LOCUS       sequence-176274-        7741 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Recombinant expression of PTBP1-RRM1 in E.coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7741)
  AUTHORS   Dorn G, Leitner A, Boudet J, Campagne S, von Schroetter C, Moursy A,
            Aebersold R, Allain FH
  TITLE     Structural modeling of protein-RNA complexes using crosslinking of 
            segmentally isotope-labeled RNA and MS/MS.
  JOURNAL   Nat Methods. 2017 Mar 27. doi: 10.1038/nmeth.4235.
  PUBMED    28346450
REFERENCE   2  (bases 1 to 7741)
  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 
            Methods. 2017 Mar 27. doi: 10.1038/nmeth.4235."
COMMENT     Sequence Label: pTYB11-PTBP1-RRM1
FEATURES             Location/Qualifiers
     source          1..7741
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      36..624
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     525..544
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     778..795
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(817..836)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     977..999
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(993..1184)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(1741..2823)
                     /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     2785..2804
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(2824..2901)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     3107..3126
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     3210..3229
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3210..3228
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    3229..3253
                     /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             3268..3290
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             3343..4161
                     /codon_start=1
                     /gene="Saccharomyces cerevisiae VMA1"
                     /product="5' region of the intein from the yeast Vma1 
                     subunit of the vacuolar ATPase (Chong et al., 1998)"
                     /label=Sce VMA intein 5' region
                     /translation="CFAKGTNVLMADGSIECIENIEVGNKVMGKDGRPREVIKLPRGRE
                     TMYSVVQKSQHRAHKSDSSREVPELLKFTCNATHELVVRTPRSVRRLSRTIKGVEYFEV
                     ITFEMGQKKAPDGRIVELVKEVSKSYPISEGPERANELVESYRKASNKAYFEWTIEARD
                     LSLLGSHVRKATYQTYAPILYENDHFFDYMQKSKFHLTIEGPKVLAYLLGLWIGDGLSD
                     RATFSVDSRDTSLMERVTEYAEKLNLCAEYKDRKEPQVAKTVNLYSKVVRG"
     CDS             4168..4320
                     /codon_start=1
                     /gene="Bacillus circulans chiA"
                     /product="chitin binding domain from chitinase A1 (Watanabe
                     et al., 1994)"
                     /label=CBD
                     /translation="TNPGVSAWQVNTAYTAGQLVTYNGKTYKCLQPHTSLAGWEPSNVP
                     ALWQLQ"
     CDS             4324..4872
                     /codon_start=1
                     /gene="Saccharomyces cerevisiae VMA1"
                     /product="modified 3' region of the intein from the yeast 
                     Vma1 subunit of the vacuolar ATPase (Chong et al., 1998)"
                     /label=Sce VMA intein 3' region
                     /note="C-terminal cleavage of this intein can be induced by
                     thiols."
                     /translation="GHGGIRNNLNTENPLWDAIVGLGFLKDGVKNIPSFLSTDNIGTRE
                     TFLAGLIDSDGYVTDEHGIKATIKTIHTSVRDGLVSLARSLGLVVSVNAEPAKVDMNVT
                     KHKISYAIYMSGGDVLLNVLSKCAGSKKFRPAPAAAFARECRGFYFELQELKEDDYYGI
                     TLSDDSDHQFLLGSQVVVQN"
     primer_bind     complement(5321..5339)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      5335..5382
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(5570..5587)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5570..5586)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5579..5601)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(5765..5782)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      5935..6021
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      6113..6140
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        6159..6250
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6251..6319,6320..7111)
                     /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(6469..6488)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(7153..7666)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7348..7369)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7560..7579
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
