LOCUS       sequence-217174-        7862 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  protein purification.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7862)
  TITLE     plasmids for expression of yeast spliceosomal proteins
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7862)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pET29b-Prp2
FEATURES             Location/Qualifiers
     source          1..7862
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     76..95
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     183..202
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        183..201
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    202..226
                     /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             241..263
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2905..2922
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2975..2993)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2989..3036
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      3073..3528
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3160..3179)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3370..3391
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3620..4435)
                     /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(3620..4435)
                     /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     4343..4362
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4427..4446
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      4557..5145
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5046..5065
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5299..5316
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5331..5470
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5396..5415)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5556..5578
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5572..5763)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(6572..7654)
                     /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     7616..7635
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(7655..7732)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 cacggggcct gccaccatac ccacgccgaa acaagcgctc atgagcccga agtggcgagc
       61 ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca gcaaccgcac ctgtggcgcc
      121 ggtgatgccg gccacgatgc gtccggcgta gaggatcgag atcgatctcg atcccgcgaa
      181 attaatacga ctcactatag gggaattgtg agcggataac aattcccctc tagaaataat
      241 tttgtttaac tttaagaagg agatatacat atgtcaagta ttacatctga aaccggtaag
      301 cgaagagtga aacgaacgta tgaagttaca agacagaatg acaatgcagt aagaatagag
      361 ccttcttcct tgggtgagga agaagacaaa gaggccaagg ataagaactc tgccttgcag
      421 ctaaaacgat cacgctatga ccccaataaa gttttttcta atacgaacca aggtccagaa
      481 aagaacaatt taaaaggaga acaacttgga tcacaaaaaa agtcatcaaa gtatgatgaa
      541 aaaataacgt caaataatga acttactaca aagaagggtc tactaggtga ctccgagaat
      601 gaaactaagt acgcctcttc taattctaaa tttaacgtag aagttactca taaaataaag
      661 aatgcaaaag aaattgacaa aataaacaga caacgcatgt gggaagaaca acaattgcga
      721 aatgctatgg cggggcaaag cgatcaccct gatgatataa cattggaagg ctctgacaag
      781 tacgattatg tgttcgacac agatgcaatg attgattata ctaatgagga ggacgattta
      841 ctacctgaag agaaacttca atatgaggca cgtctagcac aagctttaga aactgaggaa
      901 aagcgtatac tgactataca agaggcaaga aaacttttgc ctgtacatca atacaaggat
      961 gagctactac aagagatcaa aaaaaatcaa gttttgatta taatgggtga aaccggttct
     1021 ggtaaaacca cacagttacc ccaatatttg gtagaagatg gattcactga tcagggaaag
     1081 ttacagattg ccataacaca accacgtcgt gtggcggcca cttctgttgc tgcaagagta
     1141 gcagatgaaa tgaatgttgt gcttgggaag gaagtgggtt atcagatccg ttttgaagac
     1201 aaaacaaccc caaataaaac ggttctaaag tatatgacgg atggtatgtt actaagagaa
     1261 tttcttacag attccaaatt aagcaaatat tcttgtatta tgattgatga agctcatgaa
     1321 cgtactctag ccacagatat cctgataggt ttgctgaaag atattttgcc gcagaggccc
