LOCUS       sequence-176055-        5919 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Recombinant expression of PTBP1-RRM34 in E.coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5919)
  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 5919)
  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: pET28a-PTBP1-RRM34
FEATURES             Location/Qualifiers
     source          1..5919
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      18..606
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     507..526
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     760..777
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    792..931
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(857..876)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1017..1039
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(1033..1224)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(2033..3115)
                     /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     3077..3096
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3116..3193)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     3399..3418
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     3502..3521
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3502..3520
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    3521..3545
                     /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             3560..3582
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             3601..3618
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             3628..3645
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             4285..4302
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4355..4373)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      4369..4416
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      4453..4908
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4540..4559)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4750..4771
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(5000..5815)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(5000..5815)
                     /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="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     5723..5742
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5807..5826
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
ORIGIN
        1 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
       61 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
      121 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgtccttct agtgtagccg
      181 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
      241 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
      301 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
      361 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
      421 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
      481 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
      541 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
      601 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
      661 cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag
      721 tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa
      781 gcggaagagc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc
      841 aatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag tatacactcc
      901 gctatcgcta cgtgactggg tcatggctgc gccccgacac ccgccaacac ccgctgacgc
      961 gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg
     1021 gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc agctgcggta
     1081 aagctcatca gcgtggtcgt gaagcgattc acagatgtct gcctgttcat ccgcgtccag
     1141 ctcgttgagt ttctccagaa gcgttaatgt ctggcttctg ataaagcggg ccatgttaag
     1201 ggcggttttt tcctgtttgg tcactgatgc ctccgtgtaa gggggatttc tgttcatggg
     1261 ggtaatgata ccgatgaaac gagagaggat gctcacgata cgggttactg atgatgaaca
     1321 tgcccggtta ctggaacgtt gtgagggtaa acaactggcg gtatggatgc ggcgggacca
     1381 gagaaaaatc actcagggtc aatgccagcg cttcgttaat acagatgtag gtgttccaca
     1441 gggtagccag cagcatcctg cgatgcagat ccggaacata atggtgcagg gcgctgactt
     1501 ccgcgtttcc agactttacg aaacacggaa accgaagacc attcatgttg ttgctcaggt
     1561 cgcagacgtt ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg attcattctg
     1621 ctaaccagta aggcaacccc gccagcctag ccgggtcctc aacgacagga gcacgatcat
     1681 gcgcacccgt ggggccgcca tgccggcgat aatggcctgc ttctcgccga aacgtttggt
     1741 ggcgggacca gtgacgaagg cttgagcgag ggcgtgcaag attccgaata ccgcaagcga
     1801 caggccgatc atcgtcgcgc tccagcgaaa gcggtcctcg ccgaaaatga cccagagcgc
     1861 tgccggcacc tgtcctacga gttgcatgat aaagaagaca gtcataagtg cggcgacgat
     1921 agtcatgccc cgcgcccacc ggaaggagct gactgggttg aaggctctca agggcatcgg
     1981 tcgagatccc ggtgcctaat gagtgagcta acttacatta attgcgttgc gctcactgcc
