LOCUS       sequence-446771-        5771 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Mammalian expression of recombinant BMPS N143S.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5771)
  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 5771)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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 N143S-6xHis
FEATURES             Location/Qualifiers
     source          1..5771
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1134..1151
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     regulatory      1188..1197
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    1201..1335
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1238..1257)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1292..1311
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        1404..1761
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        1404..1595
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(1427..1447)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      1612..1747
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(1781..1798)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1781..1797)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1790..1812)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2010..2465
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2097..2116)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2307..2328
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2482..2501)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2601..2623
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2661..2679)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2747..2851
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2852..2920,2921..3712)
                     /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(3070..3089)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3883..4471
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4372..4391
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4625..4642
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4759..4780
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4795..4800,4801..4818,4819..4825)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4833..4849
                     /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     4838..4860
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4857..4873
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4857..4873
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     enhancer        4902..5281
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        5282..5485
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5436..5456
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     5482..5506
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        5716..5734
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     5716..5733
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     5747..5766
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
ORIGIN
        1 ctcccaggtc caactgcacc tcggttctat cgattgaatt caccatggcg caggaggtag
       61 acacggcaca gggcgccgag atgcggcggg gcgcgggcgc ggctcgggga cgcgcttcct
      121 ggtgctgggc cctggcgctg ctttggctcg cggtggttcc gggctggtcc cgggtctcgg
      181 gcatcccctc ccggcgccac tggccggtgc cctacaagcg ctttgacttc cgtccaaaac
      241 ctgatcctta ttgtcaagct aagtatactt tctgtccaac tggctcacct atcccagtta
      301 tggagggtga tgatgacatt gaagtttttc gattacaagc cccagtatgg gaatttaaat
      361 atggagacct cctgggacac ttgaaaatta tgcatgatgc cattggattc agaagtacat
      421 taactggcaa gaactacaca atggaatggt atgaactttt ccaacttggc agctgtacat
      481 ttccccatct ccgacctgaa atggatgccc ctttctggtg taatcaaggc gctgcctgct
      541 tttttgaggg aattgatgat gttcactgga aggaaaatgg gacattagtt caagtagcaa
      601 ctatatcagg aaacatgttc aaccaaatgg caaagtgggt gaaacaggac aatgaaacag
      661 gaatttatta tgagacatgg aatgtaaaag ccagcccaga aaagggggca gagacatggt
      721 ttgattccta cgactgttcc aaatttgtgt taaggacctt taacaagttg gctgaatttg
      781 gagcagagtt caagaacata gaaaccaact atacaagaat atttctttac agtggagaac
      841 ctacttatct gggaaatgaa acatctgttt ttgggccaac aggaaacaag actcttggtt
      901 tagccataaa aagattttat taccccttca aaccacattt gccaactaaa gaatttctgt
      961 tgagtctctt gcaaattttt gatgcagtga ttgtgcacaa acagttctat ttgttttata
     1021 attttgaata ttggttttta cctatgaaat tcccttttat taaaataaca tatgaagaaa
     1081 tccctttacc tatcagaaac aaaacactct ctggtttagg aggtggaggt agtcatcatc
     1141 accatcacca ctaggcggcc gctaagtaag taaggatccc cagcttggcc gccatggccc
     1201 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
     1261 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
     1321 tatcatgtct ggatcgggaa ttaattcggc gcagcaccat ggcctgaaat aacctctgaa
     1381 agaggaactt ggttaggtac cttctgaggc ggaaagaacc agctgtggaa tgtgtgtcag
     1441 ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     1501 aattagtcag caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     1561 agcatgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc
     1621 ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat
     1681 gcagaggccg aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt
     1741 ggaggcctag gcttttgcaa aaagctgtta acagcttggc actggccgtc gttttacaac
     1801 gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg ccttgcagca catccccctt
     1861 tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa cagttgcgca
     1921 gcctgaatgg cgaatggcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt
     1981 cacaccgcat acgtcaaagc aaccatagta cgcgccctgt agcggcgcat taagcgcggc
     2041 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc
     2101 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
     2161 tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact
     2221 tgatttgggt gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt
     2281 gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa
     2341 ccctatctcg ggctattctt ttgatttata agggattttg ccgatttcgg cctattggtt
     2401 aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgtttac
     2461 aattttatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagccccg
     2521 acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta
     2581 cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc
     2641 gaaacgcgcg agacgaaagg gcctcgtgat acgcctattt ttataggtta atgtcatgat
     2701 aataatggtt tcttagacgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat
     2761 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     2821 aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct
     2881 tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa
     2941 agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa
     3001 cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt
     3061 taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag agcaactcgg
     3121 tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca
     3181 tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa
     3241 cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt
     3301 gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc
     3361 cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa cgttgcgcaa
     3421 actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga
     3481 ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc
     3541 tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga
     3601 tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga
     3661 acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga
     3721 ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat
     3781 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt
     3841 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     3901 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     3961 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     4021 aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     4081 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     4141 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     4201 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     4261 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     4321 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     4381 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     4441 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     4501 cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt
     4561 ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga
     4621 gcgcagcgag tcagtgagcg aggaagcgga agagcgccca atacgcaaac cgcctctccc
     4681 cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg tttcccgact ggaaagcggg
     4741 cagtgagcgc aacgcaatta atgtgagtta gctcactcat taggcacccc aggctttaca
     4801 ctttatgctt ccggctcgta tgttgtgtgg aattgtgagc ggataacaat ttcacacagg
     4861 aaacagctat gacatgatta cgaattaatt cgagctcgcc cgacattgat tattgactag
     4921 ttattaatag taatcaatta cggggtcatt agttcatagc caatatatgg agttccgcgt
     4981 tacataactt acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac
     5041 gtcaataatg acgtatgttc ccatagtaac gccaataggg actttccatt gacgtcaatg
     5101 ggtggagtat ttacggtaaa ctgcccactt ggcagtacat caagtgtatc atatgccaag
     5161 tacgccccct attgacgtca atgacggtaa atggcccgcc tggcattatg cccagtacat
     5221 gaccttatgg gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat
     5281 ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag cggtttgact cacggggatt
     5341 tccaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa atcaacggga
     5401 ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa atgggcggta ggcgtgtacg
     5461 gtgggaggtc tatataagca gagctcgttt agtgaaccgt cagatcgcct ggagacgcca
     5521 tccacgctgt tttgacctcc atagaagaca ccgggaccga tccagcctcc gcggccggga
     5581 acggtgcatt ggaacgcgga ttccccgtgc caagagtgac gtaagtaccg cctatagagt
     5641 ctataggccc acccccttgg cttcgttaga acgcggctac aattaataca taaccttatg
     5701 tatcatacac atacgattta ggtgacacta tagaataaca tccactttgc ctttctctcc
     5761 acaggtgtcc a
//
