LOCUS       sequence-274850-        5781 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  express FUS LC with hnRNPA2-like charge patterning where all R are 
            changed to K.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5781)
  AUTHORS   Ryan VH, Perdikari TM, Naik MT, Saueressig CF, Lins J, Dignon GL, 
            Mittal J, Hart AC, Fawzi NL
  TITLE     Tyrosine phosphorylation regulates hnRNPA2 granule protein 
            partitioning and reduces neurodegeneration.
  JOURNAL   EMBO J. 2021 Feb 1;40(3):e105001. doi: 10.15252/embj.2020105001. 
            Epub 2020 Dec 22.
  PUBMED    33349959
REFERENCE   2  (bases 1 to 5781)
  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: "EMBO J."; 
            date: "2021-02-1"; volume: "40(3):e105001. doi"; pages: " 
            10.15252/embj.2020105001. Epub 2020 Dec 22"
COMMENT     Sequence Label: FUS_LC_charge_RtoK
FEATURES             Location/Qualifiers
     source          1..5781
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     44..63
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     147..166
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        147..165
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    166..190
                     /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)."
     RBS             205..227
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             252..269
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             270..290
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             792..809
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(862..880)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      876..923
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      960..1415
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1047..1066)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1257..1278
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1507..2322)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1507..2322)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2230..2249
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2314..2333
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2444..3032
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2933..2952
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3186..3203
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3218..3357
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3283..3302)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3443..3465
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3459..3650)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4459..5541)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     5503..5522
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5542..5619)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 aagcgctcat gagcccgaag tggcgagccc gatcttcccc atcggtgatg tcggcgatat
       61 aggcgccagc aaccgcacct gtggcgccgg tgatgccggc cacgatgcgt ccggcgtaga
      121 ggatcgagat ctcgatcccg cgaaattaat acgactcact ataggggaat tgtgagcgga
      181 taacaattcc cctctagaaa taattttgtt taactttaag aaggagatat accatgggct
      241 ctgataaaat tcatcatcat catcatcacg aaaatcttta ttttcaaggt catatggcga
      301 aaaacgacta cacccaacag gcgacccaaa agtatggtgc gtatccgacc cagccgggcc
      361 aaggttacag ccagcaaagc aagcaaccgt acggtcagga cagctacagc ggctataaac
      421 aaagcaccga taccagcggt tacggccaga gcagctacag cagctatggt caaagccaga
      481 acaccggtta tggcacccaa agcaccccgc agggttacgg cagcaagggt ggctatggta
      541 gcagccaaag cagccagagc agctacggcc agcaaagcag ctatccgggt tatggtgatc
      601 aaccggcgcc gagcgatacc agcggtagct acggtagcag cagccaaagc agcagctacg
      661 gtcagccgaa gagcggcagc tacagccagc aaccgaaata tggtggccag caacagagct
      721 atggtcagca acaaagctac aacccgccgc agggctacgg tcagcaaaaa caatacaaaa
      781 gctaactcga gcaccaccac caccaccact gagatccggc tgctaacaaa gcccgaaagg
      841 aagctgagtt ggctgctgcc accgctgagc aataactagc ataacccctt ggggcctcta
      901 aacgggtctt gaggggtttt ttgctgaaag gaggaactat atccggattg gcgaatggga
      961 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     1021 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     1081 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     1141 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
     1201 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     1261 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
     1321 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     1381 cgcgaatttt aacaaaatat taacgcttac aatttaggtg gcacttttcg gggaaatgtg
     1441 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaat
     1501 taattcttag aaaaactcat cgagcatcaa atgaaactgc aatttattca tatcaggatt
     1561 atcaatacca tatttttgaa aaagccgttt ctgtaatgaa ggagaaaact caccgaggca
     1621 gttccatagg atggcaagat cctggtatcg gtctgcgatt ccgactcgtc caacatcaat
     1681 acaacctatt aatttcccct cgtcaaaaat aaggttatca agtgagaaat caccatgagt
