LOCUS       sequence-403597-        6488 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  E. coli expression of HNRNPA1 low-complexity domain (186-320) with 
            an N-terminal mCherry tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6488)
  AUTHORS   Dumelie JG, Chen Q, Miller D, Attarwala N, Gross SS, Jaffrey SR
  TITLE     Biomolecular condensates create phospholipid-enriched 
            microenvironments.
  JOURNAL   Nat Chem Biol. 2023 Nov 16. doi: 10.1038/s41589-023-01474-4.
  PUBMED    37973889
REFERENCE   2  (bases 1 to 6488)
  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: "Nat Chem 
            Biol. 2023 Nov 16. doi: 10.1038/s41589-023-01474-4."
COMMENT     Sequence Label: pET28C-mCherry-HNRNPA1LCD
FEATURES             Location/Qualifiers
     source          1..6488
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             562..573
                     /codon_start=1
                     /product="WELQut protease recognition and cleavage site"
                     /label=WELQut site
                     /translation="WELQ"
     CDS             742..759
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(812..830)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      826..873
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      910..1365
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(997..1016)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1207..1228
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1457..2272)
                     /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(1457..2272)
                     /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     2180..2199
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2264..2283
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2394..2982
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2883..2902
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3136..3153
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3168..3307
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3233..3252)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3393..3415
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3409..3600)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4409..5491)
                     /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     5453..5472
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5492..5569)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5775..5794
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     5878..5897
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5878..5896
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5897..5921
                     /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             5936..5958
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             5977..5994
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             6004..6021
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             6025..6057
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             join(6064..6488,1..283)
                     /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(6209..6227)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     join(6477..6488,1..8)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
ORIGIN
        1 gaagaagacc atgggctggg aggcctcctc cgagcggatg taccccgagg acggcgccct
       61 gaagggcgag atcaagcaga ggctgaagct gaaggacggc ggccactacg acgctgaggt
      121 caagaccacc tacaaggcca agaagcccgt gcagctgccc ggcgcctaca acgtcaacat
      181 caagttggac atcacctccc acaacgagga ctacaccatc gtggaacagt acgaacgcgc
      241 cgagggccgc cactccaccg gcggcatgga cgagctgtac aagggtgcgc ctggttctgc
      301 aggctccgca gctggcggat cggggatcat ggctagtgct tcatccagcc aaagaggtcg
