LOCUS       sequence-216430-        8578 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For bacterial expression of MBP fusion of full-length Drosophila 
            Tral with Phos-mimic mutations.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8578)
  AUTHORS   Hara M, Lourido S, Petrova B, Lou HJ, Von Stetina JR, Kashevsky H, 
            Turk BE, Orr-Weaver TL
  TITLE     Identification of PNG kinase substrates uncovers interactions with 
            the translational repressor TRAL in the oocyte-to-embryo transition.
  JOURNAL   Elife. 2018 Feb 26;7. pii: 33150. doi: 10.7554/eLife.33150.
  PUBMED    29480805
REFERENCE   2  (bases 1 to 8578)
  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: "Elife."; 
            date: "2018-02-26"; volume: "7. pii: 33150. doi"; pages: " 
            10.7554/eLife.33150"
COMMENT     Sequence Label: pMAL c2x Tral FL-15D
FEATURES             Location/Qualifiers
     source          1..8578
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        3..80
                     /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             81..1163
                     /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(100..119)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1406..1411,1412..1427,1428..1434)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1442..1458
                     /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     1447..1469
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1528..2628
                     /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="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     primer_bind     2602..2625
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             2677..2688
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(4669..4686)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4669..4685)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4678..4700)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(4864..4881)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      5034..5120
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      5212..5239
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        5258..5349
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5350..5418,5419..6210)
                     /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(5568..5587)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(6252..6765)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6447..6468)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6659..6678
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     rep_origin      6876..7464
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7365..7384
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7618..7635
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    7650..7792
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(7718..7737)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7878..7900
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(7894..8085)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 ccgacaccat cgaatggtgc aaaacctttc gcggtatggc atgatagcgc ccggaagaga
       61 gtcaattcag ggtggtgaat gtgaaaccag taacgttata cgatgtcgca gagtatgccg
      121 gtgtctctta tcagaccgtt tcccgcgtgg tgaaccaggc cagccacgtt tctgcgaaaa
      181 cgcgggaaaa agtggaagcg gcgatggcgg agctgaatta cattcccaac cgcgtggcac
      241 aacaactggc gggcaaacag tcgttgctga ttggcgttgc cacctccagt ctggccctgc
      301 acgcgccgtc gcaaattgtc gcggcgatta aatctcgcgc cgatcaactg ggtgccagcg
      361 tggtggtgtc gatggtagaa cgaagcggcg tcgaagcctg taaagcggcg gtgcacaatc
      421 ttctcgcgca acgcgtcagt gggctgatca ttaactatcc gctggatgac caggatgcca
      481 ttgctgtgga agctgcctgc actaatgttc cggcgttatt tcttgatgtc tctgaccaga
      541 cacccatcaa cagtattatt ttctcccatg aagacggtac gcgactgggc gtggagcatc
      601 tggtcgcatt gggtcaccag caaatcgcgc tgttagcggg cccattaagt tctgtctcgg
      661 cgcgtctgcg tctggctggc tggcataaat atctcactcg caatcaaatt cagccgatag
      721 cggaacggga aggcgactgg agtgccatgt ccggttttca acaaaccatg caaatgctga
      781 atgagggcat cgttcccact gcgatgctgg ttgccaacga tcagatggcg ctgggcgcaa
      841 tgcgcgccat taccgagtcc gggctgcgcg ttggtgcgga tatctcagta gtgggatacg
      901 acgataccga agacagctca tgttatatcc cgccgttaac caccatcaaa caggattttc
      961 gcctgctggg gcaaaccagc gtggaccgct tgctgcaact ctctcagggc caggcggtga
     1021 agggcaatca gctgttgccc gtctcactgg tgaaaagaaa aaccaccctg gcgcccaata
     1081 cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt
