LOCUS       sequence-339776-        5945 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  bacterial experssion of a GST-tagged MEG-3 C-terminus with mutations
            in the HMG-like motif.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5945)
  AUTHORS   Schmidt H, Putnam A, Rasoloson D, Seydoux G
  TITLE     Coordination of RNA and protein condensation by the P granule 
            protein MEG-3
  JOURNAL   bioRxiv 2020.10.15.340570
REFERENCE   2  (bases 1 to 5945)
  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: "bioRxiv 
            2020.10.15.340570"
COMMENT     Sequence Label: pHS33
FEATURES             Location/Qualifiers
     source          1..5945
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      7..595
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     496..515
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     749..766
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        839..916
                     /gene="lacI"
                     /label=lacI promoter
     CDS             917..1999
                     /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     complement(936..955)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(2048..2053,2054..2071,2072..2078)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2086..2102
                     /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     2110..2126
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2110..2126
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             2122..2649
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(2142..2159)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2142..2158)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2151..2173)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     promoter        join(2751..2756,2757..2772,2773..2779)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    2787..2803
                     /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)."
     CDS             2826..3479
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     3437..3459
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             3486..3509
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(4563..4585)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4730..4748)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4816..4920
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4921..4989,4990..5781)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5139..5158)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
       61 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
      121 tggcttcagc agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca
      181 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
      241 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
      301 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
      361 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
      421 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
      481 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
      541 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
      601 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
      661 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac
      721 cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg
      781 cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca taaattccga
      841 caccatcgaa tggcgcaaaa cctttcgcgg tatggcatga tagcgcccgg aagagagtca
      901 attcagggtg gtgaatgtga aaccagtaac gttatacgat gtcgcagagt atgccggtgt
      961 ctcttatcag accgtttccc gcgtggtgaa ccaggccagc cacgtttctg cgaaaacgcg
     1021 ggaaaaagtg gaagcggcga tggcggagct gaattacatt cccaaccgcg tggcacaaca
     1081 actggcgggc aaacagtcgt tgctgattgg cgttgccacc tccagtctgg ccctgcacgc
     1141 gccgtcgcaa attgtcgcgg cgattaaatc tcgcgccgat caactgggtg ccagcgtggt
     1201 ggtgtcgatg gtagaacgaa gcggcgtcga agcctgtaaa gcggcggtgc acaatcttct
     1261 cgcgcaacgc gtcagtgggc tgatcattaa ctatccgctg gatgaccagg atgccattgc
     1321 tgtggaagct gcctgcacta atgttccggc gttatttctt gatgtctctg accagacacc
     1381 catcaacagt attattttct cccatgaaga cggtacgcga ctgggcgtgg agcatctggt
     1441 cgcattgggt caccagcaaa tcgcgctgtt agcgggccca ttaagttctg tctcggcgcg
     1501 tctgcgtctg gctggctggc ataaatatct cactcgcaat caaattcagc cgatagcgga
     1561 acgggaaggc gactggagtg ccatgtccgg ttttcaacaa accatgcaaa tgctgaatga
     1621 gggcatcgtt cccactgcga tgctggttgc caacgatcag atggcgctgg gcgcaatgcg
     1681 cgccattacc gagtccgggc tgcgcgttgg tgcggatatc tcggtagtgg gatacgacga
     1741 taccgaagac agctcatgtt atatcccgcc gttaaccacc atcaaacagg attttcgcct
     1801 gctggggcaa accagcgtgg accgcttgct gcaactctct cagggccagg cggtgaaggg
     1861 caatcagctg ttgcccgtct cactggtgaa aagaaaaacc acgctagcgc ccaatacgca
     1921 aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg
     1981 actggaaagc gggcagtgag cgcaacgcaa ttaatgtaag ttagctcact cattaggcac
     2041 cccaggcttt acactttatg cttccggctc gtatgttgtg tggaattgtg agcggataac
     2101 aatttcacac aggaaacagc tatgaccatg attacggatt cactggccgt cgttttacaa
     2161 cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc acatccccct
     2221 ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca acagttgcgc
     2281 agcctgaatg gcgaatggcg ctttgcctgg tttccggcac cagaagcggt gccggaaagc
     2341 tggctggagt gcgatcttcc tgaggccgat actgtcgtcg tcccctcaaa ctggcagatg
     2401 cacggttacg atgcgcccat ctacaccaac gtgacctatc ccattacggt caatccgccg
     2461 tttgttccca cggagaatcc gacgggttgt tactcgctca catttaatgt tgatgaaagc
     2521 tggctacagg aaggccagac gcgaattatt tttgatggcg ttggaattag cttatcgact
     2581 gcacggtgca ccaatgcttc tggcgtcagg cagccatcgg aagctgtggt atggctgtgc
     2641 aggtcgtaaa tcactgcata attcgtgtcg ctcaaggcgc actcccgttc tggataatgt
     2701 tttttgcgcc gacatcataa cggttctggc aaatattctg aaatgagctg ttgacaatta
     2761 atcatcggct cgtataatgt gtggaattgt gagcggataa caatttcaca caggaaacag
     2821 tattcatgtc ccctatacta ggttattgga aaattaaggg ccttgtgcaa cccactcgac
     2881 ttcttttgga atatcttgaa gaaaaatatg aagagcattt gtatgagcgc gatgaaggtg
     2941 ataaatggcg aaacaaaaag tttgaattgg gtttggagtt tcccaatctt ccttattata
     3001 ttgatggtga tgttaaatta acacagtcta tggccatcat acgttatata gctgacaagc
     3061 acaacatgtt gggtggttgt ccaaaagagc gtgcagagat ttcaatgctt gaaggagcgg
     3121 ttttggatat tagatacggt gtttcgagaa ttgcatatag taaagacttt gaaactctca
     3181 aagttgattt tcttagcaag ctacctgaaa tgctgaaaat gttcgaagat cgtttatgtc
     3241 ataaaacata tttaaatggt gatcatgtaa cccatcctga cttcatgttg tatgacgctc
     3301 ttgatgttgt tttatacatg gacccaatgt gcctggatgc gttcccaaaa ttagtttgtt
     3361 ttaaaaaacg tattgaagct atcccacaaa ttgataagta cttgaaatcc agcaagtata
     3421 tagcatggcc tttgcagggc tggcaagcca cgtttggtgg tggcgaccat cctccaaaat
     3481 cggatctgga agttctgttc caggggcccc tgggatcccc ggaattcccg ggtcgacaca
     3541 tgtcgtcaat catcccaact ccaaaggctt tgatcggaga aagatgtctg acatcatcca
     3601 gcaagtcttc aaagttgaac aagtcacttg gcgtggttga ttctaaagct acaaaatcac
     3661 caatgtattc cgtcacagtt tctggaaaag aaacagcttc aggaaagcga attgctcaaa
     3721 agttgactcc aaaggttgtg gcgctggaaa gctcttacat taccggaatt ccagtttcca
     3781 cggattgtaa tggttgccca actccgaaga gaagtggaat caattgtgaa attcgtgcag
     3841 ctgaagtata caatcaagct gggaaatggc ccttcgaaat caccagtgat ccggctccac
     3901 ttccttgtga atcagccgat cggatcgaat atccttctca agattgcact caagatccag
     3961 cttcaacctc gccaccacct cgcatttctg aaagcttgac agcattcctg gaagctcagc
     4021 aagacgccaa cgacgctatt gatactaacg ccaaagaaaa gacacaactc ctgaaagtga
     4081 atttggctat tcacgggatg tcacctgaaa gatggctgta cttgaattat ttttgcaccg
     4141 agacaattcc aagattggat ggtccttatg ccgatgatcc aagagttcca cctgtgcgaa
     4201 atatgttcag aaaatggttc ctccgttttg ctgaagcctg cttaggaaat cctcaccagc
     4261 ttgcagttat gcaagagatt gctgccacgt ttgttcaggc tcggctggac gatacgagca
     4321 gctccacaga ctccaccaat atgctctaca tgttgtggaa ggagtgtatc ggacagaaga
     4381 atatcatcgc aattgctgat gcgtgtttac ttgcacatct tcggaaatcg gatcctatca
     4441 aatacttgaa tgtaaaacgg gattggctgg aatctatttt tgacccaccc agagatcaat
     4501 gagcggccgc atcgtgactg actgacgatc tgcctcgcgc gtttcggtga tgacggtgaa
     4561 aacctctgac acatgcagct cccggagacg gtcacagctt gtctgtaagc ggatgccggg
     4621 agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg cgcagccatg
     4681 acccagtcac gtagcgatag cggagtgtat aattcttgaa gacgaaaggg cctcgtgata
     4741 cgcctatttt tataggttaa tgtcatgata ataatggttt cttagacgtc aggtggcact
     4801 tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg
     4861 tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt
     4921 atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct
     4981 gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca
     5041 cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc
     5101 gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc
     5161 cgtgttgacg ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg
     5221 gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta
     5281 tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc
     5341 ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt
     5401 gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg
     5461 cctgcagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct
     5521 tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc
     5581 tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct
     5641 cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac
     5701 acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc
     5761 tcactgatta agcattggta actgtcagac caagtttact catatatact ttagattgat
     5821 ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg
     5881 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     5941 aaagg
//
