LOCUS       sequence-394538-        7611 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial protein expression of Dcp1 82-129.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7611)
  AUTHORS   Currie SL, Xing W, Muhlrad D, Decker CJ, Parker R, Rosen MK
  TITLE     Quantitative reconstitution of yeast RNA processing bodies.
  JOURNAL   Proc Natl Acad Sci U S A. 2023 Apr 4;120(14):e2214064120. doi: 
            10.1073/pnas.2214064120. Epub 2023 Mar 27.
  PUBMED    36972455
REFERENCE   2  (bases 1 to 7611)
  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: "Proc Natl 
            Acad Sci U S A."; date: "2023-04-4"; volume: "120(14):e2214064120. 
            doi"; pages: " 10.1073/pnas.2214064120. Epub 2023 Mar 27"
COMMENT     Sequence Label: Dcp1 82-129-EGFP
FEATURES             Location/Qualifiers
     source          1..7611
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(123..128,129..144,145..151)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    159..175
                     /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     164..186
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             245..1345
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="MKTEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     regulatory      1202..1211
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     1319..1342
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1394..1405
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             1418..1438
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             join(1616..1618,1619..1621,1622..2332)
                     /codon_start=1
                     /product="brightly fluorescent monomeric variant of GFP"
                     /label=mEmerald
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHKVYITADKQKNGIK
                     VNFKTRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1661..1682)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1922..1941)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2269..2290
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2345..2365
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             2366..2383
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2396..2413)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2396..2412)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2405..2427)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(2591..2608)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      2761..2847
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      2939..2966
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        2985..3076
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3077..3145,3146..3937)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3295..3314)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(3979..4492)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4174..4195)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4386..4405
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     rep_origin      4603..5191
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5092..5111
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5345..5362
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(5468..5487)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5628..5650
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5644..5835)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     promoter        6331..6408
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             6409..7491
                     /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(6428..6447)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc gcactcccgt
       61 tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc tgaaatgagc
      121 tgttgacaat taatcatcgg ctcgtataat gtgtggaatt gtgagcggat aacaatttca
      181 cacaggaaac agccagtccg tttaggtgtt ttcacgagca cttcaccaac aaggaccata
      241 gattatgaaa actgaagaag gtaaactggt aatctggatt aacggcgata aaggctataa
      301 cggtctcgct gaagtcggta agaaattcga gaaagatacc ggaattaaag tcaccgttga
      361 gcatccggat aaactggaag agaaattccc acaggttgcg gcaactggcg atggccctga
      421 cattatcttc tgggcacacg accgctttgg tggctacgct caatctggcc tgttggctga
      481 aatcaccccg gacaaagcgt tccaggacaa gctgtatccg tttacctggg atgccgtacg
