LOCUS       sequence-371361-        7230 bp    DNA     circular SYN 15-DEC-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 7230)
  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 7230)
  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: "ACS Sens. 
            2022 Oct 28. doi: 10.1021/acssensors.2c00731."
COMMENT     Sequence Label: CanlonicSF
FEATURES             Location/Qualifiers
     source          1..7230
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(41..58)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    47..271
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      317..745
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(404..423)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     614..635
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(754..774)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        759..1088
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      939..1074
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1001..1020
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             1155..1949
                     /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(1209..1228)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1819..1838
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    2123..2244
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2160..2179)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2214..2233
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2293..2309)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2293..2309)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2306..2328)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2317..2333
                     /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(2341..2347,2348..2365,2366..2371))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2386..2407
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2524..2541)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2695..3283)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2775..2794)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3454..4245,4246..4314))
                     /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     4077..4096
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4315..4419)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(4494..4513)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        4685..5064
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        5065..5268
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5219..5239
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     5313..5332
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5313..5331
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5434..5458
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             6016..6267
                     /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     6204..6225
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(6331..6352)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(6592..6611)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     protein_bind    complement(join(7220..7230,1..14))
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
ORIGIN
        1 ttgtacaaag tggtaagctt aagtttaaac cgctgatcag cctcgactgt gccttctagt
       61 tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact
      121 cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat
      181 tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc
      241 aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac cagctggggc
      301 tctagggggt atccccacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt
      361 acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc
      421 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct
      481 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat
      541 ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc
      601 acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc
      661 tattcttttg atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg
      721 atttaacaaa aatttaacgc gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa
      781 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
      841 ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
      901 attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca
      961 gttccgccca ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg
     1021 ccgcctctgc ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct
     1081 tttgcaaaaa gctcccggga gcttgtatat ccattttcgg atctgatcaa gagacaggat
     1141 gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg
     1201 tggagaggct attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg
     1261 tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg
     1321 ccctgaatga actgcaggac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc
     1381 cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg
     1441 aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca
     1501 tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc
     1561 aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg
     1621 atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg
     1681 cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata
     1741 tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg
     1801 accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat
     1861 gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct
     1921 tctatcgcct tcttgacgag ttcttctgag cgggactctg gggttcgaaa tgaccgacca
     1981 agcgacgccc aacctgccat cacgagattt cgattccacc gccgccttct atgaaaggtt
     2041 gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat
     2101 gctggagttc ttcgcccacc ccaacttgtt tattgcagct tataatggtt acaaataaag
     2161 caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt
     2221 gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta gctagagctt
     2281 ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca
     2341 caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact
     2401 cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct
     2461 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc
     2521 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca
     2581 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
     2641 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     2701 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     2761 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     2821 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     2881 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     2941 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     3001 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     3061 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     3121 cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     3181 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     3241 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     3301 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     3361 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     3421 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     3481 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     3541 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
     3601 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     3661 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     3721 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
     3781 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     3841 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     3901 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     3961 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     4021 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     4081 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     4141 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     4201 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     4261 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
     4321 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
     4381 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     4441 acctgacgtc gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc
     4501 tgctctgatg ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct
     4561 gagtagtgcg cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg
     4621 aagaatctgc ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg
     4681 cgttgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat
     4741 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg
     4801 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata
     4861 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta
     4921 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc
     4981 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac
     5041 gtattagtca tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga
     5101 tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg
     5161 ttttggcacc aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg
     5221 caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact
     5281 agagaaccca ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa
     5341 gctggctagc gtttaaacgg gccctctaga ctcgagcggc cgccactgtg ctggatatct
     5401 gcagaattcc accacactgg actagtggat cccacaagtt tgtacaaaaa agcaggctac
     5461 atgttttctc ctctggcggt ggcccaagcg aggcgctacc gttggcgcat ccagaccgct
     5521 tgggatgcgg gcacggtagg ctattccctg ttccagaagt tcaccgagcg tgttaaggag
     5581 ctcaccgacg gccagttgga ggttcagccc ttccccgcag gggccgtggt gggcaccttt
     5641 gacatgtttg acgcggtgaa gacgggcgtt ttggacggga tgaacccctt caccctcgca
     5701 tgggcggggc ggatgccggt gacggccttt ctttcctcct atgccctggg tttggaccgg
     5761 cccgaccagt gggagacctg gttctacagc ctgggcggcc tggacatcgc ccggagggcc
     5821 ttcgccgagc aggggctttt ctacgtgggc cccgtgcaac acgatttgaa catcatccac
     5881 tccaaaaagc cgatccgtcg ctttgaggat ttcaaagggg taaagctgcg ggtgccgggc
     5941 ggaatgattg ctgaagtctt tgccgccgcg ggggcctcca cggtccttct actagaaaac
     6001 gtctatatca aggccgacaa gcagaagaac ggcatcaagg cgaacttcaa gatccgccac
     6061 aacatcgagg acggcggcgt gcagctcgcc taccactacc agcagaacac ccccatcggc
     6121 gacggccccg tgctgctgcc cgacaaccac tacctgagcg tccagtccaa gctgagcaaa
     6181 gaccccaacg agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc
     6241 actctcggca tggacgagct gtacaagggt ggtaccggtg gatctatggt gagcaagggc
     6301 gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc
     6361 cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa gctgaccctg
     6421 aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccctg
     6481 acctacggcg tgcagtgctt cagccgctac cccgaccaca tgaagcagca cgacttcttc
     6541 aagtccgcca tgcccgaagg ctacatccag gagcgcacca tcttcttcaa ggacgacggc
     6601 aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag
     6661 ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac
     6721 accagacccg gaggcgaggt ctatcctgcc ttggagcgtg gggtcattga tgcggccgac
     6781 tttgtggggc ctgcggtcaa ctacaatctg ggcttccacc aggtggccaa gtacatcatt
     6841 atggggcccc cggagacccc ggccatccat cagccggtgg accttatgga cttcaccatc
     6901 aacctcaatc ggtggcggtc gctacccaag cctttgcaag agcgtttcat cgccgctgtt
     6961 cacgaataca gttggattca ctacgccggg attcaaaagg ccaacctcga ggcctggccc
     7021 aagtaccgcc aggcgggtgt ggaggtcatc cgcctgtcta acgaggacgt gcgcaagttc
     7081 cgccgtctgg ccatccccat ctggttcaag tgggccaaga tggacaagta ctcgcgggag
     7141 gccttcgcca gccagctgga gtacatgaag ggcatcggct acgttacgga cgaggagctc
     7201 aaggggcttt ccctttagga cccagctttc
//