       61 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
      121 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
      181 gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
      241 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
      301 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
      361 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
      421 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
      481 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
      541 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
      601 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
      661 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
      721 cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc
      781 gagtcagtga gcgaggaagc tatggtgcac tctcagtaca atctgctctg atgccgcata
      841 gttaagccag tatacactcc gctatcgcta cgtgactggg tcatggctgc gccccgacac
      901 ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga
      961 caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa
     1021 cgcgcgaggc agctgcggta aagctcatca gcgtggtcgt gcagcgattc acagatgtct
     1081 gcctgttcat ccgcgtccag ctcgttgagt ttctccagaa gcgttaatgt ctggcttctg
     1141 ataaagcggg ccatgttaag ggcggttttt tcctgtttgg tcactgatgc ctccgtgtaa
     1201 gggggatttc tgttcatggg ggtaatgata ccgatgaaac gagagaggat gctcacgata
     1261 cgggttactg atgatgaaca tgcccggtta ctggaacgtt gtgagggtaa acaactggcg
     1321 gtatggatgc ggcgggacca gagaaaaatc actcagggtc aatgccagcc gaacgccagc
     1381 aagacgtagc ccagcgcgtc ggccgccatg ccggcgataa tggcctgctt ctcgccgaaa
     1441 cgtttggtgg cgggaccagt gacgaaggct tgagcgaggg cgtgcaagat tccgaatacc
     1501 gcaagcgaca ggccgatcat cgtcgcgctc cagcgaaagc ggtcctcgcc gaaaatgacc
     1561 cagagcgctg ccggcacctg tcctacgagt tgcatgataa agaagacagt cataagtgcg
     1621 gcgacgatag tcatgccccg cgcccaccgg aaggagctga ctgggttgaa ggctctcaag
     1681 ggcatcggtc gagatcccgg tgcctaatga gtgagctaac ttacattaat tgcgttgcgc
     1741 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     1801 cgcgcgggga gaggcggttt gcgtattggg cgccagggtg gtttttcttt tcaccagtga
     1861 gacgggcaac agctgattgc ccttcaccgc ctggccctga gagagttgca gcaagcggtc
     1921 cacgctggtt tgccccagca ggcgaaaatc ctgtttgatg gtggttaacg gcgggatata
     1981 acatgagctg tcttcggtat cgtcgtatcc cactaccgag atatccgcac caacgcgcag
     2041 cccggactcg gtaatggcgc gcattgcgcc cagcgccatc tgatcgttgg caaccagcat
     2101 cgcagtggga acgatgccct cattcagcat ttgcatggtt tgttgaaaac cggacatggc
     2161 actccagtcg ccttcccgtt ccgctatcgg ctgaatttga ttgcgagtga gatatttatg
     2221 ccagccagcc agacgcagac gcgccgagac agaacttaat gggcccgcta acagcgcgat
     2281 ttgctggtga cccaatgcga ccagatgctc cacgcccagt cgcgtaccgt cttcatggga
     2341 gaaaataata ctgttgatgg gtgtctggtc agagacatca agaaataacg ccggaacatt
     2401 agtgcaggca gcttccacag caatggcatc ctggtcatcc agcggatagt taatgatcag
     2461 cccactgacg cgttgcgcga gaagattgtg caccgccgct ttacaggctt cgacgccgct
     2521 tcgttctacc atcgacacca ccacgctggc acccagttga tcggcgcgag atttaatcgc
     2581 cgcgacaatt tgcgacggcg cgtgcagggc cagactggag gtggcaacgc caatcagcaa
     2641 cgactgtttg cccgccagtt gttgtgccac gcggttggga atgtaattca gctccgccat
     2701 cgccgcttcc actttttccc gcgttttcgc agaaacgtgg ctggcctggt tcaccacgcg
     2761 ggaaacggtc tgataagaga caccggcata ctctgcgaca tcgtataacg ttactggttt
     2821 cacattcacc accctgaatt gactctcttc cgggcgctat catgccatac cgcgaaaggt
     2881 tttgcgccat tcgatggtgt ccgggatctc gacgctctcc cttatgcgac tcctgcatta
     2941 ggaagcagcc cagtagtagg ttgaggccgt tgagcaccgc cgccgcaagg aatggtgcat
     3001 gcaaggagat ggcgcccaac agtcccccgg ccacggggcc tgccaccata cccacgccga
     3061 aacaagcgct catgagcccg aagtggcgag cccgatcttc cccatcggtg atgtcggcga
     3121 tataggcgcc agcaaccgca cctgtggcgc cggtgatgcc ggccacgatg cgtccggcgt
     3181 agaggatcga gatctcgatc ccgcgaaatt aatacgactc actatagggg aattgtgagc
     3241 ggataacaat tcccctctag aaataatttt gtttaacttt aagaaggaga tatacttatg
     3301 aaaatcgaag aaggtaaact ggtaatcggt tccctggagg gttgctttgc caagggtacc
     3361 aatgttttaa tggcggatgg gtctattgaa tgtattgaaa acattgaggt tggtaataag
     3421 gtcatgggta aagatggcag acctcgtgag gtaattaaat tgcccagagg aagagaaact
     3481 atgtacagcg tcgtgcagaa aagtcagcac agagcccaca aaagtgactc aagtcgtgaa
     3541 gtgccagaat tactcaagtt tacgtgtaat gcgacccatg agttggttgt tagaacacct
     3601 cgtagtgtcc gccgtttgtc tcgtaccatt aagggtgtcg aatattttga agttattact
     3661 tttgagatgg gccaaaagaa agcccccgac ggtagaattg ttgagcttgt caaggaagtt
     3721 tcaaagagct acccaatatc tgaggggcct gagagagcca acgaattagt agaatcctat
     3781 agaaaggctt caaataaagc ttattttgag tggactattg aggccagaga tctttctctg
     3841 ttgggttccc atgttcgtaa agctacctac cagacttacg ctccaattct ttatgagaat
     3901 gaccactttt tcgactacat gcaaaaaagt aagtttcatc tcaccattga aggtccaaaa
