LOCUS       sequence-486127-        5920 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express RILPL1 in mammalian cells with mCherry tag expressing a 
            R293A mutation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5920)
  AUTHORS   Bonet-Ponce L, Tegicho T, Fernandez-Martinez N, Rozenberg IA, 
            Ashriem M, Beilina A, Kluss JH, Li Y, Cookson MR
  TITLE     JIP4 and RILPL1 utilize opposing motor force to dynamically regulate
            lysosomal tubulation.
  JOURNAL   J Cell Biol. 2025 Nov 3;224(11):e202404018. doi: 
            10.1083/jcb.202404018. Epub 2025 Sep 24.
  PUBMED    40990465
REFERENCE   2  (bases 1 to 5920)
  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: "J Cell 
            Biol."; date: "2025-11-3"; volume: "224(11):e202404018. doi"; pages:
            " 10.1083/jcb.202404018. Epub 2025 Sep 24"
COMMENT     Sequence Label: mCherry-RILPL1 (R293A)
FEATURES             Location/Qualifiers
     source          1..5920
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        68..371
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        372..575
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     526..546
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      614..623
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             620..1327
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(765..783)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1033..1052
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    2715..2836
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2752..2771)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2806..2825
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2843..3298)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2980..3001)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3192..3211
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3325..3429
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3431..3788
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3431..3622
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3454..3474)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3639..3774
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3823..4617
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             3823..4617
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3877..3896)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4487..4506
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4805..4824)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4849..4896
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5225..5813
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5714..5733
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 catgcattag ttattaatag taatcaatta cggggtcatt agttcatagc ccatatatgg
       61 agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc
      121 gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg actttccatt
      181 gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat caagtgtatc
      241 atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc tggcattatg
      301 cccagtacat gaccttatgg gactttccta cttggcagta catctacgta ttagtcatcg
      361 ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag cggtttgact
      421 cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa
      481 atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa atgggcggta
      541 ggcgtgtacg gtgggaggtc tatataagca gagctggttt agtgaaccgt cagatccgct
      601 agcgctaccg gtcgccacca tggtgagcaa gggcgaggag gataacatgg ccatcatcaa
      661 ggagttcatg cgcttcaagg tgcacatgga gggctccgtg aacggccacg agttcgagat
      721 cgagggcgag ggcgagggcc gcccctacga gggcacccag accgccaagc tgaaggtgac
      781 caagggtggc cccctgccct tcgcctggga catcctgtcc cctcagttca tgtacggctc
      841 caaggcctac gtgaagcacc ccgccgacat ccccgactac ttgaagctgt ccttccccga
      901 gggcttcaag tgggagcgcg tgatgaactt cgaggacggc ggcgtggtga ccgtgaccca
      961 ggactcctcc ctgcaggacg gcgagttcat ctacaaggtg aagctgcgcg gcaccaactt
     1021 cccctccgac ggccccgtaa tgcagaagaa gaccatgggc tgggaggcct cctccgagcg
     1081 gatgtacccc gaggacggcg ccctgaaggg cgagatcaag cagaggctga agctgaagga
     1141 cggcggccac tacgacgctg aggtcaagac cacctacaag gccaagaagc ccgtgcagct
     1201 gcccggcgcc tacaacgtca acatcaagtt ggacatcacc tcccacaacg aggactacac
     1261 catcgtggaa cagtacgaac gcgccgaggg ccgccactcc accggcggca tggacgagct
     1321 gtacaagtcc ggactcagat ctatggagga ggagcggggg tcggcgctgg cggccgagtc
     1381 ggcgctggag aagaacgtgg ccgagctgac cgtcatggac gtgtacgaca tcgcgtcgct
     1441 tgtgggccac gagttcgagc gggtcattga ccagcacggc tgcgaggcca tcgcgcgcct
     1501 catgcccaag gtcgtgcgcg tcctggagat cctggaggtg ctggtcagcc gccaccacgt
     1561 cgcgcccgag ctggacgagc tgcgcctgga gctggaccgc ctgcgcctgg agaggatgga
     1621 ccgcatcgag aaggagcgca agcaccagaa ggagctggag ctggtggagg atgtgtggcg
     1681 aggggaggcg caggacctcc tctcccagat cgcccagctg caggaggaga acaagcagct