      361 aagtggttct ggaaactttg gtggtggtcg tggaggtggt ttcggtggga atgacaactt
      421 cggtcgtgga ggaaacttca gtggtcgtgg tggctttggt ggcagccgtg gtggtggtgg
      481 atatggtggc agtggggatg gctataatgg atttggtaat gatggaagca attttggagg
      541 tggtggaagc tacaatgatt ttgggaatta caacaatcag tcttcaaatt ttggacccat
      601 gaagggagga aattttggag gcagaagctc tggcccctat ggcggtggag gccaatactt
      661 tgcaaaacca cgaaaccaag gtggctatgg cggttccagc agcagcagta gctatggcag
      721 tggcagaaga ttttagtcga gcaccaccac caccaccact gagatccggc tgctaacaaa
      781 gcccgaaagg aagctgagtt ggctgctgcc accgctgagc aataactagc ataacccctt
      841 ggggcctcta aacgggtctt gaggggtttt ttgctgaaag gaggaactat atccggattg
      901 gcgaatggga cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca
      961 gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct
     1021 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt
     1081 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac
     1141 gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct
     1201 ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt
     1261 ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac
     1321 aaaaatttaa cgcgaatttt aacaaaatat taacgcttac aatttaggtg gcacttttcg
     1381 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
     1441 gctcatgaat taattcttag aaaaactcat cgagcatcaa atgaaactgc aatttattca
     1501 tatcaggatt atcaatacca tatttttgaa aaagccgttt ctgtaatgaa ggagaaaact
     1561 caccgaggca gttccatagg atggcaagat cctggtatcg gtctgcgatt ccgactcgtc
     1621 caacatcaat acaacctatt aatttcccct cgtcaaaaat aaggttatca agtgagaaat
     1681 caccatgagt gacgactgaa tccggtgaga atggcaaaag tttatgcatt tctttccaga
     1741 cttgttcaac aggccagcca ttacgctcgt catcaaaatc actcgcatca accaaaccgt
     1801 tattcattcg tgattgcgcc tgagcgagac gaaatacgcg atcgctgtta aaaggacaat
     1861 tacaaacagg aatcgaatgc aaccggcgca ggaacactgc cagcgcatca acaatatttt
     1921 cacctgaatc aggatattct tctaatacct ggaatgctgt tttcccgggg atcgcagtgg
     1981 tgagtaacca tgcatcatca ggagtacgga taaaatgctt gatggtcgga agaggcataa
     2041 attccgtcag ccagtttagt ctgaccatct catctgtaac atcattggca acgctacctt
     2101 tgccatgttt cagaaacaac tctggcgcat cgggcttccc atacaatcga tagattgtcg
     2161 cacctgattg cccgacatta tcgcgagccc atttataccc atataaatca gcatccatgt
     2221 tggaatttaa tcgcggccta gagcaagacg tttcccgttg aatatggctc ataacacccc
     2281 ttgtattact gtttatgtaa gcagacagtt ttattgttca tgaccaaaat cccttaacgt
     2341 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     2401 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     2461 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     2521 gcgcagatac caaatactgt ccttctagtg tagccgtagt taggccacca cttcaagaac
     2581 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     2641 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     2701 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     2761 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     2821 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     2881 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     2941 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     3001 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
     3061 cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc
     3121 cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgcct gatgcggtat
     3181 tttctcctta cgcatctgtg cggtatttca caccgcaatg gtgcactctc agtacaatct
     3241 gctctgatgc cgcatagtta agccagtata cactccgcta tcgctacgtg actgggtcat
     3301 ggctgcgccc cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc
     3361 ggcatccgct tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc
     3421 accgtcatca ccgaaacgcg cgaggcagct gcggtaaagc tcatcagcgt ggtcgtgaag