      541 ttacaacggc aagctgattg cttacccgat cgctgttgaa gcgttatcgc tgatttataa
      601 caaagatctg ctgccgaacc cgccaaaaac ctgggaagag atcccggcgc tggataaaga
      661 actgaaagcg aaaggtaaga gcgcgctgat gttcaacctg caagaaccgt acttcacctg
      721 gccgctgatt gctgctgacg ggggttatgc gttcaagtat gaaaacggca agtacgacat
      781 taaagacgtg ggcgtggata acgctggcgc gaaagcgggt ctgaccttcc tggttgacct
      841 gattaaaaac aaacacatga atgcagacac cgattactcc atcgcagaag ctgcctttaa
      901 taaaggcgaa acagcgatga ccatcaacgg cccgtgggca tggtccaaca tcgacaccag
      961 caaagtgaat tatggtgtaa cggtactgcc gaccttcaag ggtcaaccat ccaaaccgtt
     1021 cgttggcgtg ctgagcgcag gtattaacgc cgccagtccg aacaaagagc tggcaaaaga
     1081 gttcctcgaa aactatctgc tgactgatga aggtctggaa gcggttaata aagacaaacc
     1141 gctgggtgcc gtagcgctga agtcttacga ggaagagttg gcgaaagatc cacgtattgc
     1201 cgccaccatg gaaaacgccc agaaaggtga aatcatgccg aacatcccgc agatgtccgc
     1261 tttctggtat gccgtgcgta ctgcggtgat caacgccgcc agcggtcgtc agactgtcga
     1321 tgaagccctg aaagacgcgc agactaattc gagctcgaac aacaacaaca ataacaataa
     1381 caacaacctc gggatcgagg gaaggatttc agaatttgaa aacctgtatt ttcagggcca
     1441 tatgtcccca caagaagtag atatttttga ttcacaaaat ggtagtaata acattcaagt
     1501 aaacaatggt tctgacaata gcaacaggaa tagcagtgga aatgggaact cttataaaag
     1561 taatgattcg ctaacatata attgcggcgg atccgggtct ggttcaggaa tggccatggt
     1621 gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga
     1681 cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa
     1741 gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt
     1801 gaccaccttg acctacggcg tgcagtgctt cgcccgctac cccgaccaca tgaagcagca
     1861 cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa
     1921 ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa
     1981 ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct
     2041 ggagtacaac tacaacagcc acaaggtcta tatcaccgcc gacaagcaga agaacggcat
     2101 caaggtgaac ttcaagaccc gccacaacat cgaggacggc agcgtgcagc tcgccgacca
     2161 ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct
     2221 gagcacccag tccaagctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct
     2281 ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca agggaagcgg
     2341 ccgcgaaaac ctgtattttc agggccatca tcatcatcat cactgaaagc ttggcactgg
     2401 ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt tacccaactt aatcgccttg
     2461 cagcacatcc ccctttcgcc agctggcgta atagcgaaga ggcccgcacc gatcgccctt
     2521 cccaacagtt gcgcagcctg aatggcgaat ggcagcttgg ctgttttggc ggatgagata
     2581 agattttcag cctgatacag attaaatcag aacgcagaag cggtctgata aaacagaatt
     2641 tgcctggcgg cagtagcgcg gtggtcccac ctgaccccat gccgaactca gaagtgaaac
     2701 gccgtagcgc cgatggtagt gtggggtctc cccatgcgag agtagggaac tgccaggcat
     2761 caaataaaac gaaaggctca gtcgaaagac tgggcctttc gttttatctg ttgtttgtcg
     2821 gtgaacgctc tcctgagtag gacaaatccg ccgggagcgg atttgaacgt tgcgaagcaa
     2881 cggcccggag ggtggcgggc aggacgcccg ccataaactg ccaggcatca aattaagcag
     2941 aaggccatcc tgacggatgg cctttttgcg tttctacaaa ctcttttgtt tatttttcta
     3001 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata
     3061 ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
     3121 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
     3181 agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
     3241 tgagagtttt cgccccgaag aacgtttccc aatgatgagc acttttaaag ttctgctatg
     3301 tggcgcggta ttatcccgtg ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
     3361 ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat
     3421 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
     3481 acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga
     3541 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
     3601 gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga
     3661 actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc
     3721 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
     3781 cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg
     3841 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
     3901 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata
     3961 tatactttag attgatttac cccggttgat aatcagaaaa gccccaaaaa caggaagatt
     4021 gtataagcaa atatttaaat tgtaaacgtt aatattttgt taaaattcgc gttaaatttt