     1381 actttaaaat tgcttatatc atcagcaaca atgaacgcaa aaaaattttc agagtttttt
     1441 gataattgtc ccattttcaa cgttcctggg agaagatatc ctgtagatat ccactacaca
     1501 ttacagccgg aggcaaacta tatacatgcc gctataacaa ctatttttca aattcatact
     1561 acgcaaagtt taccggggga tatattggtg tttttaacgg ggcaagaaga aattgaaaga
     1621 acaaaaacca aactggagga gatcatgtct aaattaggtt ctagaacaaa acaaatgatt
     1681 ataactccta tatacgcaaa tttaccccaa gagcagcaat tgaaaatttt ccagccgact
     1741 ccggaaaatt gtaggaaagt tgttttggca actaatattg cggaaacttc attgaccatt
     1801 gatggtatta gatatgtcat tgatcctgga tttgttaagg aaaattcgta cgtaccctcc
     1861 actggaatga cccaactttt aacagttcca tgctcaagag cttcagttga tcagcgtgcc
     1921 ggtagggctg gtcgtgttgg gccgggaaaa tgttttagaa tttttaccaa atggtcatat
     1981 ttgcacgagt tggaacttat gcctaagcct gagatcacaa gaacaaatct ttctaatacg
     2041 gtgttattat tattatcgct tggcgttact gacctaataa agtttcccct gatggataag
     2101 cccagtatcc ccacattacg gaagtctttg gaaaatttat acattttagg tgccttaaat
     2161 agcaaaggga caatcactcg gttaggtaaa atgatgtgcg agtttccttg tgaacctgaa
     2221 tttgcaaaag tgctttatac agctgctacg cacgaacaat gtcaaggtgt actagaagag
     2281 tgtttgacca tagtttcgat gctacatgaa acaccttcat tatttattgg tcaaaaaaga
     2341 gatgctgcgg caagtgtgtt aagcgaagtg gaaagtgatc acatattgta tttggaaatc
     2401 ttcaaccaat ggagaaactc taaattttcc agaagttggt gtcaagacca caaaattcaa
     2461 ttcaaaacga tgttgagggt aagaaatatt agaaaccaat tattcaggtg cagtgaaaag
     2521 gttggattag ttgaaaagaa cgaccaagcc aggatgaaga taggcaatat agcgggctac
     2581 ataaatgcta gaataactcg atgcttcatc agcgggtttc cgatgaatat tgtacaatta
     2641 ggccccaccg gctatcaaac catgggtagg tcatcaggtg ggctgaacgt tagtgttcat
     2701 cctaccagta tacttttcgt taatcataag gaaaaggcgc agagaccaag caaatatgtg
     2761 ctttatcaac aattgatgct aacgtcgaag gaatttatca gggactgtct agttatccct
     2821 aaggaagaat ggttgataga catggttcca caaatcttta aagatttaat tgacgataaa
     2881 acaaataggg ggaggcggct cgagcaccac caccaccacc actgagatcc ggctgctaac
     2941 aaagcccgaa aggaagctga gttggctgct gccaccgctg agcaataact agcataaccc
     3001 cttggggcct ctaaacgggt cttgaggggt tttttgctga aaggaggaac tatatccgga
     3061 ttggcgaatg ggacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
     3121 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt
     3181 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag
     3241 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt
     3301 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
     3361 tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt
     3421 cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt
     3481 aacaaaaatt taacgcgaat tttaacaaaa tattaacgct tacaatttag gtggcacttt
     3541 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta
     3601 tccgctcatg aattaattct tagaaaaact catcgagcat caaatgaaac tgcaatttat
     3661 tcatatcagg attatcaata ccatattttt gaaaaagccg tttctgtaat gaaggagaaa
     3721 actcaccgag gcagttccat aggatggcaa gatcctggta tcggtctgcg attccgactc
     3781 gtccaacatc aatacaacct attaatttcc cctcgtcaaa aataaggtta tcaagtgaga
     3841 aatcaccatg agtgacgact gaatccggtg agaatggcaa aagtttatgc atttctttcc
     3901 agacttgttc aacaggccag ccattacgct cgtcatcaaa atcactcgca tcaaccaaac
     3961 cgttattcat tcgtgattgc gcctgagcga gacgaaatac gcgatcgctg ttaaaaggac
     4021 aattacaaac aggaatcgaa tgcaaccggc gcaggaacac tgccagcgca tcaacaatat
     4081 tttcacctga atcaggatat tcttctaata cctggaatgc tgttttcccg gggatcgcag
     4141 tggtgagtaa ccatgcatca tcaggagtac ggataaaatg cttgatggtc ggaagaggca
     4201 taaattccgt cagccagttt agtctgacca tctcatctgt aacatcattg gcaacgctac
     4261 ctttgccatg tttcagaaac aactctggcg catcgggctt cccatacaat cgatagattg
     4321 tcgcacctga ttgcccgaca ttatcgcgag cccatttata cccatataaa tcagcatcca
     4381 tgttggaatt taatcgcggc ctagagcaag acgtttcccg ttgaatatgg ctcataacac
     4441 cccttgtatt actgtttatg taagcagaca gttttattgt tcatgaccaa aatcccttaa