     2041 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     2101 gagaggcggt ttgcgtattg ggcgccaggg tggtttttct tttcaccagt gagacgggca
     2161 acagctgatt gcccttcacc gcctggccct gagagagttg cagcaagcgg tccacgctgg
     2221 tttgccccag caggcgaaaa tcctgtttga tggtggttaa cggcgggata taacatgagc
     2281 tgtcttcggt atcgtcgtat cccactaccg agatatccgc accaacgcgc agcccggact
     2341 cggtaatggc gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc atcgcagtgg
     2401 gaacgatgcc ctcattcagc atttgcatgg tttgttgaaa accggacatg gcactccagt
     2461 cgccttcccg ttccgctatc ggctgaattt gattgcgagt gagatattta tgccagccag
     2521 ccagacgcag acgcgccgag acagaactta atgggcccgc taacagcgcg atttgctggt
     2581 gacccaatgc gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg gagaaaataa
     2641 tactgttgat gggtgtctgg tcagagacat caagaaataa cgccggaaca ttagtgcagg
     2701 cagcttccac agcaatggca tcctggtcat ccagcggata gttaatgatc agcccactga
     2761 cgcgttgcgc gagaagattg tgcaccgccg ctttacaggc ttcgacgccg cttcgttcta
     2821 ccatcgacac caccacgctg gcacccagtt gatcggcgcg agatttaatc gccgcgacaa
     2881 tttgcgacgg cgcgtgcagg gccagactgg aggtggcaac gccaatcagc aacgactgtt
     2941 tgcccgccag ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc atcgccgctt
     3001 ccactttttc ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg cgggaaacgg
     3061 tctgataaga gacaccggca tactctgcga catcgtataa cgttactggt ttcacattca
     3121 ccaccctgaa ttgactctct tccgggcgct atcatgccat accgcgaaag gttttgcgcc
     3181 attcgatggt gtccgggatc tcgacgctct cccttatgcg actcctgcat taggaagcag
     3241 cccagtagta ggttgaggcc gttgagcacc gccgccgcaa ggaatggtgc atgcaaggag
     3301 atggcgccca acagtccccc ggccacgggg cctgccacca tacccacgcc gaaacaagcg
     3361 ctcatgagcc cgaagtggcg agcccgatct tccccatcgg tgatgtcggc gatataggcg
     3421 ccagcaaccg cacctgtggc gccggtgatg ccggccacga tgcgtccggc gtagaggatc
     3481 gagatctcga tcccgcgaaa ttaatacgac tcactatagg ggaattgtga gcggataaca
     3541 attcccctct agaaataatt ttgtttaact ttaagaagga gatataccat gggcagcagc
     3601 catcatcatc atcatcacag cagcggcctg gtgccgcgcg gcagccatat gggtcggatc
     3661 gccatcccgg gcctggcggg ggcaggaaat tctgtattgc tggtcagcaa cctcaaccca
     3721 gagagagtca caccccaaag cctctttatt cttttcggcg tctacggtga cgtgcagcgc
     3781 gtgaagatcc tgttcaataa gaaggagaac gccctagtgc agatggcgga cggcaaccag
     3841 gcccagctgg ccatgagcca cctgaacggg cacaagctgc acgggaagcc gatccgcatc
     3901 acgctctcga agcaccagaa cgtgcagctg ccccgcgagg gccaggagga ccagggcctg
     3961 accaaggact acggcaactc acccctgcac cgcttcaaga agccgggctc caagaacttc
     4021 cagaacatat tcccgccctc ggccactctg cacctctcca acatcccgcc ctcagtctcc
     4081 gaggaggatc tcaaggtcct gttttccagc aatgggggcg tcgtcaaagg attcaagttc
     4141 ttccagaagg accgcaagat ggcactgatc cagatgggct ccgtggagga ggcggtccag
     4201 gccctcattg acctgcacaa ccacgacctc ggggagaacc accacctgcg ggtctccttc
     4261 tccaagtcca ccatctagct cgagcaccac caccaccacc actgagatcc ggctgctaac
     4321 aaagcccgaa aggaagctga gttggctgct gccaccgctg agcaataact agcataaccc
     4381 cttggggcct ctaaacgggt cttgaggggt tttttgctga aaggaggaac tatatccgga
     4441 ttggcgaatg ggacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
     4501 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt
     4561 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag
     4621 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt
     4681 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
     4741 tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt
     4801 cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt
     4861 aacaaaaatt taacgcgaat tttaacaaaa tattaacgct tacaatttag gtggcacttt
     4921 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta
     4981 tccgctcatg aattaattct tagaaaaact catcgagcat caaatgaaac tgcaatttat
     5041 tcatatcagg attatcaata ccatattttt gaaaaagccg tttctgtaat gaaggagaaa
     5101 actcaccgag gcagttccat aggatggcaa gatcctggta tcggtctgcg attccgactc
     5161 gtccaacatc aatacaacct attaatttcc cctcgtcaaa aataaggtta tcaagtgaga
     5221 aatcaccatg agtgacgact gaatccggtg agaatggcaa aagtttatgc atttctttcc
     5281 agacttgttc aacaggccag ccattacgct cgtcatcaaa atcactcgca tcaaccaaac
     5341 cgttattcat tcgtgattgc gcctgagcga gacgaaatac gcgatcgctg ttaaaaggac
     5401 aattacaaac aggaatcgaa tgcaaccggc gcaggaacac tgccagcgca tcaacaatat
     5461 tttcacctga atcaggatat tcttctaata cctggaatgc tgttttcccg gggatcgcag
     5521 tggtgagtaa ccatgcatca tcaggagtac ggataaaatg cttgatggtc ggaagaggca
     5581 taaattccgt cagccagttt agtctgacca tctcatctgt aacatcattg gcaacgctac
     5641 ctttgccatg tttcagaaac aactctggcg catcgggctt cccatacaat cgatagattg
     5701 tcgcacctga ttgcccgaca ttatcgcgag cccatttata cccatataaa tcagcatcca
     5761 tgttggaatt taatcgcggc ctagagcaag acgtttcccg ttgaatatgg ctcataacac
     5821 cccttgtatt actgtttatg taagcagaca gttttattgt tcatgaccaa aatcccttaa
     5881 cgtgagtttt cgttccactg agcgtcagac cccgtagaa
//
