LOCUS       sequence-259995-        6899 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mammalian expression of ALK5 K232R.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6899)
  AUTHORS   Moren A, Imamura T, Miyazono K, Heldin CH, Moustakas A
  TITLE     Degradation of the tumor suppressor Smad4 by WW and HECT domain 
            ubiquitin ligases.
  JOURNAL   J Biol Chem. 2005 Jun 10;280(23):22115-23. Epub 2005 Apr 6.
  PUBMED    15817471
REFERENCE   2  (bases 1 to 6899)
  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: "J Biol 
            Chem."; date: "2005-06-10"; volume: "280(23)"; pages: "22115-23. 
            Epub 2005 Apr 6"
COMMENT     Sequence Label: pcDNA3-ALK5 K232R
FEATURES             Location/Qualifiers
     source          1..6899
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             2233..2259
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2239..2265
                     /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(2290..2308)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2291..2308)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2328..2345)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2334..2558
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2604..3032
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2691..2710)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2901..2922
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3041..3061)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3085..3295
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        3107..3303
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3154..3289
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3216..3235
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3370..4164
                     /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(3424..3443)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4034..4053
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4338..4459
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4375..4394)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4429..4448
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4508..4524)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4508..4524)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4521..4543)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4532..4548
                     /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(4556..4562,4563..4580,4581..4586))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4601..4622
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4739..4756)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4910..5498)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4990..5009)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5669..6460,6461..6529))
                     /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     6292..6311
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6530..6634)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6709..6728)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctt
      661 ggtaccgagc tcggatccac tagtaacggc cgccagtgtg ctggaattcg ggccatggag
      721 gcggcggtcg ctgctccgcg tccccggctg ctcctcctcg tgctggcggc ggcggcggcg
      781 gcggcggcgg cgctgctccc gggggcgacg gcgttacagt gtttctgcca cctctgtaca
      841 aaagacaatt ttacttgtgt gacagatggg ctctgctttg tctctgtcac agagaccaca
      901 gacaaagtta tacacaacag catgtgtata gctgaaattg acttaattcc acgagatagg
      961 ccgtttgtat gtgcaccctc ttcaaaaact gggtctgtga ctacaacata ttgctgcaat
     1021 caggaccatt gcaataaaat agaacttcca actactgtaa agtcatcacc tggccttggt
     1081 cctgtggaac tggcagctgt cattgctgga ccagtgtgct tcgtctgcat ctcactcatg
     1141 ttgatggtct atatctgcca caaccgcact gtcattcacc atcgagtgcc aaatgaagag
     1201 gacccttcat tagatcgccc ttttatttca gagggtacta cgttgaaaga cttaatttat
     1261 gatatgacaa cgtcaggttc tggctcaggt ttaccattgc ttgttcagag aacaattgcg
     1321 agaactattg tgttacaaga aagcattggc aaaggtcgat ttggagaagt ttggagagga
     1381 aagtggcggg gagaagaagt tgctgttagg atattctcct ctagagaaga acgttcgtgg
     1441 ttccgtgagg cagagattta tcaaactgta atgttacgtc atgaaaacat cctgggattt
     1501 atagcagcag acaataaaga caatggtact tggactcagc tctggttggt gtcagattat
     1561 catgagcatg gatccctttt tgattactta aacagataca cagttactgt ggaaggaatg
     1621 ataaaacttg ctctgtccac ggcgagcggt cttgcccatc ttcacatgga gattgttggt
     1681 acccaaggaa agccagccat tgctcataga gatttgaaat caaagaatat cttggtaaag
     1741 aagaatggaa cttgctgtat tgcagactta ggactggcag taagacatga ttcagccaca
     1801 gataccattg atattgctcc aaaccacaga gtgggaacaa aaaggtacat ggcccctgaa
     1861 gttctcgatg attccataaa tatgaaacat tttgaatcct tcaaacgtgc tgacatctat
     1921 gcaatgggct tagtattctg ggaaattgct cgacgatgtt ccattggtgg tattcatgaa
     1981 gattaccaac tgccttatta tgatcttgta ccttctgacc catcagttga agaaatgaga
     2041 aaagttgttt gtgaacagaa gttaaggcca aatatcccaa acagatggca gagctgtgaa
     2101 gccttgagag taatggctaa aattatgaga gaatgttggt atgccaatgg agcagctagg
     2161 cttacagcat tgcggattaa gaaaacatta tcgcaactca gtcaacagga aggcatcaaa
     2221 atgctcgagg gttatccgta tgatgttcct gattatgcta gcctctaggt cgactctaga
     2281 gggccctatt ctatagtgtc acctaaatgc tagagctcgc tgatcagcct cgactgtgcc
     2341 ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg
     2401 tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag
     2461 gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga
     2521 caatagcagg catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaaccag
     2581 ctggggctct agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
     2641 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
     2701 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
     2761 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta
     2821 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     2881 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
     2941 ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa
     3001 tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg
     3061 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     3121 tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc
     3181 gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc
     3241 tctgcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc
     3301 aaaaagctcc cgggagcttg tatatccatt ttcggatctg atcaagagac aggatgagga
     3361 tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag
     3421 aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc
     3481 cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg
     3541 aatgaactgc aggacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc
     3601 gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg
     3661 ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct
     3721 gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg
     3781 aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat
     3841 ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc
     3901 atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg
     3961 gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc
     4021 tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct
     4081 gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat
     4141 cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga
     4201 cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct
     4261 tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg
     4321 agttcttcgc ccaccccaac ttgtttattg cagcttataa tggttacaaa taaagcaata
     4381 gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca
     4441 aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag agcttggcgt
     4501 aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca
     4561 tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat
     4621 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt
     4681 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct
     4741 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
     4801 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
     4861 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     4921 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     4981 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     5041 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     5101 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     5161 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     5221 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     5281 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     5341 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     5401 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     5461 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     5521 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     5581 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     5641 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     5701 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     5761 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     5821 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     5881 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     5941 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     6001 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     6061 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     6121 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     6181 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     6241 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     6301 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     6361 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     6421 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     6481 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     6541 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     6601 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     6661 acgtcgacgg atcgggagat ctcccgatcc cctatggtgc actctcagta caatctgctc
     6721 tgatgccgca tagttaagcc agtatctgct ccctgcttgt gtgttggagg tcgctgagta
     6781 gtgcgcgagc aaaatttaag ctacaacaag gcaaggcttg accgacaatt gcatgaagaa
     6841 tctgcttagg gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat atacgcgtt
//
