LOCUS       sequence-433109-        6595 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Expresses human SKP1 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6595)
  AUTHORS   Li J, Krause GJ, Gui Q, Kaushik S, Rona G, Zhang Q, Liang FX, 
            Dhabaria A, Anerillas C, Martindale JL, Vasilyev N, Askenazi M, 
            Ueberheide B, Nudler E, Gorospe M, Cuervo AM, Pagano M
  TITLE     A noncanonical function of SKP1 regulates the switch between 
            autophagy and unconventional secretion.
  JOURNAL   Sci Adv. 2023 Oct 13;9(41):eadh1134. doi: 10.1126/sciadv.adh1134. 
            Epub 2023 Oct 13.
  PUBMED    37831778
REFERENCE   2  (bases 1 to 6595)
  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: "Sci Adv."; 
            date: "2023-10-13"; volume: "9(41):eadh1134. doi"; pages: " 
            10.1126/sciadv.adh1134. Epub 2023 Oct 13"
COMMENT     Sequence Label: Flag-SKP1-WT
FEATURES             Location/Qualifiers
     source          1..6595
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(516..1115)
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(599..619)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     1096..1115
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     promoter        complement(1125..1454)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      1139..1274
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1439..1459
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     CDS             complement(join(2269..2979,2980..2982,2983..2985))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(2269..2979,2980..2982,2983..2985))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2311..2332)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     2660..2679
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2919..2940
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      2982..2991
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             complement(3026..3442)
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     complement(3042..3058)
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     primer_bind     complement(3074..3096)
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     misc_feature    3643..3842
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     LTR             3907..4376
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     promoter        4407..4511
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4512..4580,4581..5372)
                     /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     complement(4730..4749)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5532..6120
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6021..6040
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6274..6291
                     /label=L4440
                     /note="L4440 vector, forward primer"
     LTR             join(6421..6595,1..295)
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
ORIGIN
        1 gctcggggag cagaagcgcg cgaacagaag cgagaagcga actgattggt tagttcaaat
       61 aaggcacagg gtcatttcag gtccttgggg caccctggaa acatctgatg gttctctaga
      121 aactgctgag ggctggaccg catctgggga ccatctgttc ttggccctga gccggggcag
      181 gaactgctta ccacagatat cctgtttggc ccatattcag ctgttccatc tgttcttggc
      241 cctgagccgg ggcaggaact gcttaccaca gatatcctgt ttggcccata ttcagctgtt
      301 ccatctgttc ctgaccttga tctgaacttc tctattctca gttatgtatt tttccatgcc
      361 ttgcaaaatg gcgttactta agctagcttg ccaaacctac aggtggggtc tttcattccc
      421 ccctttttct ggagactaaa taaaatcttt tattttatcg atgcatgggg tcgtgcgctc
      481 ctttcggtcg ggcgctgcgg gtcgtggggc gggcgtcagg caccgggctt gcgggtcatg
      541 caccaggtgc gcggtccttc gggcacctcg acgtcggcgg tgacggtgaa gccgagccgc
      601 tcgtagaagg ggaggttgcg gggcgcggag gtctccagga aggcgggcac cccggcgcgc
      661 tcggccgcct ccactccggg gagcacgacg gcgctgccca gacccttgcc ctggtggtcg
      721 ggcgagacgc cgacggtggc caggaaccac gcgggctcct tgggccggtg cggcgccagg
      781 aggccttcca tctgttgctg cgcggccagc cgggaaccgc tcaactcggc catgcgcggg
      841 ccgatctcgg cgaacaccgc ccccgcttcg acgctctccg gcgtggtcca gaccgccacc
      901 gcggcgccgt cgtccgcgac ccacaccttg ccgatgtcga gcccgacgcg cgtgaggaag
      961 agttcttgca gctcggtgac ccgctcgatg tggcggtccg gatcgacggt gtggcgcgtg
     1021 gcggggtagt cggcgaacgc ggcggcgagg gtgcgtacgg ccctggggac gtcgtcgcgg
     1081 gtggcgaggc gcaccgtggg cttgtactcg gtcatggtaa gctttttgca aaagcctagg
     1141 cctccaaaaa agcctcctca ctacttctgg aatagctcag aggccgaggc ggcctcggcc
     1201 tctgcataaa taaaaaaaat tagtcagcca tggggcggag aatgggcgga actgggcgga
     1261 gttaggggcg ggatgggcgg agttaggggc gggactatgg ttgctgacta attgagatgc
     1321 atgctttgca tacttctgcc tgctggggag cctggggact ttccacacct ggttgctgac
     1381 taattgagat gcatgctttg catacttctg cctgctgggg agcctgggga ctttccacac
     1441 cctaactgac acacattcca cagggtcgag ctaagccgcc gtttgttcct gataaacttc
     1501 aatcacatct tcttcctcca ttcccagttc ttttggagta tgattatcag caattctctg
     1561 accctcaaag agaaacctga gtgaattcat tggaacaccc tgtctttgac agtatgattc
     1621 tttgagtttc ttgagatgtg ttgtcatttt cactttgaag tgaatctcac tgctatcctg
     1681 tccaatgact ttgagtttaa tatattcacc ttccttctta tcccccaagt cctcagttga
     1741 aggttttgcc tcctggtcag acatcttctc ttcacaccac tggttctctt tgcgtacctg
     1801 ggcttcctcc tcttcagtaa agtcattttt gatattgaag gtcttgcgaa tctcctcagg
     1861 agttttcccc ttgatcatat tggcaacagt cttgcatgta acatcaagca aacctttgat
     1921 gtctaagtag tttgcagcca gaatgagttc aaaaagtgtt ccttggtcaa ctttcaggaa