     3961 gtacttgctt atttacttgg tttatggatt ggtgatggat tgtctgacag ggcaactttt
     4021 tcggttgatt ccagagatac ttctttgatg gaacgtgtta ctgaatatgc tgaaaagttg
     4081 aatttgtgcg ccgagtataa ggacagaaaa gaaccacaag ttgccaaaac tgttaatttg
     4141 tactctaaag ttgtcagagg tgctagcaca aatcctggtg tatccgcttg gcaggtcaac
     4201 acagcttata ctgcgggaca attggtcaca tataacggca agacgtataa atgtttgcag
     4261 ccccacacct ccttggcagg atgggaacca tccaacgttc ctgccttgtg gcagcttcaa
     4321 ggtggccatg gtggtattcg caataatctt aatactgaga atccattatg ggacgctatt
     4381 gttggcttag gattcttgaa ggacggtgtc aaaaatattc cttctttctt gtctacggac
     4441 aatatcggta ctcgtgaaac atttcttgct ggtctaattg attctgatgg ctatgttact
     4501 gatgagcatg gtattaaagc aacaataaag acaattcata cttctgtcag agatggtttg
     4561 gtttcccttg ctcgttcttt aggcttagta gtctcggtta acgcagaacc tgctaaggtt
     4621 gacatgaatg tcaccaaaca taaaattagt tatgctattt atatgtctgg tggagatgtt
     4681 ttgcttaacg ttctttcgaa gtgtgccggc tctaaaaaat tcaggcctgc tcccgccgct
     4741 gcttttgcac gtgagtgccg cggattttat ttcgagttac aagaattgaa ggaagacgat
     4801 tattatggga ttactttatc tgatgattct gatcatcagt ttttgcttgg atcccaggtt
     4861 gttgtacaga acggaaatga cagcaagaag ttcaaaggtg acagccgaag tgcaggcgtc
     4921 ccctctagag tgatccacat ccggaagctc cccatcgacg tcacggaggg ggaagtcatc
     4981 tccctggggc tgccctttgg gaaggtcacc aacctcctga tgctgaaggg gaaaaaccag
     5041 gccttcatcg agatgaacac ggaggaggct gccaacacca tggtgaacta ctacacctcg
     5101 gtgacccctg tgctgcgcgg ccagcccatc tacatccagt tctctaacca caaggagctg
     5161 aagaccgaca gctctcccaa ccaggcgcgg gcccaggcgg ccctgcaggc ggtgaactcg
     5221 gtccagtcgg ggaacctggc ctgactcgag cccgggtgac tgcaggaagg ggatccggct
     5281 gctaacaaag cccgaaagga agctgagttg gctgctgcca ccgctgagca ataactagca
     5341 taaccccttg gggcctctaa acgggtcttg aggggttttt tgctgaaagg aggaactata
     5401 tccggatatc ccgcaagagg cccggcagta ccggcataac caagcctatg cctacagcat
     5461 ccagggtgac ggtgccgagg atgacgatga gcgcattgtt agatttcata cacggtgcct
     5521 gactgcgtta gcaatttaac tgtgataaac taccgcatta aagcttggca ctggccgtcg
     5581 ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc cttgcagcac
     5641 atcccccttt cgccagctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac
     5701 agttgcgcag cctgaatggc gaatggcagc ttggctgttt tggcggatga gataagattt
     5761 tcagcctgat acagattaaa tcagaacgca gaagcggtct gataaaacag aatttgcctg
     5821 gcggcagtag cgcggtggtc ccacctgacc ccatgccgaa ctcagaagtg aaacgccgta
     5881 gcgccgatgg tagtgtgggg tctccccatg cgagagtagg gaactgccag gcatcaaata
     5941 aaacgaaagg ctcagtcgaa agactgggcc tttcgtttta tctgttgttt gtcggtgaac
     6001 gctctcctga gtaggacaaa tccgccggga gcggatttga acgttgcgaa gcaacggccc
     6061 ggagggtggc gggcaggacg cccgccataa actgccaggc atcaaattaa gcagaaggcc
     6121 atcctgacgg atggcctttt tgcgtttcta caaactcttt tgtttatttt tctaaataca
     6181 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa
     6241 aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt
     6301 ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca
     6361 gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag
     6421 ttttcgcccc gaagaacgtt tcccaatgat gagcactttt aaagttctgc tatgtggcgc
     6481 ggtattatcc cgtgttgacg ccgggcaaga gcaactcggt cgccgcatac actattctca
     6541 gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt
     6601 aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct
     6661 gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt
     6721 aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga
     6781 caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact
     6841 tactctagct tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc
     6901 acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga
     6961 gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt
     7021 agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga
     7081 gataggtgcc tcactgatta agcattggta actgtcagac caagtttact catatatact
     7141 ttagattgat ttaccccggt tgataatcag aaaagcccca aaaacaggaa gattgtataa
     7201 gcaaatattt aaattgtaaa cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa
     7261 atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa
     7321 tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac
     7381 gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa
     7441 ccatcaccca aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct
     7501 aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa
     7561 gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc
     7621 gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtaaaa ggatctaggt
     7681 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     7741 a
//
