LOCUS       sequence-454609-        6964 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of recombinant BMPS WT.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6964)
  AUTHORS   Medoh UN, Hims A, Chen JY, Ghoochani A, Nyame K, Dong W, 
            Abu-Remaileh M
  TITLE     The Batten disease gene product CLN5 is the lysosomal 
            bis(monoacylglycero)phosphate synthase.
  JOURNAL   Science. 2023 Sep 15;381(6663):1182-1189. doi: 
            10.1126/science.adg9288.
  PUBMED    37708259
REFERENCE   2  (bases 1 to 6964)
  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"; doi: "10.1126/science"; 
            journalName: "Science"; date: "2023-09-15- 15"; volume: "381"; 
            issue: "6663"; pages: "1182-1189"
COMMENT     Sequence Label: pRK5  BMPS WT-TEV-6xHis
FEATURES             Location/Qualifiers
     source          1..6964
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    6..140
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(43..62)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     97..116
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        209..566
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        209..400
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(232..252)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      417..552
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(586..603)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(586..602)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(595..617)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      815..1270
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(902..921)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1112..1133
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1401..2204)
                     /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(2205..2420)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(2444..2463)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2563..2585
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2623..2641)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2709..2813
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2814..2882,2883..3674)
                     /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(3032..3051)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3845..4433
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4334..4353
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4587..4604
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4721..4742
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4757..4762,4763..4780,4781..4787)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4795..4811
                     /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)."
     primer_bind     4800..4822
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4819..4835
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4819..4835
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     enhancer        4864..5243
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        5244..5447
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5398..5418
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     5444..5468
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        5678..5696
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     5678..5695
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     5709..5728
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             6882..6902
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             6903..6920
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     regulatory      join(6957..6964,1..2)
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 ggcccaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
       61 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
      121 tatcttatca tgtctggatc gggaattaat tcggcgcagc accatggcct gaaataacct
      181 ctgaaagagg aacttggtta ggtaccttct gaggcggaaa gaaccagctg tggaatgtgt
      241 gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
      301 atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta
      361 tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc
      421 cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta
      481 tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag tgaggaggct
      541 tttttggagg cctaggcttt tgcaaaaagc tgttaacagc ttggcactgg ccgtcgtttt
      601 acaacgtcgt gactgggaaa accctggcgt tacccaactt aatcgccttg cagcacatcc
      661 ccctttcgcc agctggcgta atagcgaaga ggcccgcacc gatcgccctt cccaacagtt
      721 gcgcagcctg aatggcgaat ggcgcctgat gcggtatttt ctccttacgc atctgtgcgg
      781 tatttcacac cgcatacgtc aaagcaacca tagtacgcgc cctgtagcgg cgcattaagc
      841 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc
      901 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
      961 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
     1021 aaacttgatt tgggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc
     1081 cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca
     1141 ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat
     1201 tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg
     1261 cttacaattt cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca
     1321 tagggtaata actgatataa ttaaattgaa gctctaattt gtgagtttag tatacatgca
     1381 tttacttata atacagtttt ttagttttgc tggccgcatc ttctcaaata tgcttcccag
     1441 cctgcttttc tgtaacgttc accctctacc ttagcatccc ttccctttgc aaatagtcct
     1501 cttccaacaa taataatgtc agatcctgta gagaccacat catccacggt tctatactgt
     1561 tgacccaatg cgtctccctt gtcatctaaa cccacaccgg gtgtcataat caaccaatcg
     1621 taaccttcat ctcttccacc catgtctctt tgagcaataa agccgataac aaaatctttg
     1681 tcgctcttcg caatgtcaac agtaccctta gtatattctc cagtagatag ggagcccttg
     1741 catgacaatt ctgctaacat caaaaggcct ctaggttcct ttgttacttc ttctgccgcc
     1801 tgcttcaaac cgctaacaat acctgggccc accacaccgt gtgcattcgt aatgtctgcc
