctgcc accgctgagc
     1501 aataactagc ataacccctt ggggcctcta aacgggtctt gaggggtttt ttgctgaaag
     1561 gaggaactat atccggatct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca
     1621 acagttgcgc agcctgaatg gcgaatggga cgcgccctgt agcggcgcat taagcgcggc
     1681 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc
     1741 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
     1801 tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact
     1861 tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt
     1921 gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa
     1981 ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt
     2041 aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgcttac
     2101 aatttaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa
     2161 atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat
     2221 tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg
     2281 gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa
     2341 gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt
     2401 gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt
     2461 ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat
     2521 tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg
     2581 acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta
     2641 cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat
     2701 catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag
     2761 cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa
     2821 ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca
     2881 ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc
     2941 ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt
     3001 atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc
     3061 gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat
     3121 atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt
     3181 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     3241 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     3301 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     3361 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta
     3421 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     3481 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     3541 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     3601 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     3661 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     3721 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     3781 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     3841 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     3901 ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc
     3961 gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg
     4021 agcgaggaag cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt
     4081 cattaatgca g
//
