LOCUS       sequence-430106-        5142 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Expresses 6xHis-tagged N- and C-terminal prion-like domains of Efg1 
            with Y4/Y8/Y16 to serine mutations.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5142)
  AUTHORS   Wang SH, Zheng T, Fawzi NL
  TITLE     Structure and position-specific interactions of prion-like domains 
            in transcription factor Efg1 phase separation.
  JOURNAL   bioRxiv [Preprint]. 2023 Nov 9:2023.11.09.566450. doi: 
            10.1101/2023.11.09.566450.
  PUBMED    37986834
REFERENCE   2  (bases 1 to 5142)
  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"; journalName: "bioRxiv 
            [Preprint]. 2023 Nov 9:2023.11.09.566450. doi: 
            10.1101/2023.11.09.566450."
COMMENT     Sequence Label: Efg1-Delta-DBD-Y4S/Y8S/Y16S
FEATURES             Location/Qualifiers
     source          1..5142
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             715..1797
                     /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(734..753)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     rep_origin      1883..2471
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2372..2391
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2625..2642
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     2740..2757
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2741..2757
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3221..3240
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3221..3239
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    3240..3264
                     /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             3325..3342
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             3343..3363
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     terminator      4517..4564
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(4787..4806)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     promoter        4873..4964
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4965..5142,1..632)
                     /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="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
ORIGIN
        1 ccaatgatgt tacagatgag atggtcagac taaactggct gacggaattt atgccacttc
       61 cgaccatcaa gcattttatc cgtactcctg atgatgcatg gttactcacc actgcgatcc
      121 ccggaaaaac agcgttccag gtattagaag aatatcctga ttcaggtgaa aatattgttg
      181 atgcgctggc agtgttcctg cgccggttgc actcgattcc tgtttgtaat tgtcctttta
      241 acagcgatcg cgtatttcgc ctcgctcagg cgcaatcacg aatgaataac ggtttggttg
      301 atgcgagtga ttttgatgac gagcgtaatg gctggcctgt tgaacaagtc tggaaagaaa
      361 tgcataaact tttgccattc tcaccggatt cagtcgtcac tcatggtgat ttctcacttg
      421 ataaccttat ttttgacgag gggaaattaa taggttgtat tgatgttgga cgagtcggaa
      481 tcgcagaccg ataccaggat cttgccatcc tatggaactg cctcggtgag ttttctcctt
      541 cattacagaa acggcttttt caaaaatatg gtattgataa tcctgatatg aataaattgc
      601 aatttcattt gatgctcgat gagtttttct aagcggcgcg ccatcgaatg gcgcaaaacc
      661 tttcgcggta tggcatgata gcgcccggaa gagagtcaat tcagggtggt gaatatgaaa
      721 ccagtaacgt tatacgatgt cgcagagtat gccggtgtct cttatcagac cgtttcccgc
      781 gtggtgaacc aggccagcca cgtttctgcg aaaacgcggg aaaaagtgga agcggcgatg
      841 gcggagctga attacattcc caaccgcgtg gcacaacaac tggcgggcaa acagtcgttg
      901 ctgattggcg ttgccacctc cagtctggcc ctgcacgcgc cgtcgcaaat tgtcgcggcg
      961 attaaatctc gcgccgatca actgggtgcc agcgtggtgg tgtcgatggt agaacgaagc
     1021 ggcgtcgaag cctgtaaagc ggcggtgcac aatcttctcg cgcaacgcgt cagtgggctg
     1081 atcattaact atccgctgga tgaccaggat gccattgctg tggaagctgc ctgcactaat
     1141 gttccggcgt tatttcttga tgtctctgac cagacaccca tcaacagtat tattttctcc
     1201 catgaggacg gtacgcgact gggcgtggag catctggtcg cattgggtca ccagcaaatc
     1261 gcgctgttag cgggcccatt aagttctgtc tcggcgcgtc tgcgtctggc tggctggcat
     1321 aaatatctca ctcgcaatca aattcagccg atagcggaac gggaaggcga ctggagtgcc
     1381 atgtccggtt ttcaacaaac catgcaaatg ctgaatgagg gcatcgttcc cactgcgatg
     1441 ctggttgcca acgatcagat ggcgctgggc gcaatgcgcg ccattaccga gtccgggctg
     1501 cgcgttggtg cggatatctc ggtagtggga tacgacgata ccgaagatag ctcatgttat
     1561 atcccgccgt taaccaccat caaacaggat tttcgcctgc tggggcaaac cagcgtggac
