LOCUS       sequence-217231-        6119 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 6119)
  TITLE     plasmids for expression of yeast spliceosomal proteins
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6119)
  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            "Unpublished"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: pET-DUET-Prp18
FEATURES             Location/Qualifiers
     source          1..6119
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     27..46
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        134..152
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    153..177
                     /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             192..214
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             233..250
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     promoter        364..382
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    383..407
                     /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)."
     CDS             1218..1262
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(1300..1318)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1314..1361
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1398..1853
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1485..1504)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1695..1716
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(join(1968..2759,2760..2828))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     2591..2610
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2829..2921)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3002..3590
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3491..3510
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3744..3761
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3776..3915
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3841..3860)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4001..4023
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4017..4208)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4780..5862)
                     /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     5824..5843
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5863..5940)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 aagtggcgag cccgatcttc cccatcggtg atgtcggcga tataggcgcc agcaaccgca
       61 cctgtggcgc cggtgatgcc ggccacgatg cgtccggcgt agaggatcga gatcgatctc
      121 gatcccgcga aattaatacg actcactata ggggaattgt gagcggataa caattcccct
      181 ctagaaataa ttttgtttaa ctttaagaag gagatatacc atgggcagca gccatcacca
      241 tcatcaccac agccaggatc cgaattcgag ctcggcgcgc ctgcaggtcg acaagcttgc
      301 ggccgcataa tgcttaagtc gaacagaaag taatcgtatt gtacacggcc gcataatcga
      361 aattaatacg actcactata ggggaattgt gagcggataa caattcccca tcttagtata
      421 ttagttaagt ataagaagga gatatacata tggacctaga tctagccagt atcttaaaag
      481 gtgaaatttc taagaagaag aaagagctgg ccaactctaa aggtgttcag ccaccatgca
      541 ctgagaaatt ccagccacat gaatccgcaa acatagacga aacaccgcga caggtagaac
      601 aagaaagtac agatgaagaa aacctgtcag acaatcagag tgacgatatt aggaccacca
      661 ttagcaaatt agaaaatcgg ccagaaagga tacaagaagc gatagctcaa gacaaaacca
      721 tctctgtaat cattgatcca tcacaaattg gttctactga aggaaaacct cttttgtcaa
      781 tgaaatgtaa tttatatatt cacgaaatat tatctcgttg gaaggcgtct ttggaagctt
      841 atcatccgga gctgttctta gacactaaaa aagcactctt tccactccta ctgcaactac
      901 gaaggaacca gcttgccccg gatttactta tttcccttgc gaccgttctc tatcatttac
      961 aacaacctaa agaaataaat ttagccgttc agtcctatat gaaactcagt atcggcaatg
     1021 tggcctggcc tattggtgtt actagcgtag gcattcatgc tcgtagtgca cattcgaaaa
     1081 ttcaaggagg ccggaatgct gctaacataa tgattgatga aagaacgcga ttatggatta
     1141 ccagtattaa aagattaata acttttgaag aatggtatac cagcaaccac gatagcttag
     1201 cctgactcga gtctggtaaa gaaaccgctg ctgcgaaatt tgaacgccag cacatggact
     1261 cgtctactag cgcagcttaa ttaacctagg ctgctgccac cgctgagcaa taactagcat
     1321 aaccccttgg ggcctctaaa cgggtcttga ggggtttttt gctgaaagga ggaactatat
     1381 ccggattggc gaatgggacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt
     1441 tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt
     1501 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc
     1561 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga
     1621 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc
     1681 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt
     1741 ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct
     1801 gatttaacaa aaatttaacg cgaattttaa caaaatatta acgtttacaa tttctggcgg
