LOCUS       sequence-136162-        7042 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Mammalian expression of flag-tagged PPAR gamma.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7042)
  AUTHORS   Hauser S, Adelmant G, Sarraf P, Wright HM, Mueller E, Spiegelman BM
  TITLE     Degradation of the peroxisome proliferator-activated receptor gamma 
            is linked to ligand-dependent activation.
  JOURNAL   J Biol Chem. 2000 Jun 16. 275(24):18527-33.
  PUBMED    10748014
REFERENCE   2  (bases 1 to 7042)
  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: "J Biol 
            Chem. 2000 Jun 16. 275(24):18527-33."
COMMENT     Sequence Label: pcDNA flag PPAR gamma
FEATURES             Location/Qualifiers
     source          1..7042
                     /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"
     CDS             986..1009
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2546..2575
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2591..2608
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2631..2648)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2637..2861
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2907..3335
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2994..3013)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3204..3225
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3344..3364)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3349..3678
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3529..3664
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3591..3610
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3745..4539
                     /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(3799..3818)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4409..4428
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4715..4836
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4752..4771)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4806..4825
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4885..4901)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4885..4901)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4898..4920)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4909..4925
                     /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(4933..4939,4940..4957,4958..4963))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4978..4999
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5116..5133)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5287..5875)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5367..5386)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6046..6837,6838..6906))
                     /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     6669..6688
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6907..7011)
                     /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 gctggctagt
      901 taagcttggt accgagctcg gatccactag tccagtgtgg tggaattctg cagatatcca
      961 gcacagtggc ggccgcacca ccatggacta caaagacgat gacgataaag caatgggtga
     1021 aactctggga gattctcctg ttgacccaga gcatggtgcc ttcgctgatg cactgcctat
     1081 gagcacttca caagaaatta ccatggttga cacagagatg ccattctggc ccaccaactt
     1141 cggaatcagc tctgtggacc tctccgtgat ggaagaccac tcgcattcct ttgacatcaa
     1201 gccctttacc acagttgatt tctccagcat ttctgctcca cactatgaag acattccatt
     1261 cacaagagct gacccaatgg ttgctgatta caaatatgac ctgaagctcc aagaatacca
     1321 aagtgcgatc aaagtagaac ctgcatctcc accttattat tctgaaaaga cccagctcta
     1381 caacaggcct catgaagaac cttctaactc cctcatggcc attgagtgcc gagtctgtgg
     1441 ggataaagca tcaggcttcc actatggagt tcatgcttgt gaaggatgca agggtttttt
     1501 ccgaagaacc atccgattga agcttattta tgataggtgt gatcttaact gccggatcca
     1561 caaaaaaagt agaaataaat gtcagtactg tcggtttcag aagtgccttg ctgtggggat
     1621 gtctcacaat gccatcaggt ttgggcggat gccacaggcc gagaaggaga agctgttggc
     1681 ggagatctcc agtgatatcg accagctgaa cccagagtct gctgatctgc gagccctggc
     1741 aaagcatttg tatgactcat acataaagtc cttcccgctg accaaagcca aggcgagggc
     1801 gatcttgaca ggaaagacaa cggacaaatc accatttgtc atctacgaca tgaattcctt
     1861 aatgatggga gaagataaaa tcaagttcaa acatatcacc cccctgcagg agcagagcaa
     1921 agaggtggcc atccgaattt ttcaagggtg ccagtttcga tccgtagaag ccgtgcaaga
     1981 gatcacagag tatgccaaaa atatccctgg tttcattaac cttgatttga atgaccaagt
     2041 gactctgctc aagtatggtg tccatgagat catctacacg atgctggcct ccctgatgaa
     2101 taaagatgga gtcctcatct cagagggcca aggattcatg accagggagt tcctcaaaag
     2161 cctgcggaag ccctttggtg actttatgga gcctaagttt gagtttgctg tgaagttcaa
     2221 tgcactggaa ttagatgaca gtgacttggc tatatttata gctgtcatta ttctcagtgg
     2281 agaccgccca ggcttgctga acgtgaagcc catcgaggac atccaagaca acctgctgca
     2341 ggccctggaa ctgcagctca agctgaatca cccagagtcc tctcagctgt tcgccaaggt
     2401 gctccagaag atgacagacc tcaggcagat cgtcacagag cacgtgcagc tactgcatgt
     2461 gatcaagaag acagagacag acatgagcct tcaccccctg ctccaggaga tctacaagga
     2521 cttgtattag tctagagggc ccttcgaaca aaaactcatc tcagaagagg atctgaatat
     2581 gcataccggt catcatcacc atcaccattg agtttaaacc cgctgatcag cctcgactgt
     2641 gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga
     2701 aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag
     2761 taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga
     2821 agacaatagc aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac
     2881 cagctggggc tctagggggt atccccacgc gccctgtagc ggcgcattaa gcgcggcggg
     2941 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt
     3001 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     3061 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     3121 ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
     3181 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc
     3241 tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa
     3301 aaatgagctg atttaacaaa aatttaacgc gaattaattc tgtggaatgt gtgtcagtta
     3361 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     3421 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     3481 atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta
     3541 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     3601 gaggccgagg ccgcctctgc ctctgagcta ttccagaagt agtgaggagg cttttttgga
     3661 ggcctaggct tttgcaaaaa gctcccggga gcttgtatat ccattttcgg atctgatcaa
     3721 gagacaggat gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc aggttctccg
     3781 gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat cggctgctct
     3841 gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac
     3901 ctgtccggtg ccctgaatga actgcaggac gaggcagcgc ggctatcgtg gctggccacg
     3961 acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg
     4021 ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa
     4081 gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca
     4141 ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt
     4201 gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc
     4261 aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc
     4321 ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg
     4381 ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt
     4441 ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag
     4501 cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg gggttcgcga
     4561 aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt
     4621 ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg
     4681 cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg
     4741 ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc
     4801 tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc
     4861 tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct
     4921 cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg
     4981 agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     5041 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     5101 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
     5161 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
     5221 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
     5281 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
     5341 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     5401 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     5461 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     5521 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     5581 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     5641 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     5701 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
     5761 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     5821 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     5881 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
     5941 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     6001 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
     6061 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     6121 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     6181 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     6241 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     6301 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
     6361 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     6421 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     6481 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     6541 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     6601 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     6661 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     6721 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     6781 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     6841 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     6901 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
     6961 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc
     7021 ccgaaaagtg ccacctgacg tc
//
