LOCUS       sequence-176079-        6435 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mammalian expression of human Skp2 with N-terminal FLAG tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6435)
  AUTHORS   Su S, Chen X, Geng J, Minges JT, Grossman G, Wilson EM
  TITLE     Melanoma antigen-A11 regulates substrate-specificity of 
            Skp2-mediated protein degradation.
  JOURNAL   Mol Cell Endocrinol. 2017 Jan 5;439:1-9. doi: 
            10.1016/j.mce.2016.10.006. Epub 2016 Oct 6.
  PUBMED    27720894
REFERENCE   2  (bases 1 to 6435)
  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: "Mol Cell 
            Endocrinol."; date: "2017-01-5"; volume: "439:1-9. doi"; pages: " 
            10.1016/j.mce.2016.10.006. Epub 2016 Oct 6"
COMMENT     Sequence Label: FLAG-Skp2
FEATURES             Location/Qualifiers
     source          1..6435
                     /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"
     CDS             618..641
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     terminator      2037..2224
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(2092..2116)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     polyA_signal    2459..3081
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(2658..2677)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     primer_bind     complement(3105..3125)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3110..3439
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3290..3425
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(3477..3496)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3478..3496)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3510..3526)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3510..3526)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3523..3545)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3534..3550
                     /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(3558..3564,3565..3582,3583..3588))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3603..3624
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3741..3758)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3912..4500)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3992..4011)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4671..5462,5463..5531))
                     /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     5294..5313
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5532..5636)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      5663..6118
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5750..5769)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5960..5981
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6244..6266
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6258..6275
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6259..6275
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
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 ccgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttacggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta caccaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaataa ccccgccccg ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc
      601 agaattgatc taccatggac tacaaagacg atgacgacaa gcttgcggcc gacgaattcc
      661 cggggatccc catggacgta tttaaaactc ccgggcctgc ggacgctatg cacaggaagc
      721 acctccagga gattccagac ctgagtagca acgttgccac cagcttcacg tggggatggg
      781 attccagcaa gacttctgaa ctgctgtcag gcatgggggt ctccgccctg gagaaagagg
      841 agcccgacag tgagaacatc ccccaggaac tgctctcaaa cctgggccac ccggagagcc
      901 ccccacggaa acggctgaag agcaaaggga gtgacaaaga ctttgtgatt gtccgcaggc
      961 ctaagctaaa tcgagagaac tttccaggtg tttcatggga ctcccttccg gatgagctgc
     1021 tcttgggaat cttttcctgt ctgtgcctcc ctgagctgct aaaggtctct ggtgtttgta
     1081 agaggtggta tcgcctagcg tctgatgagt ctctatggca gaccttagac ctcacaggta
     1141 aaaatctgca cccggatgtg actggtcggt tgctgtctca aggggtgatt gccttccgct
     1201 gcccacgatc atttatggac caaccattgg ctgaacattt cagccctttt cgtgtacagc
     1261 acatggacct atcgaactca gttatagaag tgtccaccct ccacggcata ctgtctcagt
     1321 gttccaagtt gcagaatcta agcctggaag gcctgcggct ttcggatccc attgtcaata
     1381 ctctcgcaaa aaactcaaat ttagtgcgac ttaacctttc tgggtgttct ggattctctg
     1441 aatttgccct gcagactttg ctaagcagct gttccagact ggatgagctg aacctctcct
     1501 ggtgttttga tttcactgaa aagcatgtac aggtggctgt tgcgcatgtg tcagagacca
     1561 tcacccagct gaatcttagc ggctacagaa agaatctcca gaaatcagat ctctctactt
     1621 tagttagaag atgccccaat cttgtccatc tagacttaag tgatagtgtc atgctaaaga
     1681 atgactgctt tcaggaaatt ttccagctca actacctcca acacctatca ctcagtcggt
     1741 gctatgatat aatacctgaa actttacttg aacttggaga aattcccaca ctaaaaacac
     1801 tacaagtttt tggaatcgtg ccagatggta cccttcaact gttaaaggaa gcccttcctc
     1861 atctacagat taattgctcc catttcacca ccattgccag gccaactatt ggcaacaaaa
     1921 agaaccagga gatatggggc atcaaatgcc gactgacact gcaaaagccc agttgtctat
     1981 gaagtattta ttgcaggatg gtgtctcgac gagctctgca gccaagctaa ttccgggcga