     1861 cacgatggca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     1921 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     1981 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     2041 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     2101 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     2161 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     2221 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     2281 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     2341 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     2401 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     2461 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     2521 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     2581 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     2641 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     2701 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     2761 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     2821 atactcatac tcttcctttt tcaatcatga ttgaagcatt tatcagggtt attgtctcat
     2881 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggtcatg accaaaatcc
     2941 cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt
     3001 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac
     3061 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct
     3121 tcagcagagc gcagatacca aatactgtcc ttctagtgta gccgtagtta ggccaccact
     3181 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg
     3241 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata
     3301 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga
     3361 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag
     3421 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg
     3481 agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac
     3541 ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca
     3601 acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg
     3661 cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc
     3721 gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcctga
     3781 tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcaatggt gcactctcag
     3841 tacaatctgc tctgatgccg catagttaag ccagtataca ctccgctatc gctacgtgac
     3901 tgggtcatgg ctgcgccccg acacccgcca acacccgctg acgcgccctg acgggcttgt
     3961 ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag
     4021 aggttttcac cgtcatcacc gaaacgcgcg aggcagctgc ggtaaagctc atcagcgtgg
     4081 tcgtgaagcg attcacagat gtctgcctgt tcatccgcgt ccagctcgtt gagtttctcc
     4141 agaagcgtta atgtctggct tctgataaag cgggccatgt taagggcggt tttttcctgt
     4201 ttggtcactg atgcctccgt gtaaggggga tttctgttca tgggggtaat gataccgatg
     4261 aaacgagaga ggatgctcac gatacgggtt actgatgatg aacatgcccg gttactggaa
     4321 cgttgtgagg gtaaacaact ggcggtatgg atgcggcggg accagagaaa aatcactcag
     4381 ggtcaatgcc agcgcttcgt taatacagat gtaggtgttc cacagggtag ccagcagcat
     4441 cctgcgatgc agatccggaa cataatggtg cagggcgctg acttccgcgt ttccagactt
     4501 tacgaaacac ggaaaccgaa gaccattcat gttgttgctc aggtcgcaga cgttttgcag
     4561 cagcagtcgc ttcacgttcg ctcgcgtatc ggtgattcat tctgctaacc agtaaggcaa
     4621 ccccgccagc ctagccgggt cctcaacgac aggagcacga tcatgctagt catgccccgc
     4681 gcccaccgga aggagctgac tgggttgaag gctctcaagg gcatcggtcg agatcccggt
     4741 gcctaatgag tgagctaact tacattaatt gcgttgcgct cactgcccgc tttccagtcg
     4801 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg
     4861 cgtattgggc gccagggtgg tttttctttt caccagtgag acgggcaaca gctgattgcc
     4921 cttcaccgcc tggccctgag agagttgcag caagcggtcc acgctggttt gccccagcag
     4981 gcgaaaatcc tgtttgatgg tggttaacgg cgggatataa catgagctgt cttcggtatc
     5041 gtcgtatccc actaccgaga tgtccgcacc aacgcgcagc ccggactcgg taatggcgcg
     5101 cattgcgccc agcgccatct gatcgttggc aaccagcatc gcagtgggaa cgatgccctc
     5161 attcagcatt tgcatggttt gttgaaaacc ggacatggca ctccagtcgc cttcccgttc
     5221 cgctatcggc tgaatttgat tgcgagtgag atatttatgc cagccagcca gacgcagacg
     5281 cgccgagaca gaacttaatg ggcccgctaa cagcgcgatt tgctggtgac ccaatgcgac
     5341 cagatgctcc acgcccagtc gcgtaccgtc ttcatgggag aaaataatac tgttgatggg
     5401 tgtctggtca gagacatcaa gaaataacgc cggaacatta gtgcaggcag cttccacagc
     5461 aatggcatcc tggtcatcca gcggatagtt aatgatcagc ccactgacgc gttgcgcgag
     5521 aagattgtgc accgccgctt tacaggcttc gacgccgctt cgttctacca tcgacaccac
     5581 cacgctggca cccagttgat cggcgcgaga tttaatcgcc gcgacaattt gcgacggcgc
     5641 gtgcagggcc agactggagg tggcaacgcc aatcagcaac gactgtttgc ccgccagttg
     5701 ttgtgccacg cggttgggaa tgtaattcag ctccgccatc gccgcttcca ctttttcccg
     5761 cgttttcgca gaaacgtggc tggcctggtt caccacgcgg gaaacggtct gataagagac
     5821 accggcatac tctgcgacat cgtataacgt tactggtttc acattcacca ccctgaattg
     5881 actctcttcc gggcgctatc atgccatacc gcgaaaggtt ttgcgccatt cgatggtgtc
     5941 cgggatctcg acgctctccc ttatgcgact cctgcattag gaagcagccc agtagtaggt
     6001 tgaggccgtt gagcaccgcc gccgcaagga atggtgcatg caaggagatg gcgcccaaca
     6061 gtcccccggc cacggggcct gccaccatac ccacgccgaa acaagcgctc atgagcccg
//