     4501 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     4561 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     4621 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     4681 agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag
     4741 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     4801 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     4861 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     4921 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     4981 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     5041 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     5101 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     5161 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     5221 tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc
     5281 agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cctgatgcgg
     5341 tattttctcc ttacgcatct gtgcggtatt tcacaccgca atggtgcact ctcagtacaa
     5401 tctgctctga tgccgcatag ttaagccagt atacactccg ctatcgctac gtgactgggt
     5461 catggctgcg ccccgacacc cgccaacacc cgctgacgcg ccctgacggg cttgtctgct
     5521 cccggcatcc gcttacagac aagctgtgac cgtctccggg agctgcatgt gtcagaggtt
     5581 ttcaccgtca tcaccgaaac gcgcgaggca gctgcggtaa agctcatcag cgtggtcgtg
     5641 aagcgattca cagatgtctg cctgttcatc cgcgtccagc tcgttgagtt tctccagaag
     5701 cgttaatgtc tggcttctga taaagcgggc catgttaagg gcggtttttt cctgtttggt
     5761 cactgatgcc tccgtgtaag ggggatttct gttcatgggg gtaatgatac cgatgaaacg
     5821 agagaggatg ctcacgatac gggttactga tgatgaacat gcccggttac tggaacgttg
     5881 tgagggtaaa caactggcgg tatggatgcg gcgggaccag agaaaaatca ctcagggtca
     5941 atgccagcgc ttcgttaata cagatgtagg tgttccacag ggtagccagc agcatcctgc
     6001 gatgcagatc cggaacataa tggtgcaggg cgctgacttc cgcgtttcca gactttacga
     6061 aacacggaaa ccgaagacca ttcatgttgt tgctcaggtc gcagacgttt tgcagcagca
     6121 gtcgcttcac gttcgctcgc gtatcggtga ttcattctgc taaccagtaa ggcaaccccg
     6181 ccagcctagc cgggtcctca acgacaggag cacgatcatg cgcacccgtg gggccgccat
     6241 gccggcgata atggcctgct tctcgccgaa acgtttggtg gcgggaccag tgacgaaggc
     6301 ttgagcgagg gcgtgcaaga ttccgaatac cgcaagcgac aggccgatca tcgtcgcgct
     6361 ccagcgaaag cggtcctcgc cgaaaatgac ccagagcgct gccggcacct gtcctacgag
     6421 ttgcatgata aagaagacag tcataagtgc ggcgacgata gtcatgcccc gcgcccaccg
     6481 gaaggagctg actgggttga aggctctcaa gggcatcggt cgagatcccg gtgcctaatg
     6541 agtgagctaa cttacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     6601 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     6661 gcgccagggt ggtttttctt ttcaccagtg agacgggcaa cagctgattg cccttcaccg
     6721 cctggccctg agagagttgc agcaagcggt ccacgctggt ttgccccagc aggcgaaaat
     6781 cctgtttgat ggtggttaac ggcgggatat aacatgagct gtcttcggta tcgtcgtatc
     6841 ccactaccga gatgtccgca ccaacgcgca gcccggactc ggtaatggcg cgcattgcgc
     6901 ccagcgccat ctgatcgttg gcaaccagca tcgcagtggg aacgatgccc tcattcagca
     6961 tttgcatggt ttgttgaaaa ccggacatgg cactccagtc gccttcccgt tccgctatcg
     7021 gctgaatttg attgcgagtg agatatttat gccagccagc cagacgcaga cgcgccgaga
     7081 cagaacttaa tgggcccgct aacagcgcga tttgctggtg acccaatgcg accagatgct
     7141 ccacgcccag tcgcgtaccg tcttcatggg agaaaataat actgttgatg ggtgtctggt
     7201 cagagacatc aagaaataac gccggaacat tagtgcaggc agcttccaca gcaatggcat
     7261 cctggtcatc cagcggatag ttaatgatca gcccactgac gcgttgcgcg agaagattgt
     7321 gcaccgccgc tttacaggct tcgacgccgc ttcgttctac catcgacacc accacgctgg
     7381 cacccagttg atcggcgcga gatttaatcg ccgcgacaat ttgcgacggc gcgtgcaggg
     7441 ccagactgga ggtggcaacg ccaatcagca acgactgttt gcccgccagt tgttgtgcca
     7501 cgcggttggg aatgtaattc agctccgcca tcgccgcttc cactttttcc cgcgttttcg
     7561 cagaaacgtg gctggcctgg ttcaccacgc gggaaacggt ctgataagag acaccggcat
     7621 actctgcgac atcgtataac gttactggtt tcacattcac caccctgaat tgactctctt
     7681 ccgggcgcta tcatgccata ccgcgaaagg ttttgcgcca ttcgatggtg tccgggatct
     7741 cgacgctctc ccttatgcga ctcctgcatt aggaagcagc ccagtagtag gttgaggccg
     7801 ttgagcaccg ccgccgcaag gaatggtgca tgcaaggaga tggcgcccaa cagtcccccg
     7861 gc
//
