LOCUS       sequence-87749-e        7447 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses S586A VLCAD mutant in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7447)
  AUTHORS   Kabuyama Y, Suzuki T, Nakazawa N, Yamaki J, Homma MK, Homma Y
  TITLE     Dysregulation of very long chain acyl-CoA dehydrogenase coupled with
            lipid peroxidation.
  JOURNAL   Am J Physiol Cell Physiol. 2010 Jan;298(1):C107-13. doi: 
            10.1152/ajpcell.00231.2009. Epub 2009 Nov 4.
  PUBMED    19889959
REFERENCE   2  (bases 1 to 7447)
  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: 
            "10.1152/ajpcell.00231.2009"; journalName: "Am J Physiol Cell 
            Physiol"; date: "2010-01"; volume: "298"; issue: "1"; pages: 
            "C107-13"
COMMENT     Sequence Label: pcDNA3-S586A-VLCAD-FLAG
FEATURES             Location/Qualifiers
     source          1..7447
                     /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      2918..2927
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2927..2950
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        complement(2999..3017)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3000..3017)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3037..3054)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3043..3267
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3313..3741
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3400..3419)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3610..3631
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3750..3770)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3755..4085
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3936..4071
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3998..4017
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4152..4946
                     /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(4206..4225)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4816..4835
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5120..5241
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5157..5176)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5211..5230
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5290..5306)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5290..5306)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5303..5325)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5314..5330
                     /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(5338..5344,5345..5362,5363..5368))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5383..5404
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5521..5538)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5692..6280)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5772..5791)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6451..7242,7243..7311))
                     /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     7074..7093
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7312..7416)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtcgactct 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 atgggtggac 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 gcttggtacc
      901 gagctcggat ccactagtaa cggccgccag tgtgctggaa ttcatgcagg cggctcggat
      961 ggccgcgagc ttggggcggc agctgctgag gctcgggggc ggaagctcgc ggctcacggc
     1021 gctcctgggg cagccccggc ccggccctgc ccggcggccc tatgccgggg gtgccgctca
     1081 gctggctctg gacaagtcag attcccaccc ctctgacgct ctgaccagga aaaaaccggc
     1141 caaggcggaa tctaagtcct ttgctgtggg aatgttcaaa ggccagctca ccacagatca
     1201 ggtgttccca tacccgtccg tgctcaacga agagcagaca cagtttctta aagagctggt
     1261 ggagcctgtg tcccgtttct tcgaggaagt gaacgatccc gccaagaatg acgctctgga
     1321 gatggtggag gagaccactt ggcagggcct caaggagctg ggggcctttg gtctgcaagt
     1381 gcccagtgag ctgggtggtg tgggcctttg caacacccag tacgcccgtt tggtggagat
     1441 cgtgggcatg catgaccttg gcgtgggcat taccctgggg gcccatcaga gcatcggttt
     1501 caaaggcatc ctgctctttg gcacaaaggc ccagaaagaa aaatacctcc ccaagctggc
     1561 atctggggag actgtggccg ctttctgtct aaccgagccc tcaagcgggt cagatgcagc
     1621 ctccatccga acctctgctg tgcccagccc ctgtggaaaa tactataccc tcaatggaag
     1681 caagctttgg atcagtaatg ggggcctagc agacatcttc acggtctttg ccaagacacc
     1741 agttacagat ccagccacag gagccgtgaa ggagaagatc acagcttttg tggtggagag
     1801 gggcttcggg ggcattaccc atgggccccc tgagaagaag atgggcatca aggcttcaaa
     1861 cacagcagag gtgttctttg atggagtacg ggtgccatcg gagaacgtgc tgggtgaggt
     1921 tgggagtggc ttcaaggttg ccatgcacat cctcaacaat ggaaggtttg gcatggctgc
     1981 ggccctggca ggtaccatga gaggcatcat tgctaaggcg gtagatcatg ccactaatcg
     2041 tacccagttt ggggagaaaa ttcacaactt tgggctgatc caggagaagc tggcacggat
     2101 ggttatgctg cagtatgtaa ctgagtccat ggcttacatg gtgagtgcta acatggacca
     2161 gggagccacg gacttccaga tagaggccgc catcagcaaa atctttggct cggaggcagc
     2221 ctggaaggtg acagatgaat gcatccaaat catggggggt atgggcttca tgaaggaacc
     2281 tggagtagag cgtgtgctcc gagatcttcg catcttccgg atctttgagg ggacaaatga
     2341 cattcttcgg ctgtttgtgg ctctgcaggg ctgtatggac aaaggaaagg agctctctgg
     2401 gcttggcagt gctctaaaga atccctttgg gaatgctggc ctcctgctag gagaggcagg
     2461 caaacagctg aggcggcggg cagggctggg cagcggcctg agtctcagcg gacttgtcca
     2521 cccggagttg agtcggagtg gcgagctggc agtacgggct ctggagcagt ttgccactgt
     2581 ggtggaggcc aagctgataa aacacaagaa ggggattgtc aatgaacagt ttctgctgca
     2641 gcggctggca gacggggcca tcgacctcta tgccatggtg gtggttctct cgagggccgc
     2701 aagatccctg agtgagggcc accccacggc ccagcatgag aaaatgctct gtgacacctg
     2761 gtgtatcgag gctgcagctc ggatccgaga gggcatggcc gccctgcagt ctgacccctg
     2821 gcagcaagag ctctaccgca acttcaaaag catctccaag gccttggtgg agcggggtgg
     2881 tgtggtcacc agcaacccac ttggcttcag agcggccgcc accatggatt acaaggatga
     2941 cgacgataag agcccgggcg gatcctctag agtcgagcat gcatctagag ggccctattc
     3001 tatagtgtca cctaaatgct agagctcgct gatcagcctc gactgtgcct tctagttgcc
     3061 agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca
     3121 ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta
     3181 ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc
     3241 atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc tggggctcta
     3301 gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
     3361 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt
     3421 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggc atccctttag
     3481 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt
     3541 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
     3601 tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt
     3661 cttttgattt ataagggatt ttggggattt cggcctattg gttaaaaaat gagctgattt
     3721 aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt cagttagggt gtggaaagtc
     3781 cccaggctcc ccaggcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     3841 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     3901 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt
     3961 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg
     4021 cctctgcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt
     4081 gcaaaaagct cccgggagct tgtatatcca ttttcggatc tgatcaagag acaggatgag
     4141 gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg
     4201 agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt
     4261 tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc
     4321 tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt
     4381 gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag
     4441 tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg
     4501 ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag
     4561 cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg
     4621 atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc
     4681 gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca
     4741 tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc
     4801 gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg
     4861 ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct
     4921 atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc
     4981 gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg
     5041 cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct
     5101 ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa
     5161 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
     5221 caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc
     5281 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
     5341 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
     5401 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
     5461 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
     5521 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
     5581 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     5641 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     5701 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     5761 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     5821 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     5881 ttctcaatgc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     5941 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     6001 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     6061 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     6121 ctacactaga aggacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     6181 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     6241 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     6301 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     6361 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     6421 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     6481 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     6541 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg
     6601 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     6661 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     6721 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     6781 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     6841 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     6901 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     6961 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     7021 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     7081 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     7141 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     7201 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     7261 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     7321 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     7381 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     7441 tgacgtc
//
