LOCUS       sequence-234126-        6987 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses sphingosine kinase 1, which catalyzes the phosphorylation 
            of sphingosine to form sphingosine 1-phosphate in mice.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6987)
  AUTHORS   Kharel Y, Agah S, Huang T, Mendelson AJ, Eletu OT, Barkey-Bircann P,
            Gesualdi J, Smith JS, Santos WL, Lynch KR
  TITLE     Saccharomyces cerevisiae as a platform for assessing sphingolipid 
            lipid kinase inhibitors.
  JOURNAL   PLoS One. 2018 Apr 19;13(4):e0192179. doi: 
            10.1371/journal.pone.0192179. eCollection 2018.
  PUBMED    29672528
REFERENCE   2  (bases 1 to 6987)
  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: "PLoS One.";
            date: "2018-04-19"; volume: "13(4):e0192179. doi"; pages: " 
            10.1371/journal.pone.0192179. eCollection 2018"
COMMENT     Sequence Label: MmSphk1
FEATURES             Location/Qualifiers
     source          1..6987
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        37..478
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    37..154
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     449..472
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     primer_bind     510..529
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        510..528
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             548..571
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     terminator      1771..2018
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(1787..1805)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(2095..2112)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2266..2854)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2346..2365)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3025..3816,3817..3885))
                     /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     3648..3667
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     CDS             complement(3981..4784)
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     promoter        complement(4785..5000)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     rep_origin      5601..6478
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     rep_origin      complement(join(6489..6987,1..15))
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6684..6705)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6896..6915
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 cgctacaggg cgcgtgggga tcgatccact agtccacgga ttagaagccg ccgagcgggt
       61 gacagccctc cgaaggaaga ctctcctccg tgcgtcctcg tcttcaccgg tcgcgttcct
      121 gaaacgcaga tgtgcctcgc gccgcactgc tccgaacaat aaagattcta caatactagc
      181 ttttatggtt atgaagagga aaaattggca gtaacctggc cccacaaacc ttcaaatgaa
      241 cgaatcaaat taacaaccat aggatgataa tgcgattagt tttttagcct tatttctggg
      301 gtaattaatc agcgaagcga tgatttttga tctattaaca gatatataaa tgcaaaaact
      361 gcataaccac tttaactaat actttcaaca ttttcggttt gtattacttc ttattcaaat
      421 gtaataaaag tatcaacaaa aaattgttaa tatacctcta tactttaacg tcaaggagaa
      481 aaaaccccgg atcggactac tagcagctgt aatacgactc actataggga atattaagct
      541 taccatggac tacaaggacg atgacgataa gggatccatg tggtggtgct gtgtcttgtt
      601 cgttgtcgaa tgcccaaggg gtttgttacc aagaccatgc agagtcttag tcttgttgaa
      661 ccctcagggt ggtaagggta aggctttgca gttgttccag tcaagggtcc aaccattctt
      721 ggaagaggca gaaattactt ttaaattaat tttgactgaa agaaagaacc acgctaggga
      781 attggtctgc gcagaggaat tgggtcactg ggatgcttta gctgtcatgt caggtgacgg
      841 tttgatgcac gaagttgtta atggtttgat ggagaggcca gattgggaga cagctattca
      901 aaagcctttg tgttctttgc caggtggatc tggtaacgct ttggctgctt cagtcaacca
      961 ctacgctggt tatgaacaag tcactaatga agatttgttg attaattgta cattgttatt
     1021 gtgcagaaga agattgtctc caatgaattt attgtctttg catacagctt ctggtttgag
     1081 attgtactct gttttgtcat tgtcttgggg tttcgttgct gacgttgact tggagtctga
     1141 gaaatataga agattgggag aaattagatt cactgttggt acttttttca gattggcttc
     1201 tttgagaatt taccaaggac agttggcata cttgccagtc ggtactgtcg cttctaagag
     1261 acctgcttct actttggtcc agaagggtcc agtcgatact catttggtcc cattggagga
     1321 acctgtccca tctcattgga ctgttgtccc agaacaagat tttgttttag ttttggtttt
     1381 gttacataca catttgtctt ctgaattgtt cgcagctcca atgggtaggt gtgaggctgg
     1441 tgttatgcac ttgttttatg ttagagctgg agtctctagg gctgctttgt tgagattgtt
     1501 cttagctatg caaaaaggta aacatatgga gttggactgc ccatacttgg tccatgtccc
     1561 tgtcgttgct ttcaggttgg aaccaaggtc acaaagggga gtcttctctg ttgacggtga
     1621 gttgatggtt tgcgaagcag tccaaggtca ggttcatcca aactacttgt ggatggtttg
     1681 cggttctaga gacgcaccat caggtaggga ttctagaaga ggtcctccac ctgaagaacc
     1741 ataactcgag catgcatcta gagggccgca tcatgtaatt agttatgtca cgcttacatt
     1801 cacgccctcc ccccacatcc gctctaaccg aaaaggaagg agttagacaa cctgaagtct
     1861 aggtccctat ttattttttt atagttatgt tagtattaag aacgttattt atatttcaaa
     1921 tttttctttt ttttctgtac agacgcgtgt acgcatgtaa cattatactg aaaaccttgc
     1981 ttgagaaggt tttgggacgc tcgaaggctt taatttgcaa gctgcggccc tgcattaatg
     2041 aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct
     2101 cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc
     2161 ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg
