LOCUS       sequence-438872-        7294 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  SPNS1 codon-optimized orf with n-terminal 3xFlag in pMXs-IRES-Blast 
            vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7294)
  AUTHORS   Scharenberg SG, Dong W, Ghoochani A, Nyame K, Levin-Konigsberg R, 
            Krishnan AR, Rawat ES, Spees K, Bassik MC, Abu-Remaileh M
  TITLE     An SPNS1-dependent lysosomal lipid transport pathway that enables 
            cell survival under choline limitation.
  JOURNAL   Sci Adv. 2023 Apr 21;9(16):eadf8966. doi: 10.1126/sciadv.adf8966. 
            Epub 2023 Apr 19.
  PUBMED    37075117
REFERENCE   2  (bases 1 to 7294)
  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: "Sci Adv."; 
            date: "2023-04-21"; volume: "9(16):eadf8966. doi"; pages: " 
            10.1126/sciadv.adf8966. Epub 2023 Apr 19"
COMMENT     Sequence Label: pMXs-3xFlag-SPNS1
FEATURES             Location/Qualifiers
     source          1..7294
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     21..40
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(259..363)
                     /gene="bla"
                     /label=AmpR promoter
     LTR             393..986
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     misc_feature    1049..1406
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             1465..1881
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1811..1833
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1849..1865
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     misc_feature    1891..2265
                     /label=pol region
                     /note="Moloney murine leukemia virus (MMLV) pol region 
                     containing the splice acceptor site"
     primer_bind     2203..2219
                     /label=pBMN 5'
                     /note="MMLV sequence, for inserts in pBMN retroviral 
                     vector"
     regulatory      2294..2303
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2303..2368
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     misc_feature    join(4017..4591,4592..4594,4595..4603)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(4184..4201)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     4411..4430
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             4604..4999
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MVPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTGV
                     NVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGIK
                     AIVKDSDGQPTAVGIRELLPSGYVWEG"
     LTR             5170..5639
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(5769..5786)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5940..6528)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6020..6039)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6692..7294,1..189,190..258))
                     /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"
ORIGIN
        1 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
       61 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
      121 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
      181 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
      241 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
      301 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
      361 gcgcacattt ccccgaaaag tgccacctgc ataatgaaag accccacctg taggtttggc
      421 aagctagctt aagtaacgcc attttgcaag gcatggaaaa atacataact gagaatagaa
      481 aagttcagat caaggtcagg aacagatgga acagctgaat atgggccaaa caggatatct
      541 gtggtaagca gttcctgccc cggctcaggg ccaagaacag atggaacagc tgaatatggg
      601 ccaaacagga tatctgtggt aagcagttcc tgccccggct cagggccaag aacagatggt
      661 ccccagatgc ggtccagccc tcagcagttt ctagagaacc atcagatgtt tccagggtgc
      721 cccaaggacc tgaaatgacc ctgtgcctta tttgaactaa ccaatcagtt cgcttctcgc
      781 ttctgttcgc gcgcttctgc tccccgagct caataaaaga gcccacaacc cctcactcgg
