LOCUS       sequence-306233-        8591 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mammalian expression of HA tagged HIF1alpha (full length) with 
            P402A,P564A mutation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8591)
  AUTHORS   Kageyama Y, Koshiji M, To KK, Tian YM, Ratcliffe PJ, Huang LE
  TITLE     Leu-574 of human HIF-1alpha is a molecular determinant of prolyl 
            hydroxylation.
  JOURNAL   FASEB J. 2004 Jun;18(9):1028-30. Epub 2004 Apr 14.
  PUBMED    15084514
REFERENCE   2  (bases 1 to 8591)
  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: "FASEB J."; 
            date: "2004-06"; volume: "18(9)"; pages: "1028-30. Epub 2004 Apr 14"
COMMENT     Sequence Label: p(HA)HIF1alpha (P402A,P564A)
FEATURES             Location/Qualifiers
     source          1..8591
                     /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             937..963
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             943..969
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YDVPDYASL"
     promoter        complement(4144..4162)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(4145..4162)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(4182..4199)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4188..4412
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4458..4886
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4545..4564)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4755..4776
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4895..4915)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4900..5229
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5080..5215
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5142..5161
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5296..6090
                     /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(5350..5369)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5960..5979
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    6264..6385
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6301..6320)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6355..6374
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6434..6450)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6434..6450)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6447..6469)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6458..6474
                     /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(6482..6488,6489..6506,6507..6512))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6527..6548
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6665..6682)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6836..7424)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6916..6935)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7595..8386,8387..8455))
                     /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     8218..8237
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8456..8560)
                     /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 gcttggtacc
      901 gggccccccc tcgaggtcga ccgattcacc atggcgtacc catacgacgt gcctgactac
      961 gcctccctca tggagggcgc cggcggcgcg aacgacaaga aaaagataag ttctgaacgt
     1021 cgaaaagaaa agtctcgaga tgcagccaga tctcggcgaa gtaaagaatc tgaagttttt
     1081 tatgagcttg ctcatcagtt gccacttcca cataatgtga gttcgcatct tgataaggcc
     1141 tctgtgatga ggcttaccat cagctatttg cgtgtgagga aacttctgga tgctggtgat
     1201 ttggatattg aagatgacat gaaagcacag atgaattgct tttatttgaa agccttggat
     1261 ggttttgtta tggttctcac agatgatggt gacatgattt acatttctga taatgtgaac
     1321 aaatacatgg gattaactca gtttgaacta actggacaca gtgtgtttga ttttactcat
     1381 ccatgtgacc atgaggaaat gagagaaatg cttacacaca gaaatggcct tgtgaaaaag
     1441 ggtaaagaac aaaacacaca gcgaagcttt tttctcagaa tgaagtgtac cctaactagc
     1501 cgaggaagaa ctatgaacat aaagtctgca acatggaagg tattgcactg cacaggccac
     1561 attcacgtat atgataccaa cagtaaccaa cctcagtgtg ggtataagaa accacctatg
     1621 acctgcttgg tgctgatttg tgaacccatt cctcacccat caaatattga aattccttta
     1681 gatagcaaga ctttcctcag tcgacacagc ctggatatga aattttctta ttgtgatgaa
     1741 agaattaccg aattgatggg atatgagcca gaagaacttt taggccgctc aatttatgaa
     1801 tattatcatg ctttggactc tgatcatctg accaaaactc atcatgatat gtttactaaa
     1861 ggacaagtca ccacaggaca gtacaggatg cttgccaaaa gaggtggata tgtctgggtt
     1921 gaaactcaag caactgtcat atataacacc aagaattctc aaccacagtg cattgtatgt
     1981 gtgaattacg ttgtgagtgg tattattcag cacgacttga ttttctccct tcaacaaaca
     2041 gaatgtgtcc ttaaaccggt tgaatcttca gatatgaaaa tgactcagct attcaccaaa
     2101 gttgaatcag aagatacaag tagcctcttt gacaaactta agaaggaacc tgatgcttta
     2161 actttgctgg ccgcagccgc tggagacaca atcatatctt tagattttgg cagcaacgac
     2221 acagaaactg atgaccagca acttgaggaa gtaccattat ataatgatgt aatgctcccc
     2281 tcacccaacg aaaaattaca gaatataaat ttggcaatgt ctccattacc caccgctgaa
     2341 acgccaaagc cacttcgaag tagtgctgac cctgcactca atcaagaagt tgcattaaaa
     2401 ttagaaccaa atccagagtc actggaactt tcttttacca tgccccagat tcaggatcag
     2461 acacctagtc cttccgatgg aagcactaga caaagttcac ctgagcctaa tagtcccagt
     2521 gaatattgtt tttatgtgga tagtgatatg gtcaatgaat tcaagttgga attggtagaa
     2581 aaactttttg ctgaagacac agaagcaaag aacccatttt ctactcagga cacagattta
     2641 gacttggaga tgttagctgc ctatatccca atggatgatg acttccagtt acgttccttc
     2701 gatcagttgt caccattaga aagcagttcc gcaagccctg aaagcgcaag tcctcaaagc
     2761 acagttacag tattccagca gactcaaata caagaaccta ctgctaatgc caccactacc
     2821 actgccacca ctgatgaatt aaaaacagtg acaaaagacc gtatggaaga cattaaaata
     2881 ttgattgcat ctccatctcc tacccacata cataaagaaa ctactagtgc cacatcatca
     2941 ccatatagag atactcaaag tcggacagcc tcaccaaaca gagcaggaaa aggagtcata
     3001 gaacagacag aaaaatctca tccaagaagc cctaacgtgt tatctgtcgc tttgagtcaa
     3061 agaactacag ttcctgagga agaactaaat ccaaagatac tagctttgca gaatgctcag
     3121 agaaagcgaa aaatggaaca tgatggttca ctttttcaag cagtaggaat tggaacatta
     3181 ttacagcagc cagacgatca tgcagctact acatcacttt cttggaaacg tgtaaaagga
