LOCUS       sequence-460265-        5752 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of mouse MPC1 in which K45 and K46 motif sites were 
            replaced with alanine blocking acetylation with a C terminal FLAG 
            tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5752)
  AUTHORS   Tiwari A, Myeong J, Hashemiaghdam A, Stunault MI, Zhang H, Niu X, 
            Laramie MA, Sponagel J, Shriver LP, Patti GJ, Klyachko VA, Ashrafi G
  TITLE     Mitochondrial pyruvate transport regulates presynaptic metabolism 
            and neurotransmission.
  JOURNAL   Sci Adv. 2024 Nov 15;10(46):eadp7423. doi: 10.1126/sciadv.adp7423. 
            Epub 2024 Nov 15.
  PUBMED    39546604
REFERENCE   2  (bases 1 to 5752)
  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: "Sci Adv."; 
            date: "2024-11-15"; volume: "10(46):eadp7423. doi"; pages: " 
            10.1126/sciadv.adp7423. Epub 2024 Nov 15"
COMMENT     Sequence Label: MPC1-AA-FLAG
FEATURES             Location/Qualifiers
     source          1..5752
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     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"
     regulatory      937..946
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1279..1302
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(1343..1360)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1349..1573
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1619..2047
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1706..1725)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1916..1937
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2056..2076)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2061..2390
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2241..2376
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2303..2322
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2457..3251
                     /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(2511..2530)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3121..3140
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3425..3546
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3462..3481)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3516..3535
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3595..3611)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3595..3611)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3608..3630)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3619..3635
                     /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(3643..3649,3650..3667,3668..3673))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3688..3709
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3826..3843)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3997..4585)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4077..4096)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4756..5547,5548..5616))
                     /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     5379..5398
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5617..5721)
                     /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 gtttaaactt aagcttggta ccgagctcgg atccccgcca ccatggcagg cgctctcgtt
      961 agaaaggccg ccgactacgt gcagagcaag gacttcaggg actacctgat gtctacccac
     1021 ttctggggcc ctgtggctaa ctggggactg cctatcgccg ccatcaacga catggctgcc
     1081 tctccagaga tcatcagcgg caggatgaca ttcgccctgt gctgctacag cctgaccttc
     1141 atgagattcg cctacaaggt gcagcctagg aactggctgc tgttcgcctg ccacgtgaca
     1201 aatgaagtgg cccagctgat ccaaggcggc aggctgatca actacgagat gagcaagagg
     1261 ccctctgccg gcggaggcga ctacaaggac gacgacgata agtgactcga gtctagaggg
     1321 cccgtttaaa cccgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt
     1381 tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa
     1441 taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg
     1501 gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg
     1561 gtgggctcta tggcttctga ggcggaaaga accagctggg gctctagggg gtatccccac
     1621 gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct
     1681 acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg
     1741 ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt
     1801 gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca
     1861 tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga
     1921 ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa
     1981 gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac
     2041 gcgaattaat tctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag
     2101 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc
     2161 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag
     2221 tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc
     2281 cccatggctg actaattttt tttatttatg cagaggccga ggccgcctct gcctctgagc
     2341 tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctcccgg
     2401 gagcttgtat atccattttc ggatctgatc aagagacagg atgaggatcg tttcgcatga
     2461 ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct
     2521 atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc
     2581 aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcagg
     2641 acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg
     2701 acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc
     2761 tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc
     2821 ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg
     2881 agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc
     2941 atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg
     3001 aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc
     3061 gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag
     3121 cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg
     3181 tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg
     3241 agttcttctg agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc
     3301 atcacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt
     3361 ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca
     3421 ccccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
     3481 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
     3541 atcttatcat gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata
     3601 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     3661 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     3721 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     3781 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     3841 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     3901 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     3961 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     4021 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     4081 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     4141 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     4201 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     4261 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     4321 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     4381 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     4441 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     4501 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     4561 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     4621 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     4681 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     4741 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     4801 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     4861 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     4921 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     4981 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     5041 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     5101 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     5161 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     5221 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     5281 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     5341 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     5401 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     5461 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     5521 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     5581 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     5641 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     5701 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tc
//