      841 cgcgccagtc ctccgattga ctgagtcgcc cgggtacccg tgtatccaat aaaccctctt
      901 gcagttgcat ccgacttgtg gtctcgctgt tccttgggag ggtctcctct gagtgattga
      961 ctacccgtca gcgggggtct ttcatttggg ggctcgtccg ggatcgggag acccctgccc
     1021 agggaccacc gacccaccac cgggaggtaa gctggccagc aacttatctg tgtctgtccg
     1081 attgtctagt gtctatgact gattttatgc gcctgcgtcg gtactagtta gctaactagc
     1141 tctgtatctg gcggacccgt ggtggaactg acgagttcgg aacacccggc cgcaaccctg
     1201 ggagacgtcc cagggacttc gggggccgtt tttgtggccc gacctgagtc caaaaatccc
     1261 gatcgttttg gactctttgg tgcacccccc taataggagg gatatgtggt tctggtagga
     1321 gacgagaacc taaaacagtt cccgcctccg tctgaatttt tgctttcggt ttgggaccga
     1381 agccgcgccg cgcgtcttgt ctgctgcagc atcgttctgt gttgtctctg tctgactgtg
     1441 tttctgtatt tgtctgaaaa ttagggccag actgttacca ctcccttaag tttgacctta
     1501 ggtcactgga aagatgtcga gcggatcgct cacaaccagt cggtagatgt caagaagaga
     1561 cgttgggtta ccttctgctc tgcagaatgg ccaaccttta acgtcggatg gccgcgagac
     1621 ggcaccttta accgagacct catcacccag gttaagatca aggtcttttc acctggcccg
     1681 catggacacc cagaccaggt cccctacatc gtgacctggg aagccttggc ttttgacccc
     1741 cctccctggg tcaagccctt tgtacaccct aagcctccgc ctcctcttcc tccatccgcc
     1801 ccgtctctcc cccttgaacc tcctcgttcg accccgcctc gatcctccct ttatccagcc
     1861 ctcactcctt ctctaggcgc ccccatatgg ccatatgaga tcttatatgg ggcacccccg
     1921 ccccttgtaa acttccctga ccctgacatg acaagagtta ctaacagccc ctctctccaa
     1981 gctcacttac aggctctcta cttagtccag cacgaagtct ggagacctct ggcggcagcc
     2041 taccaagaac aactggaccg accggtggta cctcaccctt accgagtcgg cgacacagtg
     2101 tgggtccgcc gacaccagac taagaaccta gaacctcgct ggaaaggacc ttacacagtc
     2161 ctgctgacca cccccaccgc cctcaaagta gacggcatcg cagcttggat acacgccgcc
     2221 cacgtgaagg ctgccgaccc cgggggtgga ccatcctcta gactgccgga tctagctagt
     2281 taattaagga tccgccacca tggactacaa ggaccacgac ggtgactaca aggaccacga
     2341 catcgactac aaggacgacg acgacaaggg gcatggcacc ggcagcaccg gcagcgaatt
     2401 catggccggc tctgacaccg cccctttcct gagccaggcc gacgatccag acgatggacc
     2461 agtgccagga acccctggac tgccaggctc tacaggcaac cccaagagcg aggagccaga
     2521 ggtgcctgac caggagggac tgcagaggat cacaggactg tcccctggcc ggtctgccct
     2581 gatcgtggcc gtgctgtgct acatcaacct gctgaattat atggatcgct ttaccgtggc
     2641 aggcgtgctg ccagacatcg agcagttctt taacatcggc gatagctcct ctggcctgat
     2701 ccagacagtg ttcatcagct cctacatggt gctggccccc gtgttcggct atctgggcga
     2761 tcgctacaat cggaagtatc tgatgtgcgg cggcatcgcc ttctggagcc tggtgaccct
     2821 gggctctagc ttcatcccag gcgagcactt ttggctgctg ctgctgacaa ggggactggt
     2881 gggagtggga gaggcaagct actccaccat cgcacctaca ctgatcgccg acctgtttgt
     2941 ggccgatcag agaagcagga tgctgtccat cttctacttt gccatcccag tgggaagcgg
     3001 actgggatat atcgcaggat ccaaggtgaa ggacatggcc ggcgattggc actgggcact
     3061 gagagtgacc cctggactgg gagtggtggc cgtgctgctg ctgttcctgg tggtgagaga
     3121 gccacctagg ggagcagtgg agaggcactc tgacctgcca cccctgaacc caacaagctg
     3181 gtgggcagat ctgcgcgccc tggcaaggaa tccatccttc gtgctgtcct ctctgggctt
     3241 caccgccgtg gcctttgtga caggatccct ggccctgtgg gcacctgcct ttctgctgcg
     3301 ctctagggtg gtgctgggag agacccctcc atgcctgcca ggcgactcct gtagctcctc
     3361 tgattctctg atctttggcc tgatcacctg cctgacaggc gtgctgggag tgggactggg
     3421 agtggagatc agccggagac tgagacactc caacccaagg gcagacccac tggtgtgcgc
     3481 aaccggactg ctgggctccg cccccttcct gtttctgtct ctggcatgtg caaggggaag
     3541 catcgtggca acctacatct tcatctttat cggcgagaca ctgctgagca tgaattgggc
     3601 catcgtggcc gatatcctgc tgtatgtggt catcccaacc aggcgctcca cagccgaggc
     3661 cttccagatc gtgctgtctc acctgctggg cgacgcagga agcccatatc tgatcggcct
     3721 gatctccgat cgcctgcgga gaaattggcc cccttctttc ctgagcgagt ttcgggccct
     3781 gcagttcagc ctgatgctgt gcgcctttgt gggcgccctg ggaggagcag ccttcctggg
     3841 caccgccatc tttatcgagg ccgacaggcg ccgggcacag ctgcacgtgc agggactgct
     3901 gcacgaggca ggctccaccg acgatagaat cgtggtgccc cagagaggca ggtccacaag