     1741 catgaccaac ctctcccaca aggatgtcaa cttctcagag gaggagttcc agaagcatga
     1801 aggcatgtca gagcgggagc gacaggtgat gaagaagctg aaggaggtgg tggacaaaca
     1861 acgcgacgag atccgcgcca aggacaggga gctgggcctg aaaaatgagg acgttgaggc
     1921 tttacagcag cagcagacac ggctgatgaa gatcaaccat gaccttcggc accgggtcac
     1981 ggtggtggag gcccagggga aagccctgat cgaacagaag gtggagctgg aggcagacct
     2041 gcagaccaag gagcaggaga tgggcagcct gcgagcagag ctggggaagt tgcgagagag
     2101 gctgcagggg gagcacagcc agaatgggga ggaggagcct gagacggagc cggtgggaga
     2161 ggagagcatc tccgacgcag agaaggtggc catggatctc aaggacccca accgccccgc
     2221 gttcaccctg caggagctgc gggacgtgct gcacgagagg aacgagctca agtccaaggt
     2281 gttcttgctg caggaggagc tggcttacta taagagtgaa gaaatggaag aggaaaaccg
     2341 aataccccaa cccccaccca tcgcccaccc gaggacgtcc ccccagccgg agtcgggcat
     2401 caagcgactg tttagcttct tctcccgaga taagaagcgc ctggccaaca cacagagaaa
     2461 cgtgcacatc caggagtcct ttggacagtg ggcaaacacc caccgcgatg acggttacac
     2521 agagcaagga caggaagccc tgcagcatct gtgagaattc tgcagtcgac ggtaccgcgg
     2581 gcccgggatc caccggatct agataactga tcataatcag ccataccaca tttgtagagg
     2641 ttttacttgc tttaaaaaac ctcccacacc tccccctgaa cctgaaacat aaaatgaatg
     2701 caattgttgt tgttaacttg tttattgcag cttataatgg ttacaaataa agcaatagca
     2761 tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac
     2821 tcatcaatgt atcttaacgc gtaaattgta agcgttaata ttttgttaaa attcgcgtta
     2881 aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat
     2941 aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca
     3001 ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc
     3061 ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg taaagcacta
     3121 aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg
     3181 gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg
     3241 gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtca
     3301 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     3361 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
     3421 aggaagagtc ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa
     3481 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     3541 ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
     3601 attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca
     3661 gttccgccca ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg
     3721 ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct
     3781 tttgcaaaga tcgatcaaga gacaggatga ggatcgtttc gcatgattga acaagatgga
     3841 ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa
     3901 cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt
     3961 ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaagacga ggcagcgcgg
     4021 ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa
     4081 gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac
     4141 cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt
     4201 gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact
     4261 cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca ggggctcgcg
     4321 ccagccgaac tgttcgccag gctcaaggcg agcatgcccg acggcgagga tctcgtcgtg
     4381 acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc
     4441 atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt
     4501 gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc
     4561 gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgagcg
     4621 ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg
     4681 attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct
     4741 ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccaccct agggggaggc
     4801 taactgaaac acggaaggag acaataccgg aaggaacccg cgctatgacg gcaataaaaa
     4861 gacagaataa aacgcacggt gttgggtcgt ttgttcataa acgcggggtt cggtcccagg
     4921 gctggcactc tgtcgatacc ccaccgagac cccattgggg ccaatacgcc cgcgtttctt
     4981 ccttttcccc accccacccc ccaagttcgg gtgaaggccc agggctcgca gccaacgtcg
     5041 gggcggcagg ccctgccata gcctcaggtt actcatatat actttagatt gatttaaaac
     5101 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     5161 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     5221 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     5281 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     5341 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
     5401 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     5461 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     5521 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     5581 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     5641 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     5701 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     5761 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     5821 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     5881 ctgcgttatc ccctgattct gtggataacc gtattaccgc
//