     3241 tgcaaatcta gtgaacagaa tggaatggag caaaagacaa ttattttaat accctctgat
     3301 ttagcatgta gactgctggg gcaatcaatg gatgaaagtg gattaccaca gctgaccagt
     3361 tatgattgtg aagttaatgc tcctatacaa ggcagcagaa acctactgca gggtgaagaa
     3421 ttactcagag ctttggatca agttaactga gctttttctt aatttcattc ctttttttgg
     3481 acactggtgg ctcactacct aaagcagtct atttatattt tctacatcta attttagaag
     3541 cctggctaca atactgcaca aacttggtta gttcaatttt tgatcccctt tctacttaat
     3601 ttacattaat gctctttttt agtatgttct ttaatgctgg atcacagaca gctcattttc
     3661 tcagtttttt ggtatttaaa ccattgcatt gcagtagcat cattttaaaa aatgcacctt
     3721 tttatttatt tatttttggc tagggagttt atcccttttt cgaattattt ttaagaagat
     3781 gccaatataa tttttgtaag aaggcagtaa cctttcatca tgatcatagg cagttgaaaa
     3841 atttttacac cttttttttc acattttaca taaataataa tgctttgcca gcagtacgtg
     3901 gtagccacaa ttgcacaata tattttctta aaaaatacca gcagttactc atggaatata
     3961 ttctgcgttt ataaaactag tttttaagaa gaaatttttt ttggcctatg aaattgttaa
     4021 acctggaaca tgacattgtt aatcatataa taatgattct taaatgctgt atggtttatt
     4081 atttaaatgg gtaaagccat ttacataata tagaaagata tgcatatatc tagagggccc
     4141 tattctatag tgtcacctaa atgctagagc tcgctgatca gcctcgactg tgccttctag
     4201 ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac
     4261 tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca
     4321 ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag
     4381 caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa ccagctgggg
     4441 ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt
     4501 tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt
     4561 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc
     4621 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga
     4681 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc
     4741 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt
     4801 ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct
     4861 gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga
     4921 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     4981 accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     5041 aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc
     5101 agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag
     5161 gccgcctctg cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc
     5221 ttttgcaaaa agctcccggg agcttgtata tccattttcg gatctgatca agagacagga
     5281 tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg
     5341 gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc
     5401 gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt
     5461 gccctgaatg aactgcagga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt
     5521 ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc
     5581 gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc
     5641 atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac
     5701 caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag
     5761 gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag
     5821 gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat
     5881 atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg
     5941 gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa
     6001 tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc
     6061 ttctatcgcc ttcttgacga gttcttctga gcgggactct ggggttcgaa atgaccgacc
     6121 aagcgacgcc caacctgcca tcacgagatt tcgattccac cgccgccttc tatgaaaggt
     6181 tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca
     6241 tgctggagtt cttcgcccac cccaacttgt ttattgcagc ttataatggt tacaaataaa
     6301 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
     6361 tgtccaaact catcaatgta tcttatcatg tctgtatacc gtcgacctct agctagagct
     6421 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     6481 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     6541 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     6601 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
     6661 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     6721 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     6781 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     6841 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     6901 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     6961 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     7021 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     7081 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     7141 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     7201 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     7261 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     7321 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     7381 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     7441 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     7501 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     7561 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     7621 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     7681 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     7741 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     7801 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     7861 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     7921 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     7981 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     8041 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     8101 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     8161 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     8221 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     8281 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     8341 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     8401 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     8461 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     8521 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     8581 cacctgacgt c
//