     1741 gacgactgaa tccggtgaga atggcaaaag tttatgcatt tctttccaga cttgttcaac
     1801 aggccagcca ttacgctcgt catcaaaatc actcgcatca accaaaccgt tattcattcg
     1861 tgattgcgcc tgagcgagac gaaatacgcg atcgctgtta aaaggacaat tacaaacagg
     1921 aatcgaatgc aaccggcgca ggaacactgc cagcgcatca acaatatttt cacctgaatc
     1981 aggatattct tctaatacct ggaatgctgt tttcccgggg atcgcagtgg tgagtaacca
     2041 tgcatcatca ggagtacgga taaaatgctt gatggtcgga agaggcataa attccgtcag
     2101 ccagtttagt ctgaccatct catctgtaac atcattggca acgctacctt tgccatgttt
     2161 cagaaacaac tctggcgcat cgggcttccc atacaatcga tagattgtcg cacctgattg
     2221 cccgacatta tcgcgagccc atttataccc atataaatca gcatccatgt tggaatttaa
     2281 tcgcggccta gagcaagacg tttcccgttg aatatggctc ataacacccc ttgtattact
     2341 gtttatgtaa gcagacagtt ttattgttca tgaccaaaat cccttaacgt gagttttcgt
     2401 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc
     2461 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc
     2521 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac
     2581 caaatactgt ccttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac
     2641 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt
     2701 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct
     2761 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat
     2821 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt
     2881 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg
     2941 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt
     3001 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt
     3061 tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg
     3121 tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg
     3181 agcgcagcga gtcagtgagc gaggaagcgg aagagcgcct gatgcggtat tttctcctta
     3241 cgcatctgtg cggtatttca caccgcaatg gtgcactctc agtacaatct gctctgatgc
     3301 cgcatagtta agccagtata cactccgcta tcgctacgtg actgggtcat ggctgcgccc
     3361 cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct
     3421 tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca
     3481 ccgaaacgcg cgaggcagct gcggtaaagc tcatcagcgt ggtcgtgaag cgattcacag
     3541 atgtctgcct gttcatccgc gtccagctcg ttgagtttct ccagaagcgt taatgtctgg
     3601 cttctgataa agcgggccat gttaagggcg gttttttcct gtttggtcac tgatgcctcc
     3661 gtgtaagggg gatttctgtt catgggggta atgataccga tgaaacgaga gaggatgctc
     3721 acgatacggg ttactgatga tgaacatgcc cggttactgg aacgttgtga gggtaaacaa
     3781 ctggcggtat ggatgcggcg ggaccagaga aaaatcactc agggtcaatg ccagcgcttc
     3841 gttaatacag atgtaggtgt tccacagggt agccagcagc atcctgcgat gcagatccgg
     3901 aacataatgg tgcagggcgc tgacttccgc gtttccagac tttacgaaac acggaaaccg
     3961 aagaccattc atgttgttgc tcaggtcgca gacgttttgc agcagcagtc gcttcacgtt
     4021 cgctcgcgta tcggtgattc attctgctaa ccagtaaggc aaccccgcca gcctagccgg
     4081 gtcctcaacg acaggagcac gatcatgcgc acccgtgggg ccgccatgcc ggcgataatg
     4141 gcctgcttct cgccgaaacg tttggtggcg ggaccagtga cgaaggcttg agcgagggcg
     4201 tgcaagattc cgaataccgc aagcgacagg ccgatcatcg tcgcgctcca gcgaaagcgg
     4261 tcctcgccga aaatgaccca gagcgctgcc ggcacctgtc ctacgagttg catgataaag
     4321 aagacagtca taagtgcggc gacgatagtc atgccccgcg cccaccggaa ggagctgact
     4381 gggttgaagg ctctcaaggg catcggtcga gatcccggtg cctaatgagt gagctaactt
     4441 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     4501 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ccagggtggt
     4561 ttttcttttc accagtgaga cgggcaacag ctgattgccc ttcaccgcct ggccctgaga
     4621 gagttgcagc aagcggtcca cgctggtttg ccccagcagg cgaaaatcct gtttgatggt
     4681 ggttaacggc gggatataac atgagctgtc ttcggtatcg tcgtatccca ctaccgagat
     4741 atccgcacca acgcgcagcc cggactcggt aatggcgcgc attgcgccca gcgccatctg
     4801 atcgttggca accagcatcg cagtgggaac gatgccctca ttcagcattt gcatggtttg
     4861 ttgaaaaccg gacatggcac tccagtcgcc ttcccgttcc gctatcggct gaatttgatt
     4921 gcgagtgaga tatttatgcc agccagccag acgcagacgc gccgagacag aacttaatgg
     4981 gcccgctaac agcgcgattt gctggtgacc caatgcgacc agatgctcca cgcccagtcg
     5041 cgtaccgtct tcatgggaga aaataatact gttgatgggt gtctggtcag agacatcaag
     5101 aaataacgcc ggaacattag tgcaggcagc ttccacagca atggcatcct ggtcatccag
     5161 cggatagtta atgatcagcc cactgacgcg ttgcgcgaga agattgtgca ccgccgcttt
     5221 acaggcttcg acgccgcttc gttctaccat cgacaccacc acgctggcac ccagttgatc
     5281 ggcgcgagat ttaatcgccg cgacaatttg cgacggcgcg tgcagggcca gactggaggt
     5341 ggcaacgcca atcagcaacg actgtttgcc cgccagttgt tgtgccacgc ggttgggaat
     5401 gtaattcagc tccgccatcg ccgcttccac tttttcccgc gttttcgcag aaacgtggct
     5461 ggcctggttc accacgcggg aaacggtctg ataagagaca ccggcatact ctgcgacatc
     5521 gtataacgtt actggtttca cattcaccac cctgaattga ctctcttccg ggcgctatca
     5581 tgccataccg cgaaaggttt tgcgccattc gatggtgtcc gggatctcga cgctctccct
     5641 tatgcgactc ctgcattagg aagcagccca gtagtaggtt gaggccgttg agcaccgccg
     5701 ccgcaaggaa tggtgcatgc aaggagatgg cgcccaacag tcccccggcc acggggcctg
     5761 ccaccatacc cacgccgaaa c
//