     3481 cgattcacag atgtctgcct gttcatccgc gtccagctcg ttgagtttct ccagaagcgt
     3541 taatgtctgg cttctgataa agcgggccat gttaagggcg gttttttcct gtttggtcac
     3601 tgatgcctcc gtgtaagggg gatttctgtt catgggggta atgataccga tgaaacgaga
     3661 gaggatgctc acgatacggg ttactgatga tgaacatgcc cggttactgg aacgttgtga
     3721 gggtaaacaa ctggcggtat ggatgcggcg ggaccagaga aaaatcactc agggtcaatg
     3781 ccagcgcttc gttaatacag atgtaggtgt tccacagggt agccagcagc atcctgcgat
     3841 gcagatccgg aacataatgg tgcagggcgc tgacttccgc gtttccagac tttacgaaac
     3901 acggaaaccg aagaccattc atgttgttgc tcaggtcgca gacgttttgc agcagcagtc
     3961 gcttcacgtt cgctcgcgta tcggtgattc attctgctaa ccagtaaggc aaccccgcca
     4021 gcctagccgg gtcctcaacg acaggagcac gatcatgcgc acccgtgggg ccgccatgcc
     4081 ggcgataatg gcctgcttct cgccgaaacg tttggtggcg ggaccagtga cgaaggcttg
     4141 agcgagggcg tgcaagattc cgaataccgc aagcgacagg ccgatcatcg tcgcgctcca
     4201 gcgaaagcgg tcctcgccga aaatgaccca gagcgctgcc ggcacctgtc ctacgagttg
     4261 catgataaag aagacagtca taagtgcggc gacgatagtc atgccccgcg cccaccggaa
     4321 ggagctgact gggttgaagg ctctcaaggg catcggtcga gatcccggtg cctaatgagt
     4381 gagctaactt acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     4441 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     4501 ccagggtggt ttttcttttc accagtgaga cgggcaacag ctgattgccc ttcaccgcct
     4561 ggccctgaga gagttgcagc aagcggtcca cgctggtttg ccccagcagg cgaaaatcct
     4621 gtttgatggt ggttaacggc gggatataac atgagctgtc ttcggtatcg tcgtatccca
     4681 ctaccgagat atccgcacca acgcgcagcc cggactcggt aatggcgcgc attgcgccca
     4741 gcgccatctg atcgttggca accagcatcg cagtgggaac gatgccctca ttcagcattt
     4801 gcatggtttg ttgaaaaccg gacatggcac tccagtcgcc ttcccgttcc gctatcggct
     4861 gaatttgatt gcgagtgaga tatttatgcc agccagccag acgcagacgc gccgagacag
     4921 aacttaatgg gcccgctaac agcgcgattt gctggtgacc caatgcgacc agatgctcca
     4981 cgcccagtcg cgtaccgtct tcatgggaga aaataatact gttgatgggt gtctggtcag
     5041 agacatcaag aaataacgcc ggaacattag tgcaggcagc ttccacagca atggcatcct
     5101 ggtcatccag cggatagtta atgatcagcc cactgacgcg ttgcgcgaga agattgtgca
     5161 ccgccgcttt acaggcttcg acgccgcttc gttctaccat cgacaccacc acgctggcac
     5221 ccagttgatc ggcgcgagat ttaatcgccg cgacaatttg cgacggcgcg tgcagggcca
     5281 gactggaggt ggcaacgcca atcagcaacg actgtttgcc cgccagttgt tgtgccacgc
     5341 ggttgggaat gtaattcagc tccgccatcg ccgcttccac tttttcccgc gttttcgcag
     5401 aaacgtggct ggcctggttc accacgcggg aaacggtctg ataagagaca ccggcatact
     5461 ctgcgacatc gtataacgtt actggtttca cattcaccac cctgaattga ctctcttccg
     5521 ggcgctatca tgccataccg cgaaaggttt tgcgccattc gatggtgtcc gggatctcga
     5581 cgctctccct tatgcgactc ctgcattagg aagcagccca gtagtaggtt gaggccgttg
     5641 agcaccgccg ccgcaaggaa tggtgcatgc aaggagatgg cgcccaacag tcccccggcc
     5701 acggggcctg ccaccatacc cacgccgaaa caagcgctca tgagcccgaa gtggcgagcc
     5761 cgatcttccc catcggtgat gtcggcgata taggcgccag caaccgcacc tgtggcgccg
     5821 gtgatgccgg ccacgatgcg tccggcgtag aggatcgaga tctcgatccc gcgaaattaa
     5881 tacgactcac tataggggaa ttgtgagcgg ataacaattc ccctctagaa ataattttgt
     5941 ttaactttaa gaaggagata taccatgggc agcagccatc atcatcatca tcacagcagc
     6001 ggcctggtgc cgcgcggcag ccatatggct agcatgactg gtggacagca aatgggtcgg
     6061 atcatggtga gcaagggcga ggaggataac atggccatca tcaaggagtt catgcgcttc
     6121 aaggtgcaca tggagggctc cgtgaacggc cacgagttcg agatcgaggg cgagggcgag
     6181 ggccgcccct acgagggcac ccagaccgcc aagctgaagg tgaccaaggg tggccccctg
     6241 cccttcgcct gggacatcct gtcccctcag ttcatgtacg gctccaaggc ctacgtgaag
     6301 caccccgccg acatccccga ctacttgaag ctgtccttcc ccgagggctt caagtgggag
     6361 cgcgtgatga acttcgagga cggcggcgtg gtgaccgtga cccaggactc ctccctgcag
     6421 gacggcgagt tcatctacaa ggtgaagctg cgcggcacca acttcccctc cgacggcccc
     6481 gtaatgca
//