     3961 ggtgcctgtg gcctctgtgc tgatctgagc ggccgctacg taaattccgc cccccccccc
     4021 tctccctccc ccccccctaa cgttactggc cgaagccgct tggaataagg ccggtgtgcg
     4081 tttgtctata tgttattttc caccatattg ccgtcttttg gcaatgtgag ggcccggaaa
     4141 cctggccctg tcttcttgac gagcattcct aggggtcttt cccctctcgc caaaggaatg
     4201 caaggtctgt tgaatgtcgt gaaggaagca gttcctctgg aagcttcttg aagacaaaca
     4261 acgtctgtag cgaccctttg caggcagcgg aaccccccac ctggcgacag gtgcctctgc
     4321 ggccaaaagc cacgtgtata agatacacct gcaaaggcgg cacaacccca gtgccacgtt
     4381 gtgagttgga tagttgtgga aagagtcaaa tggctctcct caagcgtatt caacaagggg
     4441 ctgaaggatg cccagaaggt accccattgt atgggatctg atctggggcc tcggtgcaca
     4501 tgctttacat gtgtttagtc gaggttaaaa aaacgtctag gccccccgaa ccacggggac
     4561 gtggttttcc tttgaaaaac acgatgataa tatggccaca accatggttc ctttgtctca
     4621 agaagaatcc accctcattg aaagagcaac ggctacaatc aacagcatcc ccatctctga
     4681 agactacagc gtcgccagcg cagctctctc tagcgacggc cgcatcttca ctggtgtcaa
     4741 tgtatatcat tttactgggg gaccttgtgc agaactcgtg gtgctgggca ctgctgctgc
     4801 tgcggcagct ggcaacctga cttgtatcgt cgcgatcgga aatgagaaca ggggcatctt
     4861 gagcccctgc ggacggtgcc gacaggtgct tctcgatctg catcctggga tcaaagccat
     4921 agtgaaggac agtgatggac agccgacggc agttgggatt cgtgaattgc tgccctctgg
     4981 ttatgtgtgg gagggctaag tcgacgataa aataaaagat tttatttagt ctccagaaaa
     5041 aggggggaat gaaagacccc acctgtaggt ttggcaagct agcttaagta acgccatttt
     5101 gcaaggcatg gaaaaataca taactgagaa tagagaagtt cagatcaagg tcaggaacag
     5161 atggaacagc tgaatatggg ccaaacagga tatctgtggt aagcagttcc tgccccggct
     5221 cagggccaag aacagatgga acagctgaat atgggccaaa caggatatct gtggtaagca
     5281 gttcctgccc cggctcaggg ccaagaacag atggtcccca gatgcggtcc agccctcagc
     5341 agtttctaga gaaccatcag atgtttccag ggtgccccaa ggacctgaaa tgaccctgtg
     5401 ccttatttga actaaccaat cagttcgctt ctcgcttctg ttcgcgcgct tctgctcccc
     5461 gagctcaata aaagagccca caacccctca ctcggggcgc cagtcctccg attgactgag
     5521 tcgcccgggt acccgtgtat ccaataaacc ctcttgcagt tgcatccgac ttgtggtctc
     5581 gctgttcctt gggagggtct cctctgagtg attgactacc cgtcagcggg ggtctttcac
     5641 atgcagcatg tatcaaaatt aatttggttt tttttcttaa gtatttacat taaatggcca
     5701 tagttgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct
     5761 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     5821 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     5881 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     5941 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     6001 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     6061 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     6121 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     6181 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     6241 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     6301 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     6361 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc
     6421 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     6481 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
     6541 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
     6601 atgagattat caaaaaggat cttcacctag atccttttgc ggccggccgc aaatcaatct
     6661 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     6721 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     6781 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
     6841 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     6901 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     6961 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
     7021 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     7081 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     7141 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     7201 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     7261 tctgagaata gtgtatgcgg cgaccgagtt gctc
//