     4081 tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca
     4141 aaagaatagc ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta
     4201 aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta
     4261 cgtgaaccat cacccaaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg
     4321 aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga
     4381 aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg
     4441 ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtaaaaggat
     4501 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt
     4561 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     4621 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     4681 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     4741 aaatactgtc cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     4801 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     4861 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     4921 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     4981 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     5041 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     5101 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     5161 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     5221 cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt
     5281 ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga
     5341 gcgcagcgag tcagtgagcg aggaagcgga agagcgcctg atgcggtatt ttctccttac
     5401 gcatctgtgc ggtatttcac accgcatgtg gtgcactctc agtacaccgc atgtggtgca
     5461 ctctcagtac aatctgctct gatgccgcat agttaagcca gtatacactc cgctatcgct
     5521 acgtgactgg gtcatggctg cgccccgaca cccgccaaca cccgctgacg cgccctgacg
     5581 ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat
     5641 gtgtcagagg ttttcaccgt catcaccgaa acgcgcgagg cagctgcggt aaagctcatc
     5701 agcgtggtcg tgcagcgatt cacagatgtc tgcctgttca tccgcgtcca gctcgttgag
     5761 tttctccaga agcgttaatg tctggcttct gataaagcgg gccatgttaa gggcggtttt
     5821 ttcctgtttg gtcactgatg cctccgtgta agggggattt ctgttcatgg gggtaatgat
     5881 accgatgaaa cgagagagga tgctcacgat acgggttact gatgatgaac atgcccggtt
     5941 actggaacgt tgtgagggta aacaactggc ggtatggatg cggcgggacc agagaaaaat
     6001 cactcagggt caatgccagc gcttcgttaa tacagatgta ggtgttccac agggtagcca
     6061 gcagcatcct gcgatgcaga tccggaacat aatggtgcag ggcgctgact tccgcgtttc
     6121 cagactttac gaaacacgga aaccgaagac cattcatgtt gttgctcagg tcgcagacgt
     6181 tttgcagcag cagtcgcttc acgttcgctc gcgtatcggt gattcattct gctaaccagt
     6241 aaggcaaccc cgccagccta gccgggtcct caacgacagg agcacgatca tgcgcacccg
     6301 tggccaggac ccaacgctgc ccgaaattcc gacaccatcg aatggtgcaa aacctttcgc
     6361 ggtatggcat gatagcgccc ggaagagagt caattcaggg tggtgaatgt gaaaccagta
     6421 acgttatacg atgtcgcaga gtatgccggt gtctcttatc agaccgtttc ccgcgtggtg
     6481 aaccaggcca gccacgtttc tgcgaaaacg cgggaaaaag tggaagcggc gatggcggag
     6541 ctgaattaca ttcccaaccg cgtggcacaa caactggcgg gcaaacagtc gttgctgatt
     6601 ggcgttgcca cctccagtct ggccctgcac gcgccgtcgc aaattgtcgc ggcgattaaa
     6661 tctcgcgccg atcaactggg tgccagcgtg gtggtgtcga tggtagaacg aagcggcgtc
     6721 gaagcctgta aagcggcggt gcacaatctt ctcgcgcaac gcgtcagtgg gctgatcatt
     6781 aactatccgc tggatgacca ggatgccatt gctgtggaag ctgcctgcac taatgttccg
     6841 gcgttatttc ttgatgtctc tgaccagaca cccatcaaca gtattatttt ctcccatgaa
     6901 gacggtacgc gactgggcgt ggagcatctg gtcgcattgg gtcaccagca aatcgcgctg
     6961 ttagcgggcc cattaagttc tgtctcggcg cgtctgcgtc tggctggctg gcataaatat
     7021 ctcactcgca atcaaattca gccgatagcg gaacgggaag gcgactggag tgccatgtcc
     7081 ggttttcaac aaaccatgca aatgctgaat gagggcatcg ttcccactgc gatgctggtt
     7141 gccaacgatc agatggcgct gggcgcaatg cgcgccatta ccgagtccgg gctgcgcgtt
     7201 ggtgcggata tctcggtagt gggatacgac gataccgaag acagctcatg ttatatcccg
     7261 ccgttaacca ccatcaaaca ggattttcgc ctgctggggc aaaccagcgt ggaccgcttg
     7321 ctgcaactct ctcagggcca ggcggtgaag ggcaatcagc tgttgcccgt ctcactggtg
     7381 aaaagaaaaa ccaccctggc gcccaatacg caaaccgcct ctccccgcgc gttggccgat
     7441 tcattaatgc agctggcacg acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc
     7501 aattaatgta agttagctca ctcattaggc acaattctca tgtttgacag cttatcatcg
     7561 actgcacggt gcaccaatgc ttctggcgtc aggcagccat cggaagctgt g
//
