LOCUS       sequence-465590-        7480 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses the G-PTEN sensor under a CAG promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7480)
  AUTHORS   Kagan T, Gabay M, Meenakshisundaram A, Levi Y, Eid S, Malchenko N, 
            Maman M, Nitzan A, Ravotto L, Zaidel-Bar R, Eickholt BJ, Gal M, 
            Laviv T
  TITLE     Genetically encoded biosensor for fluorescence lifetime imaging of 
            PTEN dynamics in the intact brain.
  JOURNAL   Nat Methods. 2025 Apr;22(4):764-777. doi: 
            10.1038/s41592-025-02610-9. Epub 2025 Feb 20.
  PUBMED    39979596
REFERENCE   2  (bases 1 to 7480)
  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: 
            "10.1038/s41592-025-02610-9"; journalName: "Nat Methods"; date: 
            "2025-04"; volume: "22"; issue: "4"; pages: "764-777"
COMMENT     Sequence Label: CAG_G-PTEN
FEATURES             Location/Qualifiers
     source          1..7480
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        9..388
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        390..665
                     /label=chicken beta-actin promoter
     intron          666..1686
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1606..1628
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1694..1713
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1757..1766
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1763..1765,1766..1768,1769..2479)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(3719..3721,3722..4432)
                     /codon_start=1
                     /product="enhanced variant of YFP (Heikal et al., 2001)"
                     /label=Citrine
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     LICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNWNSVNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     RVTAAGITLGMDELYK"
     primer_bind     complement(4502..4521)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4567..4622
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4621..4640)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(4983..4999)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4983..4999)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4996..5018)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5007..5023
                     /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(5031..5037,5038..5055,5056..5061))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5076..5097
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        5156..5351
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      5202..5337
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    5357..5491
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5394..5413)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5448..5467
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5559..5576)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5730..6318)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5810..5829)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6489..7280,7281..7349))
                     /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     7112..7131
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7350..7454)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggtcgatcga cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt
       61 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
      121 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
      181 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
      241 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
      301 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat
      361 ctacgtatta gtcatcgcta ttaccatggt cgaggtgagc cccacgttct gcttcactct
      421 ccccatctcc cccccctccc cacccccaat tttgtattta tttatttttt aattattttg
      481 tgcagcgatg ggggcggggg gggggggggg gcgcgccagg cggggcgggg cggggcgagg
      541 ggcggggcgg ggcgaggcgg agaggtgcgg cggcagccaa tcagagcggc gcgctccgaa
      601 agtttccttt tatggcgagg cggcggcggc ggcggcccta taaaaagcga agcgcgcggc
      661 gggcgggagt cgctgcgcgc tgccttcgcc ccgtgccccg ctccgccgcc gcctcgcgcc
      721 gcccgccccg gctctgactg accgcgttac tcccacaggt gagcgggcgg gacggccctt
      781 ctcctccggg ctgtaattag cgcttggttt aatgacggct tgtttctttt ctgtggctgc
      841 gtgaaagcct tgaggggctc cgggagggcc ctttgtgcgg ggggagcggc tcggggggtg
      901 cgtgcgtgtg tgtgtgcgtg gggagcgccg cgtgcggctc cgcgctgccc ggcggctgtg
      961 agcgctgcgg gcgcggcgcg gggctttgtg cgctccgcag tgtgcgcgag gggagcgcgg
     1021 ccgggggcgg tgccccgcgg tgcggggggg gctgcgaggg gaacaaaggc tgcgtgcggg
     1081 gtgtgtgcgt gggggggtga gcagggggtg tgggcgcgtc ggtcgggctg caaccccccc
     1141 tgcacccccc tccccgagtt gctgagcacg gcccggcttc gggtgcgggg ctccgtacgg
     1201 ggcgtggcgc ggggctcgcc gtgccgggcg gggggtggcg gcaggtgggg gtgccgggcg
     1261 gggcggggcc gcctcgggcc ggggagggct cgggggaggg gcgcggcggc ccccggagcg
     1321 ccggcggctg tcgaggcgcg gcgagccgca gccattgcct tttatggtaa tcgtgcgaga
     1381 gggcgcaggg acttcctttg tcccaaatct gtgcggagcc gaaatctggg aggcgccgcc
     1441 gcaccccctc tagcgggcgc ggggcgaagc ggtgcggcgc cggcaggaag gaaatgggcg
     1501 gggagggcct tcgtgcgtcg ccgcgccgcc gtccccttct ccctctccag cctcggggct
     1561 gtccgcgggg ggacggctgc cttcgggggg gacggggcag ggcggggttc ggcttctggc
     1621 gtgtgaccgg cggctctagc tagagcctct gctaaccatg ttcatgcctt cttctttttc
     1681 ctacagctcc tgggcaacgt gctggttatt gtgctgtctc atcattttgg caaagaattc
     1741 gctagcgcta ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt
     1801 gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga
     1861 gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa
     1921 gctgcccgtg ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag
     1981 ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta
     2041 cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt
     2101 gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga
     2161 ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat
     2221 catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga
     2281 ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc
     2341 cgtgctgctg cccgacaacc actacctgag cacccagtcc aagctgagca aagaccccaa
     2401 cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg
     2461 catggacgag ctgtacaagt ccggactcag atctcgagct atgacagcca tcatcaaaga
     2521 gatcgttagc agaaacaaag ggagatatca agaggatgga ttcgacttag acttgaccta
     2581 tatttatcca aatattattg ctatgggatt tcctgcagaa agacttgaag gtgtatacag
     2641 gaacaatatt gatgatgtag taaggttttt ggattcaaag cataaaaacc attacaagat
     2701 atacaatcta tgtgctgaga gacattatga caccgccaaa tttaactgca gagttgcaca
     2761 gtatcctttt gaagaccata acccaccaca gctagaactt atcaaaccct tttgtgaaga
     2821 tcttgaccaa tggctaagtg aagacgacaa tcatgttgca gcaattcact gtaaagctgg
     2881 gaaaggacgg actggtgtaa tgatttgtgc atatttattg catcggggca agtttttaaa
     2941 ggcacaagag gccctggatt tttatgggga agtaaggacc agagataaaa agggagtaac
     3001 tattcccagt cagaggcgct atgtatatta ttatagctac ctgttaaaga atcacctgga
     3061 ttacagacca gtggcactgt tgtttcacaa gatgatgttt gaaactattc caatgttcag
     3121 tggcggaact tgcaatcccc agtttgtggt ctgccagcta aaggtgaaga tctactcctc
     3181 caactcagga cccacgcggc gggaggacaa gctcatgtac tttgagttcc ctcagccatt
     3241 gcctgtgtgt ggtgacatca aagtagagtt cttccacaaa cagaacaaga tgctcaaaaa
     3301 ggacaaaatg tttcactttt gggtaaatac gttcttcata ccaggaccag aggaaacctc
     3361 agaaaaagtg gaaaatggaa gtctttgtga tcaggaaatc gatagcattt gtagtataga
     3421 gcgtgcggat aatgacaagg agtatcttgt gctcaccctg acaaaaaatg atcttgacaa
     3481 agcaaacaaa gacaaggcca accgatactt ctctccaaat tttaaggtga aattatactt
     3541 tacaaaaaca gtagaggaac catcaaatcc agaggctagc agttcaactt ctgtgactcc
     3601 agacgttagt gacaatgaac ctgatcatta tagatattct gacaccactg actctgatcc
     3661 agagaatgaa ccttttgatg aagatcagca ttcacaaatt acaaaagtcc ctacccctgt
     3721 gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga
     3781 cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa
     3841 gctgaccctg aagctcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt
     3901 gaccaccttc ggctacggcc tgatgtgctt cgcccgctac cccgaccaca tgaagcagca
     3961 cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa
     4021 ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa
     4081 ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct
     4141 ggagtacaac tggaacagcg tcaacgtcta tatcatggcc gacaagcaga agaacggcat
     4201 caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca
     4261 ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct
     4321 gagctaccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct
     4381 ggagagagtg accgccgccg ggatcactct cggcatggac gagctgtaca agtgaagata
     4441 actgatcata atcagccgcg ggcccgggat cctaagcggc cgcgactcta gagtcgacgg
     4501 atctttttcc ctctgccaaa aattatgggg acatcatgaa gccccttgag catctgactt
     4561 ctggctaata aaggaaattt attttcattg caatagtgtg ttggaatttt ttgtgtctct
     4621 cactcggaag gacatatggg agggcaaatc atttaaaaca tcagaatgag tatttggttt
     4681 agagtttggc aacatatgcc catatgctgg ctgccatgaa caaaggttgg ctataaagag
     4741 gtcatcagta tatgaaacag ccccctgctg tccattcctt attccataga aaagccttga
     4801 cttgaggtta gatttttttt atattttgtt ttgtgttatt tttttcttta acatccctaa
     4861 aattttcctt acatgtttta ctagccagat ttttcctcct ctcctgacta ctcccagtca
     4921 tagctgtccc tcttctctta tggagatccc tcgacctgca gcccaagctt ggcgtaatca
     4981 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     5041 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     5101 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagcg gatccgcatc
     5161 tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc
     5221 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     5281 aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     5341 gcttttgcaa aaagctaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     5401 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     5461 actcatcaat gtatcttatc atgtctggat ccgctgcatt aatgaatcgg ccaacgcgcg
     5521 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     5581 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     5641 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     5701 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     5761 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     5821 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     5881 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     5941 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     6001 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     6061 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     6121 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     6181 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     6241 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     6301 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     6361 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     6421 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     6481 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     6541 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     6601 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     6661 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     6721 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     6781 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     6841 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     6901 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     6961 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     7021 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     7081 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     7141 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     7201 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     7261 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     7321 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     7381 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     7441 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
//