     1861 cattctgcta ttctgtatac acccgcagag tactgcaatt tgactgtatt accaatgtca
     1921 gcaaattttc tgtcttcgaa gagtaaaaaa ttgtacttgg cggataatgc ctttagcggc
     1981 ttaactgtgc cctccatgga aaaatcagtc aagatatcca catgtgtttt tagtaaacaa
     2041 attttgggac ctaatgcttc aactaactcc agtaattcct tggtggtacg aacatccaat
     2101 gaagcacaca agtttgtttg cttttcgtgc atgatattaa atagcttggc agcaacagga
     2161 ctaggatgag tagcagcacg ttccttatat gtagctttcg acatgattta tcttcgtttc
     2221 ctgcaggttt ttgttctgtg cagttgggtt aagaatactg ggcaatttca tgtttcttca
     2281 acactacata tgcgtatata taccaatcta agtctgtgct ccttccttcg ttcttccttc
     2341 tgttcggaga ttaccgaatc aaaaaaattt caaggaaacc gaaatcaaaa aaaagaataa
     2401 aaaaaaaatg atgaattgaa aaggtggtat ggtgcactct cagtacaatc tgctctgatg
     2461 ccgcatagtt aagccagccc cgacacccgc caacacccgc tgacgcgccc tgacgggctt
     2521 gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc
     2581 agaggttttc accgtcatca ccgaaacgcg cgagacgaaa gggcctcgtg atacgcctat
     2641 ttttataggt taatgtcatg ataataatgg tttcttagac gtcaggtggc acttttcggg
     2701 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     2761 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
     2821 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
     2881 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
     2941 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
     3001 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg
     3061 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
     3121 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
     3181 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac
     3241 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
     3301 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag
     3361 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
     3421 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     3481 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     3541 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     3601 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     3661 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
     3721 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     3781 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     3841 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     3901 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     3961 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
     4021 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     4081 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     4141 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     4201 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     4261 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     4321 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     4381 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     4441 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     4501 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg
     4561 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc
     4621 caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca
     4681 ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt tagctcactc
     4741 attaggcacc ccaggcttta cactttatgc ttccggctcg tatgttgtgt ggaattgtga
     4801 gcggataaca atttcacaca ggaaacagct atgacatgat tacgaattaa ttcgagctcg
     4861 cccgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
     4921 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
     4981 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
     5041 ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac
     5101 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
     5161 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
     5221 tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat
     5281 agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt
     5341 tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc
     5401 aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctcgt ttagtgaacc
     5461 gtcagatcgc ctggagacgc catccacgct gttttgacct ccatagaaga caccgggacc
     5521 gatccagcct ccgcggccgg gaacggtgca ttggaacgcg gattccccgt gccaagagtg
     5581 acgtaagtac cgcctataga gtctataggc ccaccccctt ggcttcgtta gaacgcggct
     5641 acaattaata cataacctta tgtatcatac acatacgatt taggtgacac tatagaataa
     5701 catccacttt gcctttctct ccacaggtgt ccactcccag gtccaactgc acctcggttc
     5761 tatcgattga attcaccatg gctcaggagg ttgataccgc acaaggagcc gagatgcgac
     5821 gcggtgcagg agccgccaga ggcagggcgt cttggtgctg ggcccttgca ctgctttggc
     5881 ttgccgtggt gcctgggtgg tcacgggtta gcggaattcc ctcccggcgc cactggcctg
     5941 tgccatataa gcggttcgac tttcgaccta agccagaccc ttattgccaa gccaaataca
     6001 ccttctgccc caccggctct cccattccag tgatggaggg ggacgatgat atcgaagtgt
     6061 tcaggctcca ggctccagtc tgggaattca agtacggaga cttgctgggg catctcaaaa
     6121 ttatgcatga cgcgatcgga tttcggtcta ctctcacggg aaaaaattat accatggaat
     6181 ggtatgagct cttccaactc gggaattgca cttttcctca tttgaggcca gagatggatg
     6241 cacccttctg gtgtaaccaa ggcgctgcct gctttttcga gggaatagac gatgtgcatt
     6301 ggaaggagaa tgggacattg gtgcaggtgg ctaccattag cggcaatatg tttaatcaga
     6361 tggctaaatg ggtgaagcag gacaacgaaa cgggtattta ctatgagacc tggaacgtga
     6421 aagcatcccc tgagaagggc gctgagacat ggttcgactc ttatgactgc agcaagttcg
     6481 ttctcaggac tttcaataaa cttgcagagt ttggggccga gtttaagaac atcgagacaa
     6541 attacacccg catattcctg tactcaggcg agcccacata tctgggaaac gagacgtccg
     6601 tatttggccc aacaggcaac aagacgctgg gcttggccat caaacggttc tattatcctt
     6661 tcaagcccca tttacccaca aaggaattcc tgctgtctct tctccagatc tttgatgcag
     6721 tgatcgtgca caagcaattt tatttgtttt ataactttga gtactggttt ctgccaatga
     6781 agttcccatt cattaaaatt acatacgagg agatccctct tccgatccga aacaagaccc
     6841 ttagcggcct cggaggggga ggaagtcttc cagaaactgg agagaatctg tacttccagt
     6901 cccatcacca ccaccaccat taggcggccg ctaagtaagt aaggatcccc agcttggccg
     6961 ccat
//