     1981 ttcttggtcc caaacaggga tatcatctgt tcgcttttct ttgttctcat catcttcagg
     2041 aggaggaggg tcatccttgt ggtgggtgca ccactgaatg acctttttta atattgctgc
     2101 attcacattt ggtagaggaa ctgggtcatc atctccttca tcatccattc ccaaatcttc
     2161 caacatggtc ttaatagtca cagattgttt ggcaatttcc acatcaactt caaatatctc
     2221 tccatcagaa ctctgcaact taattgaagg catagatctg agtccggact tgtacagctc
     2281 gtccatgccg agagtgatcc cggcggcggt cacgaactcc agcaggacca tgtgatcgcg
     2341 cttctcgttg gggtctttgc tcagggcgga ctgggtgctc aggtagtggt tgtcgggcag
     2401 cagcacgggg ccgtcgccga tgggggtgtt ctgctggtag tggtcggcga gctgcacgct
     2461 gccgtcctcg atgttgtggc ggatcttgaa gttcaccttg atgccgttct tctgcttgtc
     2521 ggccatgata tagacgttgt ggctgttgta gttgtactcc agcttgtgcc ccaggatgtt
     2581 gccgtcctcc ttgaagtcga tgcccttcag ctcgatgcgg ttcaccaggg tgtcgccctc
     2641 gaacttcacc tcggcgcggg tcttgtagtt gccgtcgtcc ttgaagaaga tggtgcgctc
     2701 ctggacgtag ccttcgggca tggcggactt gaagaagtcg tgctgcttca tgtggtcggg
     2761 gtagcggctg aagcactgca cgccgtaggt cagggtggtc acgagggtgg gccagggcac
     2821 gggcagcttg ccggtggtgc agatgaactt cagggtcagc ttgccgtagg tggcatcgcc
     2881 ctcgccctcg ccggacacgc tgaacttgtg gccgtttacg tcgccgtcca gctcgaccag
     2941 gatgggcacc accccggtga acagctcctc gcccttgctc accatggtgg cgaccggtag
     3001 cgtaccacca cactgggatc cggccggcgc ctagagaagg agtgagggct ggataaaggg
     3061 aggatcgagg cggggtcgaa cgaggaggtt caagggggag agacggggcg gatggaggaa
     3121 gaggaggcgg aggcttaggg tgtacaaagg gcttgaccca gggagggggg tcaaaagcca
     3181 aggcttccca ggtcacgatg taggggacct ggtctgggtg tccatgcggg ccaggtgaaa
     3241 agaccttgat cttaacctgg gtgatgaggt ctcggttaaa ggtgccgtct cgcggccatc
     3301 cgacgttaaa ggttggccat tctgcagagc agaaggtaac ccaacgtctc ttcttgacat
     3361 ctaccgactg gttgtgagcg atccgctcga catctttcca gtgacctaag gtcaaactta
     3421 agggagtggt aacagtctgg ccctaatttt cagacaaata cagaaacaca gtcagacaga
     3481 gacaacacag aacgatgctg cagcgctgca gcagacaaga cgcgcggctt cggttccaaa
     3541 ccgaaagcaa aaattcagac ggaggcggga actgttttag gttctcgtct cctaccagaa
     3601 ccacatatcc tgacggggtc ggattccaca tcgactccct tcctcaggtc gggccacaaa
     3661 aacggccccc aaagtccctg ggacgtctcc cagggttgcg gccgggtgtt cagaactcgt
     3721 cagttccacc acgggtccgc cagatacaga gctagttagc taactagtac cgacgcaggc
     3781 gcataaaatc agtcatagac actagacaat cggacagaca cagataagtt gctggccagc
     3841 ttgcctcccg gtggtgggtc ggtggtccct gggcaggggt ctcccgatcc cggacgagcc
     3901 cccaaatgaa agacccccgc tgacgggtag tcaatcactc agaggagacc ctcccaagga
     3961 acagcgagac cacaagtcgg atgcaactgc aagagggttt attggataca cgggtacccg
     4021 ggcgactcag tcaatcggag gactggcgcc ccgagtgagg ggttgtgggc tcttttattg
     4081 agctcgggga gcagaagcgc gcgaacagaa gcgagaagcg aactgattgg ttagttcaaa
     4141 taaggcacag ggtcatttca ggtccttggg gcaccctgga aacatctgat ggttctctag
     4201 aaactgctga gggctggacc gcatctgggg accatctgtt cttggccctg agccggggca
     4261 ggaactgctt accacagata tcctgtttgg cccatattca gctgttccat ctgttcttgg
     4321 ccctgagccg gggcaggaac tgcttaccac agatatcctg tttggcccat attcaggctg
     4381 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     4441 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     4501 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
     4561 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
     4621 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
     4681 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
     4741 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc
     4801 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
     4861 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
     4921 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
     4981 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
     5041 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
     5101 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
     5161 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
     5221 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
     5281 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
     5341 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     5401 tttagattga tttgcggccg caaaaggatc taggtgaaga tcctttttga taatctcatg
     5461 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     5521 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     5581 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     5641 gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta
     5701 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     5761 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     5821 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     5881 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     5941 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     6001 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     6061 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     6121 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     6181 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
     6241 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
     6301 gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcaactat
     6361 ggccatttaa tgtaaatact taagaaaaaa aaccaaatta attttgatac atgctgcatg
     6421 tgaaagaccc ccgctgacgg gtagtcaatc actcagagga gaccctccca aggaacagcg
     6481 agaccacaag tcggatgcaa ctgcaagagg gtttattgga tacacgggta cccgggcgac
     6541 tcagtcaatc ggaggactgg cgccccgagt gaggggttgt gggctctttt attga
//