     1141 cccgactgga aagcgggcag tgagcgcaac gcaattaatg taagttagct cactcattag
     1201 gcacaattct catgtttgac agcttatcat cgactgcacg gtgcaccaat gcttctggcg
     1261 tcaggcagcc atcggaagct gtggtatggc tgtgcaggtc gtaaatcact gcataattcg
     1321 tgtcgctcaa ggcgcactcc cgttctggat aatgtttttt gcgccgacat cataacggtt
     1381 ctggcaaata ttctgaaatg agctgttgac aattaatcat cggctcgtat aatgtgtgga
     1441 attgtgagcg gataacaatt tcacacagga aacagccagt ccgtttaggt gttttcacga
     1501 gcacttcacc aacaaggacc atagcatatg aaaatcgaag aaggtaaact ggtaatctgg
     1561 attaacggcg ataaaggcta taacggtctc gctgaagtcg gtaagaaatt cgagaaagat
     1621 accggaatta aagtcaccgt tgagcatccg gataaactgg aagagaaatt cccacaggtt
     1681 gcggcaactg gcgatggccc tgacattatc ttctgggcac acgaccgctt tggtggctac
     1741 gctcaatctg gcctgttggc tgaaatcacc ccggacaaag cgttccagga caagctgtat
     1801 ccgtttacct gggatgccgt acgttacaac ggcaagctga ttgcttaccc gatcgctgtt
     1861 gaagcgttat cgctgattta taacaaagat ctgctgccga acccgccaaa aacctgggaa
     1921 gagatcccgg cgctggataa agaactgaaa gcgaaaggta agagcgcgct gatgttcaac
     1981 ctgcaagaac cgtacttcac ctggccgctg attgctgctg acgggggtta tgcgttcaag
     2041 tatgaaaacg gcaagtacga cattaaagac gtgggcgtgg ataacgctgg cgcgaaagcg
     2101 ggtctgacct tcctggttga cctgattaaa aacaaacaca tgaatgcaga caccgattac
     2161 tccatcgcag aagctgcctt taataaaggc gaaacagcga tgaccatcaa cggcccgtgg
     2221 gcatggtcca acatcgacac cagcaaagtg aattatggtg taacggtact gccgaccttc
     2281 aagggtcaac catccaaacc gttcgttggc gtgctgagcg caggtattaa cgccgccagt
     2341 ccgaacaaag agctggcaaa agagttcctc gaaaactatc tgctgactga tgaaggtctg
     2401 gaagcggtta ataaagacaa accgctgggt gccgtagcgc tgaagtctta cgaggaagag
     2461 ttggcgaaag atccacgtat tgccgccact atggaaaacg cccagaaagg tgaaatcatg
     2521 ccgaacatcc cgcagatgtc cgctttctgg tatgccgtgc gtactgcggt gatcaacgcc
     2581 gccagcggtc gtcagactgt cgatgaagcc ctgaaagacg cgcagactaa ttcgagctcg
     2641 aacaacaaca acaataacaa taacaacaac ctcgggatcg agggaaggat ttcagaattc
     2701 atgagcgggg gattaccgga gttgggctcc aaaatcagcc tcatttccaa ggcggacatt
     2761 cgatacgagg gccgcctgta cacggtggat ccgcaggagt gcaccattgc gctctcgagt
     2821 gtgcgatcct tcggtacaga agatcgggac acccagttcc agatagcgcc gcaatcgcag
     2881 atctacgact acatcctatt ccgtggctcg gacatcaagg acattcgcgt tgtcaacaac
     2941 cacacgctgc cgcaccacaa tgacccggcg atcatgcagg cgcagctcca gaacggaccg
     3001 ccccagatgc cgcagcactt ccccatgccc agcgggatga gcggtccgcc gcagcagcaa
     3061 caggtgccgt cgcagcaacc gccccaaatg ccaagcggtg gaggcagtgg tggtgctggt
     3121 ggagcaggag ctcctggcgg cggcggaccc ggctacggca atggcaatcc gttcggcaac
     3181 ctcggcggtc ccaatctggc caacatggtg ggcaatgcgg gcggatcgct tgcacccggt
     3241 tccggtgctc cgggcagcgg acccttcatg cacatcggtg gcaatcagca gcaaccgaag
     3301 ccccagcagc agaaacagcc aaacatgctg gccggtgcct cgcgatcgac gacgcccatt
     3361 agcctgattg tgagtcccac ggcggagctg actcagcagc agcagctgca ccagcaacag
     3421 aacgctggcc agggtggcaa cggacgcgat gcaggacata agcgccagaa ccaccagcag
     3481 caacagaatc agcagcatca gcaacgtggc ggacccagcc acaacaatat gcagcagcag
     3541 cagcaacgag gcggcggcag tggaacggac ttctataacc agcagcggga tcgtcgggac
     3601 tccggacgtc aaatggacaa caactatagc aacaactaca acaacaataa taataatcag
     3661 cgcaatcgcg gcggcggcaa cggaatgcaa cagcagcagc gaggaggaaa cggcggcagc
     3721 ggcggcggcg gtggaaacgg aggtggaaac aacccggcct ggaacatgca tcgcggcaac
     3781 cagaactcga acaacatgat gaacatgcgc aaccgcggca tgggatcccg cggccccatg
     3841 cgacccaatc agggctatcg tccgcagtcg gccaataatc agaacaagcc gcggaacaag
     3901 atcaagttcg agggcgactt cgatttcgag caggcaaaca acaagttcga ggaactgcgc
     3961 tcccaactgg ccaagctcaa ggtggccgag gatggtgcac ccaagccagc cgacaatgca
     4021 gatgccgccg atgcagatgc agacaatgag caggtgggtg agaaggttga aggcgttcac
     4081 acactgaatg gcgagaccga caagaaggat gattctggca acgagaccgg cgctggagag
     4141 cacgagcctg aggaggatga tgttgctgtg tgctacgaca agaccaaatc gttcttcgac
     4201 aacatctcgt gcgaggctgc ccaggatcgc agcaagaaca agaagaacga ttggcgccag
     4261 gagcgcaagt tgaacgatga ggacttcgga gtggacgacg atcgacgtgg cagttaccgc
     4321 ggacgcaacc aatactacaa caataatggc aacggcggca ttaactcggg atatggagga
     4381 gcgccaggct acaacaggaa caattatcgc atgggtggcg gcggcggaaa cttccgaaac
     4441 aggagcaaca atcgcaacaa cggcggcggt cgtggcggaa acggaatgcc aaacatcacc
     4501 aatggcaaca cggctgctgc gctgaaggcg gccaacaatg ctgctggcca cggatccaat
     4561 gccacggact ccagtgcacc aaatgccgat gacgcggatg acaagtcgac gtccctcttg
     4621 ccagagcagg atcaacaggt ggcggcagtt tcacaataga agcttggcac tggccgtcgt
     4681 tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca
     4741 tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca
     4801 gttgcgcagc ctgaatggcg aatggcagct tggctgtttt ggcggatgag ataagatttt
     4861 cagcctgata cagattaaat cagaacgcag aagcggtctg ataaaacaga atttgcctgg
     4921 cggcagtagc gcggtggtcc cacctgaccc catgccgaac tcagaagtga aacgccgtag
     4981 cgccgatggt agtgtggggt ctccccatgc gagagtaggg aactgccagg catcaaataa
     5041 aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat ctgttgtttg tcggtgaacg
     5101 ctctcctgag taggacaaat ccgccgggag cggatttgaa cgttgcgaag caacggcccg
     5161 gagggtggcg ggcaggacgc ccgccataaa ctgccaggca tcaaattaag cagaaggcca
     5221 tcctgacgga tggccttttt gcgtttctac aaactctttt gtttattttt ctaaatacat
     5281 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
     5341 aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt
     5401 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag
     5461 ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt
     5521 tttcgccccg aagaacgttt cccaatgatg agcactttta aagttctgct atgtggcgcg
     5581 gtattatccc gtgttgacgc cgggcaagag caactcggtc gccgcataca ctattctcag
     5641 aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta
     5701 agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg
     5761 acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta
     5821 actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac
     5881 accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt
     5941 actctagctt cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca
     6001 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
     6061 cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta
     6121 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag
     6181 ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt
     6241 tagattgatt taccccggtt gataatcaga aaagccccaa aaacaggaag attgtataag
     6301 caaatattta aattgtaaac gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa
     6361 tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat
     6421 agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg
     6481 tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac
     6541 catcacccaa atcaagtttt ttggggtcga ggtgccgtaa agcactaaat cggaacccta
     6601 aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg agaaaggaag
     6661 ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg
     6721 taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtaaaag gatctaggtg
     6781 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     6841 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     6901 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     6961 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     7021 gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     7081 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     7141 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     7201 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     7261 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     7321 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     7381 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     7441 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     7501 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     7561 cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc
     7621 gagtcagtga gcgaggaagc ggaagagcgc ctgatgcggt attttctcct tacgcatctg
     7681 tgcggtattt cacaccgcat atatggtgca ctctcagtac aatctgctct gatgccgcat
     7741 agttaagcca gtatacactc cgctatcgct acgtgactgg gtcatggctg cgccccgaca
     7801 cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag
     7861 acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa
     7921 acgcgcgagg cagctgcggt aaagctcatc agcgtggtcg tgcagcgatt cacagatgtc
     7981 tgcctgttca tccgcgtcca gctcgttgag tttctccaga agcgttaatg tctggcttct
     8041 gataaagcgg gccatgttaa gggcggtttt ttcctgtttg gtcactgatg cctccgtgta
     8101 agggggattt ctgttcatgg gggtaatgat accgatgaaa cgagagagga tgctcacgat
     8161 acgggttact gatgatgaac atgcccggtt actggaacgt tgtgagggta aacaactggc
     8221 ggtatggatg cggcgggacc agagaaaaat cactcagggt caatgccagc gcttcgttaa
     8281 tacagatgta ggtgttccac agggtagcca gcagcatcct gcgatgcaga tccggaacat
     8341 aatggtgcag ggcgctgact tccgcgtttc cagactttac gaaacacgga aaccgaagac
     8401 cattcatgtt gttgctcagg tcgcagacgt tttgcagcag cagtcgcttc acgttcgctc
     8461 gcgtatcggt gattcattct gctaaccagt aaggcaaccc cgccagccta gccgggtcct
     8521 caacgacagg agcacgatca tgcgcacccg tggccaggac ccaacgctgc ccgaaatt
//
