LOCUS       Exported                6930 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Highly resposive GFP-based pyruvate nanosensor..
ACCESSION   .
VERSION     .
KEYWORDS    PyronicSF/pcDNA3.1(-)
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6930)
  AUTHORS   Arce-Molina R, Cortes-Molina F, Sandoval PY, Galaz A, Alegria K, 
            Schirmeier S, Barros LF, San Martin A
  TITLE     A highly responsive pyruvate sensor reveals pathway-regulatory role 
            of the mitochondrial pyruvate carrier MPC.
  JOURNAL   Elife. 2020 Mar 6;9. pii: 53917. doi: 10.7554/eLife.53917.
  PUBMED    32142409
REFERENCE   2  (bases 1 to 6930)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6930
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2521..2538)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2527..2751
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2797..3225
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2884..2903)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3094..3115
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3234..3254)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3239..3568
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3419..3554
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3481..3500
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3635..4429
                     /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(3689..3708)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4299..4318
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4603..4724
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4640..4659)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4694..4713
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4773..4789)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4773..4789)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4786..4808)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4797..4813
                     /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(4821..4851)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4866..4887
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5004..5021)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5175..5763)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5255..5274)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5934..6794)
                     /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     6557..6576
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6795..6899)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaacgg gccctctaga ctcgagcggc cgccactgtg ctggatatct gcagaattcc
      961 accacactgg actagtggat ccctcgagat ggcatatagc aaaattcggc agcccaaact
     1021 gagcgatgtg attgagcagc agctggagtt tctgattctg gaaggcaccc tgagacctgg
     1081 agagaaactg ccccctgagc gcgaactcgc caagcagttc gacgtgagtc gaccatcact
     1141 gagggaggct atccagaggc tggaagcaaa gggactgctc ctgaggagac agggaggagg
     1201 gactttcgtg cagagctccc tgtggcagag cttcagcgac cccctggtcg agctcctgtc
     1261 tgaccaccca gaaagtcagt acgatctcct ggagacaaga catgctctgg aaggcatcgc
     1321 cgcttactat gcagccctgc ggtccactga cgaggataag gaacgcatcc gagagctgca
     1381 ccatgccatt gaactcgctc agcagtcagg agacctggat gcagagagca acgccgtgct
     1441 gcagtaccag attgcagtca ccgaggctgc acacaatgtg gtcctcctgc atctcctgag
     1501 gtgcatggag ccaatgctgg cccagaacgt gagacagaat tttgagctcc tgctcgaaaa
     1561 cgtctatatc aaggccgaca agcagaaaaa cggcattaag gctaacttca agatcagaca
     1621 caacatcgag gatggtggcg tgcagctggc ctaccattat cagcagaaca caccaatcgg
     1681 agacggacca gtgctgctcc cagataatca ctacctgagc gtccagtcca agctgtctaa
     1741 agaccctaac gagaagcggg atcatatggt gctgctcgaa tttgtcacag ccgctgggat
     1801 cactctgggt atggacgagc tctataaagg agggaccggt ggcagtatgg tgtcaaaggg
     1861 cgaggaactg ttcacaggag tggtccccat tctggtggag ctcgacggcg atgtcaatgg
     1921 acacaaattt tccgtgtctg gcgagggcga aggagatgct acctacggga agctgacact
     1981 caaattcatc tgcaccacag gcaagctgcc agtgccctgg cctactctgg tcactaccct
     2041 cacctacggg gtgcagtgtt tctccagata tcccgaccac atgaagcagc atgatttctt
     2101 taaatctgct atgcctgagg ggtacatcca ggaacggaca attttcttta aggacgatgg
     2161 taactacaaa acacgcgcag aggtgaagtt cgaaggcgac actctggtca atcgaatcga
     2221 gctgaaggga attgacttta aagaagatgg gaacatcctg ggtcacaagc tggagtacaa
     2281 tactaggtat tctcggcgcg aaatgctgcc actcgtgtct agtcacagga ccagaatctt
     2341 tgaggcaatt atggccggaa agcccgagga agctagagaa gcaagtcacc ggcatctggc
     2401 cttcatcgag gaaattctgc tcgaccggag ccgcgaggaa tcccgaaggg agcgcagcct
     2461 gagacgactc gaacagcgaa agaactgact cgagaagctt aagtttaaac cgctgatcag
     2521 cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct
     2581 tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc
     2641 attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg
     2701 aggattggga agacaatagc aggcatgctg gggatgcggt gggctctatg gcttctgagg
     2761 cggaaagaac cagctggggc tctagggggt atccccacgc gccctgtagc ggcgcattaa
     2821 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc
     2881 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
     2941 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     3001 aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
     3061 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
     3121 cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct
     3181 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattaattc tgtggaatgt
     3241 gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat
     3301 gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag
     3361 tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat
     3421 cccgccccta actccgccca gttccgccca ttctccgccc catggctgac taattttttt
     3481 tatttatgca gaggccgagg ccgcctctgc ctctgagcta ttccagaagt agtgaggagg
     3541 cttttttgga ggcctaggct tttgcaaaaa gctcccggga gcttgtatat ccattttcgg
     3601 atctgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc
     3661 aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat
     3721 cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt
     3781 caagaccgac ctgtccggtg ccctgaatga actgcaggac gaggcagcgc ggctatcgtg
     3841 gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag
     3901 ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc
     3961 tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc
     4021 tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga
     4081 agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga
     4141 actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg
     4201 cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg
     4261 tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc
     4321 tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc
     4381 cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg
     4441 gggttcgaaa tgaccgacca agcgacgccc aacctgccat cacgagattt cgattccacc
     4501 gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc
     4561 ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct
     4621 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     4681 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg
     4741 tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt
     4801 tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt
     4861 gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg
     4921 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg
     4981 cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
     5041 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
     5101 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
     5161 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
     5221 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
     5281 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
     5341 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg
     5401 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
     5461 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
     5521 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
     5581 ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg
     5641 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
     5701 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
     5761 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
     5821 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa
     5881 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa
     5941 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
     6001 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct
     6061 gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca
     6121 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
     6181 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
     6241 gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
     6301 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
     6361 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
     6421 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
     6481 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
     6541 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
     6601 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
     6661 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
     6721 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
     6781 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
     6841 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc
     6901 acatttcccc gaaaagtgcc acctgacgtc
//