     1621 cgcttgctgc aactctctca gggccaggcg gtgaagggca atcagctgtt gccagtctca
     1681 ctggtgaaaa gaaaaaccac cctggcgccc aatacgcaaa ccgcctctcc ccgcgcgttg
     1741 gccgattcat taatgcagct ggcacgacag gtttcccgac tggaaagcgg gcagtgactc
     1801 atgaccaaaa tcccttaacg tgagttacgc gcgcgtcgtt ccactgagcg tcagaccccg
     1861 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     1921 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     1981 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt
     2041 agccgtagtt agcccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     2101 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     2161 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     2221 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     2281 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     2341 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     2401 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     2461 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     2521 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
     2581 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg
     2641 aggaagcgga aggcgagagt agggaactgc caggcatcaa actaagcaga aggcccctga
     2701 cggatggcct ttttgcgttt ctacaaactc tttctgtgtt gtaaaacgac ggccagtctt
     2761 aagctcgggc cccctgggcg gttctgataa cgagtaatcg ttaatccgca aataacgtaa
     2821 aaacccgctt cggcgggttt ttttatgggg ggagtttagg gaaagagcat ttgtcagaat
     2881 atttaagggc gcctgtcact ttgcttgata tatgagaatt atttaacctt ataaatgaga
     2941 aaaaagcaac gcactttaaa taagatacgt tgctttttcg attgatgaac acctataatt
     3001 aaactattca tctattattt atgatttttt gtatatacaa tatttctagt ttgttaaaga
     3061 gaattaagaa aataaatctc gaaaataata aagggaaaat cagtttttga tatcaaaatt
     3121 atacatgtca acgataatac aaaatataat acaaactata agatgttatc agtatttatt
     3181 atgcatttag aataaatttt gtgtcgccct tccgcgaaat taatacgact cactataggg
     3241 gaattgtgag cggataacaa ttcccctcta gaaataattt tgtttaactt ttgaaggaga
     3301 tataccatgg gcagcgacaa gatccaccat caccaccatc acgaaaatct gtactttcaa
     3361 ggtcatatga gcacctctag cattccgtct tataatcaga tgaacggcaa ctccaataat
     3421 ggtatgccgc agcagaccac cgcagcaaat cagcaggcat ttcctcagca gcagcaaccg
     3481 accaccaccg gtaatgcaag ccagcaacag caacaggcag ccgcaaccgc agcagcggca
     3541 gcagttcagc agccgtataa ctatatgttt tatcagcagc aaggtcagcc tggtcagcag
     3601 accggtcaga ccgcaggtca gcagcaacaa cagcaacagc agcagcagca atatgattat
     3661 aacacctata accgctatca gtatccggca gcaaccagcc agggtaatta ctatcagcag
     3721 acaattccga atcagctgag ccagccgcag cctcagcatt ataacggtag caatcgtaat
     3781 tataccagcg caccgagtgg tgcaccgatt ccgagcaata gcaccagcgg tccgagccag
     3841 cagcctccgc tgcctggaca gcaggccgtt ccgattccgc ctcatgttag caccatgcag
     3901 cagccgacat caggtaatgg taatagcatt agcgcgacca gcggtggtag caatgttagt
     3961 ggtgcaagcg gtgcaggtag caccaccagt ccggttaata ccaaagcagc gaccgcagcc
     4021 ggtattccgc agggcaacta ttatcagacc tataatcagc aacagtaccc gcagcagtat
     4081 ggtcagtata atgcaccggg taaaaatcag aatacaccgg catcacagcc tggtagtacc
     4141 accaatgatc agtatctgca gcagcagcaa cagcaacaac aacaaatgta tggttatcag
     4201 agcaactact atcaaggtgg tgcagccaat agcagctatt atccgaatta ttaccaacaa
     4261 cagcagccga attatgcaag cagctatccg tatcaacagc agcaacaaaa acagcagcaa
     4321 caacaaccga atcagcagca acagagtgat cagcagcaga cctcaacccc gagcggtggt
     4381 gccggtacac gtagcgttca tcagagtccg caggttcaga gcctgaccca gggtagcgtg
     4441 catccgagcc cacagcagca tcaggcaaat cagagcgcaa gcaccgttgc aaaagaagag
     4501 aaataactcg agccccctag cataacccct tggggcctct aaacgggtct tgaggggttt
     4561 tttgcccctg agacgcgtca atcgagttcg tacctaaggg cgacaccccc taattagccc
     4621 gggcgaaagg cccagtcttt cgactgagcc tttcgtttta tttgatgcct ggcagttccc
     4681 tactctcgca tggggagtcc ccacactacc atcggcgcta cggcgtttca cttctgagtt
     4741 cggcatgggg tcaggtggga ccaccgcgct actgccgcca ggcaaacaag gggtgttatg
     4801 agccatattc aggtataaat gggctcgcga taatgttcag aattggttaa ttggttgtaa
     4861 cactgacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac
     4921 aataaccctg ataaatgctt caataatatt gaaaaaggaa gaatatgagc catattcaac
     4981 gggaaacgtc gaggccgcga ttaaattcca acatggatgc tgatttatat gggtataaat
     5041 gggctcgcga taatgtcggg caatcaggtg cgacaatcta tcgcttgtat gggaagcccg
     5101 atgcgccaga gttgtttctg aaacatggca aaggtagcgt tg
//