     2221 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     2281 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     2341 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     2401 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     2461 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     2521 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     2581 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     2641 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     2701 tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     2761 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
     2821 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg
     2881 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
     2941 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
     3001 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
     3061 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
     3121 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc
     3181 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
     3241 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
     3301 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg
     3361 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
     3421 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
     3481 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
     3541 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
     3601 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc
     3661 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
     3721 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
     3781 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
     3841 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
     3901 atgggtaata actgatataa ttaaattgaa gctctaattt gtgagtttag tatacatgca
     3961 tttacttata atacagtttt ttagttttgc tggccgcatc ttctcaaata tgcttcccag
     4021 cctgcttttc tgtaacgttc accctctacc ttagcatccc ttccctttgc aaatagtcct
     4081 cttccaacaa taataatgtc agatcctgta gagaccacat catccacggt tctatactgt
     4141 tgacccaatg cgtctccctt gtcatctaaa cccacaccgg gtgtcataat caaccaatcg
     4201 taaccttcat ctcttccacc catgtctctt tgagcaataa agccgataac aaaatctttg
     4261 tcgctcttcg caatgtcaac agtaccctta gtatattctc cagtagatag ggagcccttg
     4321 catgacaatt ctgctaacat caaaaggcct ctaggttcct ttgttacttc ttctgccgcc
     4381 tgcttcaaac cgctaacaat acctgggccc accacaccgt gtgcattcgt aatgtctgcc
     4441 cattctgcta ttctgtatac acccgcagag tactgcaatt tgactgtatt accaatgtca
     4501 gcaaattttc tgtcttcgaa gagtaaaaaa ttgtacttgg cggataatgc ctttagcggc
     4561 ttaactgtgc cctccatgga aaaatcagtc aagatatcca catgtgtttt tagtaaacaa
     4621 attttgggac ctaatgcttc aactaactcc agtaattcct tggtggtacg aacatccaat
     4681 gaagcacaca agtttgtttg cttttcgtgc atgatattaa atagcttggc agcaacagga
     4741 ctaggatgag tagcagcacg ttccttatat gtagctttcg acatgattta tcttcgtttc
     4801 ctgcaggttt ttgttctgtg cagttgggtt aagaatactg ggcaatttca tgtttcttca
     4861 acactacata tgcgtatata taccaatcta agtctgtgct ccttccttcg ttcttccttc
     4921 tgttcggaga ttaccgaatc aaaaaaattt caaggaaacc gaaatcaaaa aaaagaataa
     4981 aaaaaaaatg atgaattgaa aagctagctt atcgatgata agctgtcaaa catgagaatt
     5041 aattccacgg actatagact atacctagta tactccgtct actgtacgat acacttccgc
     5101 tcaggtcctt gtcctttaac gaggccttac cactcttttg ttactctatt gatccagctc
     5161 agcaaaggca gtgtgatcta agattctatc ttcgcgatgt agtaaaacta gctagaccga
     5221 gaaagagact agaaatgcaa aaggcacttc tacaatggct gccatcatta ttatccgatg
     5281 tgacgctgca gcttctcaat gatattcgaa tacgctttga ggagatacag cctaatatcc
     5341 gacaaactgt tttacagatt tacgatcgta cttgttaccc atcattgaat tttgaacatc
     5401 cgaacctggg agttttccct gaaacagata gtatatttga acctgtataa taatatatag
     5461 tctagcgctt tacggaagac aatgtatgta tttcggttcc tggagaaact attgcatcta
     5521 ttgcataggt aatcttgcac gtcgcatccc cggttcattt tctgcgtttc catcttgcac
     5581 ttcaatagca tatctttgtt aacgaagcat ctgtgcttca ttttgtagaa caaaaatgca
     5641 acgcgagagc gctaattttt caaacaaaga atctgagctg catttttaca gaacagaaat
     5701 gcaacgcgaa agcgctattt taccaacgaa gaatctgtgc ttcatttttg taaaacaaaa
     5761 atgcaacgcg agagcgctaa tttttcaaac aaagaatctg agctgcattt ttacagaaca
     5821 gaaatgcaac gcgagagcgc tattttacca acaaagaatc tatacttctt ttttgttcta
     5881 caaaaatgca tcccgagagc gctatttttc taacaaagca tcttagatta ctttttttct
     5941 cctttgtgcg ctctataatg cagtctcttg ataacttttt gcactgtagg tccgttaagg
     6001 ttagaagaag gctactttgg tgtctatttt ctcttccata aaaaaagcct gactccactt
     6061 cccgcgttta ctgattacta gcgaagctgc gggtgcattt tttcaagata aaggcatccc
     6121 cgattatatt ctataccgat gtggattgcg catactttgt gaacagaaag tgatagcgtt
     6181 gatgattctt cattggtcag aaaattatga acggtttctt ctattttgtc tctatatact
     6241 acgtatagga aatgtttaca ttttcgtatt gttttcgatt cactctatga atagttctta
     6301 ctacaatttt tttgtctaaa gagtaatact agagataaac ataaaaaatg tagaggtcga
     6361 gtttagatgc aagttcaagg agcgaaaggt ggatgggtag gttatatagg gatatagcac
     6421 agagatatat agcaaagaga tacttttgag caatgtttgt ggaagcggta ttcgcaatgg
     6481 gaagctccac cccggttgat aatcagaaaa gccccaaaaa caggaagatt gtataagcaa
     6541 atatttaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca
     6601 gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga
     6661 ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg
     6721 actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat
     6781 caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag
     6841 ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga
     6901 agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa
     6961 ccaccacacc cgccgcgctt aatgcgc
//
