LOCUS       sequence-458381-        7004 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expressing the human version of TINGL biosensor for glucose (named 
            THINGL) in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7004)
  AUTHORS   Botman D, Tielman A, Goedhart J, Teusink B
  TITLE     An mTurquoise2-based glucose biosensor
  JOURNAL   bioRxiv 2024.11.29.626064
REFERENCE   2  (bases 1 to 7004)
  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: "bioRxiv 
            2024.11.29.626064"
COMMENT     Sequence Label: THINGL pcDNA3.1
FEATURES             Location/Qualifiers
     source          1..7004
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(706..727)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(967..986)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     CDS             2053..2304
                     /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     2241..2262
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     promoter        complement(2323..2341)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2324..2341)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2361..2378)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2367..2591
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2637..3065
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2724..2743)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2934..2955
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3074..3094)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3079..3408
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3259..3394
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3321..3340
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3475..4269
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3529..3548)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4139..4158
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4443..4564
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4480..4499)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4534..4553
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4613..4629)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4613..4629)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4626..4648)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4637..4653
                     /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)."
     promoter        complement(join(4661..4667,4668..4685,4686..4691))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4706..4727
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4844..4861)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5015..5603)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5095..5114)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5774..6565,6566..6634))
                     /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     6397..6416
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6635..6739)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6814..6833)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctt
      661 atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac
      721 ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac
      781 ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc ctggcccacc
      841 ctcgtgacca ccctgtcctg gggcgtgcag tgcttcgccc gctaccccga ccacatgaag
      901 cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc
      961 ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg
     1021 gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac
     1081 aagctggagt acaactacat ggttgatact agcattggtg taacaatcta taagtacgac
     1141 gattattata tgtctgtagt gcgcaaggct attgagcaag atgcgaaagc cgcgccagat
     1201 gttcagctgc tgatgaatga ttctcagaat gaccagtcca agcagaacga tcagatcgac
     1261 gtattgctgg cgaaaggggt gaaggcactg gcaatcaacc tggttgaccc ggcagctgcg
     1321 ggtacggtga ttgagaaagc gcgtgggcaa aacgtgccgg tggttttctt caacaaagaa
     1381 ccgtctcgta aggcgctgga tagctacgac aaagcctact acgttggcac tgactccaaa
     1441 gagtccggca ttattcaagg cgatttgatt gctaaacact gggcggcgaa tcagggttgg
     1501 gatctgaaca aagacggtca gattcagttc gtactgctga aaggtgaacc gggccatccg
     1561 gatgcagaag cacgtaccac ttacgtgatt aaagaattga acgataaagg catcaaaact
     1621 gaacagttac agttagatac cgcaatgtgg gacaccgctc aggcgaaaga taagatggac
     1681 gcctggctgt ctggcccgaa cgccaacaaa atcgaagtgg ttatcgccaa caacgatgcg
     1741 atggcaatgg gcgcggttga agcgctgaaa gcacacaaca agtccagcat tccggtgttt
     1801 ggcgtcgatg cgctgccaga agcgctggcg ctggtgaaat ccggtgcact ggcgggcacc
     1861 gtactgaacg atgctaacaa ccaggcgaaa gcgacctttg atctggcgaa aaacctggcc
     1921 gatggtaaag gtgcggctga tggcaccaac tggaaaatcg acaacaaagt ggtccgcgta
     1981 ccttatgttg gcgtagataa agacaacctg gctgagttca gctactttag cgacaacgtc
     2041 tatatcaccg ccgacaagca gaagaacggc atcaaggcca acttcaagat ccgccacaac
     2101 atcgaggacg gcggcgtgca gctcgccgac cactaccagc agaacacccc catcggcgac
     2161 ggccccgtgc tgctgcccga caaccactac ctgagcaccc agtccaagct gagcaaagac
     2221 cccaacgaga agcgcgatca catggtcctg ctggagttcg tgaccgccgc cgggatcact
     2281 ctcggcatgg acgagctgta caagtaatct agagggccct attctatagt gtcacctaaa
     2341 tgctagagct cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg
     2401 cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata
     2461 aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt
     2521 ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt
     2581 gggctctatg gcttctgagg cggaaagaac cagctggggc tctagggggt atccccacgc
     2641 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     2701 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     2761 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     2821 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
     2881 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     2941 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
     3001 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     3061 gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca
     3121 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca
     3181 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc
     3241 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
     3301 catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta
     3361 ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccggga
     3421 gcttgtatat ccattttcgg atctgatcaa gagacaggat gaggatcgtt tcgcatgatt
     3481 gaacaagatg gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat
     3541 gactgggcac aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag
     3601 gggcgcccgg ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaggac
     3661 gaggcagcgc ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac
     3721 gttgtcactg aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc
     3781 ctgtcatctc accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg
     3841 ctgcatacgc ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag
     3901 cgagcacgta ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat
     3961 caggggctcg cgccagccga actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag
     4021 gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc
     4081 ttttctggat tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg
     4141 ttggctaccc gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg
     4201 ctttacggta tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag
     4261 ttcttctgag cgggactctg gggttcgaaa tgaccgacca agcgacgccc aacctgccat
     4321 cacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc
     4381 gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc
     4441 ccaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca
     4501 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat
     4561 cttatcatgt ctgtataccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc
     4621 tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca
     4681 taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct
     4741 cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac
     4801 gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc
     4861 tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt
     4921 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
     4981 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
     5041 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
     5101 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
     5161 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
     5221 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
     5281 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
     5341 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
     5401 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag
     5461 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
     5521 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
     5581 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
     5641 agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca
     5701 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa
     5761 cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
     5821 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
     5881 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
     5941 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
     6001 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
     6061 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg
     6121 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
     6181 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
     6241 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
     6301 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
     6361 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa
     6421 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
     6481 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
     6541 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
     6601 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa
     6661 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
     6721 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc gacggatcgg
     6781 gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt
     6841 aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg cgagcaaaat
     6901 ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc ttagggttag
     6961 gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgtt
//
