LOCUS       sequence-371413-        6739 bp    DNA     circular SYN 14-APR-2025
DEFINITION  To explore lactate dynamics in intact systems.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6739)
  AUTHORS   Aburto C, Galaz A, Bernier A, Sandoval PY, Holtheuer-Gallardo S, 
            Ruminot I, Soto-Ojeda I, Hertenstein H, Schweizer JA, Schirmeier S, 
            Pastor TP, Mardones GA, Barros LF, San Martin A
  TITLE     Single-Fluorophore Indicator to Explore Cellular and Sub-cellular 
            Lactate Dynamics.
  JOURNAL   ACS Sens. 2022 Oct 28. doi: 10.1021/acssensors.2c00731.
  PUBMED    36306435
REFERENCE   2  (bases 1 to 6739)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "ACS Sens. 
            2022 Oct 28. doi: 10.1021/acssensors.2c00731."
COMMENT     Sequence Label: NLS-CanlonicSF
FEATURES             Location/Qualifiers
     source          1..6739
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        12..388
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        390..593
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     544..564
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      650..659
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1235..1486
                     /codon_start=1
                     /product="C-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VC155
                     /translation="DKQKNGIKANFKIRHNIEDGGVQLAYHYQQNTPIGDGPVLLPDNH
                     YLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYK"
     primer_bind     1423..1444
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1550..1571)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1811..1830)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     CDS             2450..2470
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2474..2494
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2498..2518
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2531..2560
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(2596..2613)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2602..2826
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2872..3300
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2959..2978)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3169..3190
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3309..3329)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3314..3643
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3494..3629
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3556..3575
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3669..4463
                     /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="MVEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3723..3742)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4333..4352
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4637..4758
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4674..4693)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4728..4747
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4791..4808)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4962..5550)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5042..5061)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5721..6512,6513..6581))
                     /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     6344..6363
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6582..6686)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtaccgaatt cacattgatt attgagtagt tattaatagt aatcaattac ggggtcatta
       61 gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc
      121 tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg
      181 ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg
      241 gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa
      301 tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac
      361 atctacggtt agtcatcgct attaccatag tgatgcggtt ttggcagtac atcaatgggc
      421 gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga
      481 gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat
      541 tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctttctggc
      601 taactagaga acccactgct tactggcacg tggaaattaa tacgacgtgg ccaccatggc
      661 ccaggtgcag ctgcagacca tgttttctcc tctggcggtg gcccaagcga ggcgctaccg
      721 ttggcgcatc cagaccgctt gggatgcggg cacggtaggc tattccctgt tccagaagtt
      781 caccgagcgt gttaaggagc tcaccgacgg ccagttggag gttcagccct tccccgcagg
      841 ggccgtggtg ggcacctttg acatgtttga cgcggtgaag acgggcgttt tggacgggat
      901 gaaccccttc accctcgcat gggcggggcg gatgccggtg acggcctttc tttcctccta
      961 tgccctgggt ttggaccggc ccgaccagtg ggagacctgg ttctacagcc tgggcggcct
     1021 ggacatcgcc cggagggcct tcgccgagca ggggcttttc tacgtgggcc ccgtgcaaca
     1081 cgatttgaac atcatccact ccaaaaagcc gatccgtcgc tttgaggatt tcaaaggggt
     1141 aaagctgcgg gtgccgggcg gaatgattgc tgaagtcttt gccgccgcgg gggcctccac
     1201 ggtccttcta ctagaaaacg tctatatcaa ggccgacaag cagaagaacg gcatcaaggc
     1261 gaacttcaag atccgccaca acatcgagga cggcggcgtg cagctcgcct accactacca
     1321 gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcgt
     1381 ccagtccaag ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt
     1441 cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaagggtg gtaccggtgg
     1501 atctatggtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct
     1561 ggacggcgac gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac
     1621 ctacggcaag ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc
     1681 caccctcgtg accaccctga cctacggcgt gcagtgcttc agccgctacc ccgaccacat
     1741 gaagcagcac gacttcttca agtccgccat gcccgaaggc tacatccagg agcgcaccat
     1801 cttcttcaag gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac
     1861 cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg
     1921 gcacaagctg gagtacaaca ccagacccgg aggcgaggtc tatcctgcct tggagcgtgg
     1981 ggtcattgat gcggccgact ttgtggggcc tgcggtcaac tacaatctgg gcttccacca
     2041 ggtggccaag tacatcatta tggggccccc ggagaccccg gccatccatc agccggtgga
     2101 ccttatggac ttcaccatca acctcaatcg gtggcggtcg ctacccaagc ctttgcaaga
     2161 gcgtttcatc gccgctgttc acgaatacag ttggattcac tacgccggga ttcaaaaggc
     2221 caacctcgag gcctggccca agtaccgcca ggcgggtgtg gaggtcatcc gcctgtctaa
     2281 cgaggacgtg cgcaagttcc gccgtctggc catccccatc tggttcaagt gggccaagat
     2341 ggacaagtac tcgcgggagg ccttcgccag ccagctggag tacatgaagg gcatcggcta
     2401 cgttacggac gaggagctca aggggctttc ccttgaggcg gccgcagatc caaaaaagaa
     2461 gagaaaggta gatccaaaaa agaagagaaa ggtagatcca aaaaagaaga gaaaggtaga
     2521 tacggccgca gaacaaaaac tcatctcaga agaggatctg aatggggccg catagtctag
     2581 aagctcgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct
     2641 cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg
     2701 aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc
     2761 aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat ggcccgggct
     2821 ctatggcttc tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct
     2881 gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg
     2941 ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg
     3001 gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac
     3061 ggcacctctc ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct
     3121 gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt
     3181 tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta taagggattt
     3241 tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt
     3301 aattctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag
     3361 aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc
     3421 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc
     3481 cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc cgcccctagg
     3541 ctgactaatt ttttttattt atgcagaggc cgaggccgcc tctgcctctg agctattcca
     3601 gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctcc ccccccggga
     3661 ggtccacaat ggttgaacaa gatggattgc acgcaggttc tccggccgct tgggtggaga
     3721 ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc gccgtgttcc
     3781 ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga
     3841 atgaactcca ggacgaggca gcgcggctat cgtggctggc cacgacgggc gttccttgcg
     3901 cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg ggcgaagtgc
     3961 cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc atcatggctg
     4021 atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac caccaagcga
     4081 aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat caggatgatc
     4141 tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgta
     4201 tgcccgacgg cgaggatctc gtcgtgactc atggcgatgc ctgcttgccg aatatcatgg
     4261 tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg gcggaccgct
     4321 atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc gaatgggctg
     4381 accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc gccttctatc
     4441 gccttcttga cgagttcttc tgagcgggac tctggggttc gaaatgaccg accaagcgac
     4501 gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt
     4561 cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga
     4621 gttcttcgcc caccccaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     4681 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     4741 actcatcaat gtatcttatc atgtctgtat accgtcgatc tttccgcttc ctcgctcact
     4801 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     4861 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     4921 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     4981 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     5041 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     5101 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     5161 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtatgcac
     5221 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     5281 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     5341 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     5401 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     5461 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     5521 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     5581 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     5641 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     5701 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     5761 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     5821 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     5881 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     5941 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     6001 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     6061 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     6121 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     6181 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     6241 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     6301 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     6361 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     6421 atcttaccgc tgttgagatc cagttcgatg taacccactc gggcacccaa ctgatcttca
     6481 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     6541 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     6601 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     6661 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtcaga
     6721 tcgacggatc gggagatcg
//