     2041 atttcttatg atttatgatt tttattatta aataagttat aaaaaaaata agtgtataca
     2101 aattttaaag tgactcttag gttttaaaac gaaaattctt attcttgagt aactctttcc
     2161 tgtaggtcag gttgctttct caggtatagc atgaggtcgc tcttattgac cacacctcta
     2221 ccggcatgcc ggcaagtgca caaacaatac ttaaataaat actactcagt aataacctat
     2281 ttcttagcat ttttgacgaa atttgctatt ttgttagagt cttttacacc atttgtctcc
     2341 acacctccgc ttacatcaac accaataacg ccatttaatc taagcgcatc accaacattt
     2401 tctggcgtca gtccaccagc taacataaaa tgtaagcttc tcgactctag aggatcccgg
     2461 gtggcatccc tgtgacccct ccccagtgcc tctcctggcc ctggaagttg ccactccagt
     2521 gcccaccagc cttgtcctaa taaaattaag ttgcatcatt ttgtctgact aggtgtcctt
     2581 ctataatatt atggggtgga ggggggtggt atggagcaag gggcaagttg ggaagacaac
     2641 ctgtagggcc tgcggggtct attgggaacc aagctggagt gcagtggcac aatcttggct
     2701 cactgcaatc tccgcctcct gggttcaagc gattctcctg cctcagcctc ccgagttgtt
     2761 gggattccag gcatgcatga ccaggctcag ctaatttttg tttttttggt agagacgggg
     2821 tttcaccata ttggccaggc tggtctccaa ctcctaatct caggtgatct acccaccttg
     2881 gcctcccaaa ttgctgggat tacaggcgtg aaccactgct cccttccctg tccttctgat
     2941 tttaaaataa ctataccagc aggaggacgt ccagacacag cataggctac ctggccatgc
     3001 ccaaccggtg ggacatttga gttgcttgct tggcactgtc ctctcatgcg ttgggtccac
     3061 tcagtagatg cctgttgaat tgggtacgcg gccagcttgg ctgtggaatg tgtgtcagtt
     3121 agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     3181 ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     3241 catgcatctc aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct
     3301 aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc
     3361 agaggccgag gccgcctcgg cctctgagct attccagaag tagtgaggag gcttttttgg
     3421 aggcctaggc ttttgcaaaa agctcctcga ggaactgaaa aaccagaaag ttaattccct
     3481 atagtgagtc gtattaaatt cgtaatcatg tcatagctgt ttcctgtgtg aaattgttat
     3541 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
     3601 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     3661 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     3721 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     3781 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     3841 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     3901 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     3961 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     4021 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     4081 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     4141 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     4201 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     4261 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     4321 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     4381 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     4441 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     4501 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     4561 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     4621 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     4681 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     4741 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     4801 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     4861 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     4921 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     4981 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     5041 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     5101 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     5161 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     5221 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     5281 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
     5341 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
     5401 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
     5461 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     5521 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     5581 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     5641 tttccccgaa aagtgccacc tgacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg
     5701 gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct
     5761 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
     5821 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag
     5881 ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg
     5941 gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc
     6001 tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat
     6061 gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgct tacaatttcc
     6121 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
     6181 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
     6241 tttcccagtc acgacgttgt aaaacgacgg ccagtgccaa gctgatctat acattgaatc
     6301 aatattggca attagccata ttagtcattg gttatatagc ataaatcaat attggctatt
     6361 ggccattgca tacgttgtat ctatatcata atatgtacat ttatattggc tcatgtccaa
     6421 tatgaccgcc atgtt
//
