LOCUS       sequence-150790-        6540 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Wild-type S. cerevisiae Cus2p gene for recombinant expression and 
            crystallization.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6540)
  AUTHORS   Rodgers ML, Tretbar US, Dehaven A, Alwan AA, Luo G, Mast HM, Hoskins
            AA
  TITLE     Conformational dynamics of stem II of the U2 snRNA.
  JOURNAL   RNA. 2015 Dec 2.
  PUBMED    26631165
REFERENCE   2  (bases 1 to 6540)
  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: "RNA. 2015 
            Dec 2."
COMMENT     Sequence Label: wt Cus2p
FEATURES             Location/Qualifiers
     source          1..6540
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      899..947
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(1122..1127,1128..1144,1145..1150))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(1177..1195)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1263..1367
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1368..1436,1437..2228)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1586..1605)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2399..2987
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2888..2907
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3141..3158
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3173..3315
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3241..3260)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3401..3423
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3417..3608)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4920..6002)
                     /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     5964..5983
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6003..6080)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6286..6305
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     6389..6408
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6389..6407
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    6408..6432
                     /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             6447..6469
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             6488..6505
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             6518..6538
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
ORIGIN
        1 tatggatgct gatgaattgg aattgaaagg tcatttgaaa aaattgaaaa aagaagaatt
       61 gcttagaagg aaacagttaa aagagagtaa tcttcagaaa agagagcttg aatataataa
      121 tgcctcaaaa aatacttcaa tatatatttc tggtcttccc acagacaaaa caacgaaaga
      181 aggactcact gagcagtttt gcaagtatgg aatgataaga accaatcgag atggggaacc
      241 cctttgcaag ctatatgtga atgataaagg cgctttcaaa ggggatgcct tgatcactta
      301 ttccaaggaa gaaagtgtta cattagcaat cgaaatgatg aacgagtcga tcttcctggg
      361 aaaacagatc agggtcgaaa gagcccaatt tcaaaacaag gaaggtgaca atatgcacgg
      421 gaaagaaaat gatttaaagg agttcaacgg acctgagcct ccaataaaga ggctaaaaaa
      481 agcgaagtct gaaggggagg gggaagtaat agattacaat gacgatgaaa gcctagcaaa
      541 agctgatcga acggtgatat ttgccaacgt attcaatatt tacaaatcat acacaaatga
      601 tgatataaat gatattcagg aagatctttt ggaaggctgt gaagagatag gtcaagtaga
      661 cagtatatct gtatctccca ataaggggga agcgacggtc gtctttaaga acaataaagt
      721 tgcattgcaa tgttgcaaaa tcatgaccgg acgttatttt gatgggcaaa agttgcttgc
      781 tttcatttct ggagatgaga acacttcaag cacaagtgac aagaatgagg acagcgaagt
      841 ggaagatgac cttatatagg gatcctaata actaagtaaa ctagtgcctg agcaataact
      901 agcataaccc ccttggggcc tctaaacggg tcttgagggg ttttttgctg aaaggaggaa
      961 ctatatccgg atatcccgca agaggcccgg cagtaccggc ataaccaagc ctatgcctac
     1021 agcatccagg gtgacggtgc cgaggatgac gatgagcgca ttgttagatt tcatacacgg
     1081 tgcctgactg cgttagcaat ttaactgtga taaactaccg cattaaagct tatcgatgat
     1141 aagctgtcaa acatgagaat tcttgaagac gaaagggcct cgtgatacgc ctatttttat
     1201 aggttaatgt catgataata atggtttctt agacgtcagg tggcactttt cggggaaatg
     1261 tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga
     1321 gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac
     1381 atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc
     1441 cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca
     1501 tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc
     1561 caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt gttgacgccg
     1621 ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac
     1681 cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca
     1741 taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg
     1801 agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac
     1861 cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct gcagcaatgg
     1921 caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat
     1981 taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg
     2041 ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg
     2101 cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc
     2161 aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc
     2221 attggtaact gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt
     2281 tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt
     2341 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
     2401 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
     2461 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
     2521 gcagagcgca gataccaaat actgtccttc tagtgtagcc gtagttaggc caccacttca
     2581 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
     2641 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
     2701 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
     2761 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
     2821 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
     2881 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
     2941 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
     3001 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt
     3061 tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc
     3121 gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agcggaagag cgcctgatgc
     3181 ggtattttct ccttacgcat ctgtgcggta tttcacaccg catatatggt gcactctcag
     3241 tacaatctgc tctgatgccg catagttaag ccagtataca ctccgctatc gctacgtgac
     3301 tgggtcatgg ctgcgccccg acacccgcca acacccgctg acgcgccctg acgggcttgt
     3361 ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag
     3421 aggttttcac cgtcatcacc gaaacgcgcg aggcagctgc ggtaaagctc atcagcgtgg
     3481 tcgtgaagcg attcacagat gtctgcctgt tcatccgcgt ccagctcgtt gagtttctcc
     3541 agaagcgtta atgtctggct tctgataaag cgggccatgt taagggcggt tttttcctgt
     3601 ttggtcactg atgcctccgt gtaaggggga tttctgttca tgggggtaat gataccgatg
     3661 aaacgagaga ggatgctcac gatacgggtt actgatgatg aacatgcccg gttactggaa
     3721 cgttgtgagg gtaaacaact ggcggtatgg atgcggcggg accagagaaa aatcactcag
     3781 ggtcaatgcc agcgcttcgt taatacagat gtaggtgttc cacagggtag ccagcagcat
     3841 cctgcgatgc agatccggaa cataatggtg cagggcgctg acttccgcgt ttccagactt
     3901 tacgaaacac ggaaaccgaa gaccattcat gttgttgctc aggtcgcaga cgttttgcag
     3961 cagcagtcgc ttcacgttcg ctcgcgtatc ggtgattcat tctgctaacc agtaaggcaa
     4021 ccccgccagc ctagccgggt cctcaacgac aggagcacga tcatgcgcac ccgtggccag
     4081 gacccaacgc tgcccgagat gcgccgcgtg cggctgctgg agatggcgga cgcgatggat
     4141 atgttctgcc aagggttggt ttgcgcattc acagttctcc gcaagaattg attggctcca
     4201 attcttggag tggtgaatcc gttagcgagg tgccgccggc ttccattcag gtcgaggtgg
     4261 cccggctcca tgcaccgcga cgcaacgcgg ggaggcagac aaggtatagg gcggcgccta
     4321 caatccatgc caacccgttc catgtgctcg ccgaggcggc ataaatcgcc gtgacgatca
     4381 gcggtccagt gatcgaagtt aggctggtaa gagccgcgag cgatccttga agctgtccct
     4441 gatggtcgtc atctacctgc ctggacagca tggcctgcaa cgcgggcatc ccgatgccgc
     4501 cggaagcgag aagaatcata atggggaagg ccatccagcc tcgcgtcgcg aacgccagca
     4561 agacgtagcc cagcgcgtcg gccgccatgc cggcgataat ggcctgcttc tcgccgaaac
     4621 gtttggtggc gggaccagtg acgaaggctt gagcgagggc gtgcaagatt ccgaataccg
     4681 caagcgacag gccgatcatc gtcgcgctcc agcgaaagcg gtcctcgccg aaaatgaccc
     4741 agagcgctgc cggcacctgt cctacgagtt gcatgataaa gaagacagtc ataagtgcgg
     4801 cgacgatagt catgccccgc gcccaccgga aggagctgac tgggttgaag gctctcaagg
     4861 gcatcggtcg agatcccggt gcctaatgag tgagctaact tacattaatt gcgttgcgct
     4921 cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac
     4981 gcgcggggag aggcggtttg cgtattgggc gccagggtgg tttttctttt caccagtgag
     5041 acgggcaaca gctgattgcc cttcaccgcc tggccctgag agagttgcag caagcggtcc
     5101 acgctggttt gccccagcag gcgaaaatcc tgtttgatgg tggttaacgg cgggatataa
     5161 catgagctgt cttcggtatc gtcgtatccc actaccgaga tatccgcacc aacgcgcagc
     5221 ccggactcgg taatggcgcg cattgcgccc agcgccatct gatcgttggc aaccagcatc
     5281 gcagtgggaa cgatgccctc attcagcatt tgcatggttt gttgaaaacc ggacatggca
     5341 ctccagtcgc cttcccgttc cgctatcggc tgaatttgat tgcgagtgag atatttatgc
     5401 cagccagcca gacgcagacg cgccgagaca gaacttaatg ggcccgctaa cagcgcgatt
     5461 tgctggtgac ccaatgcgac cagatgctcc acgcccagtc gcgtaccgtc ttcatgggag
     5521 aaaataatac tgttgatggg tgtctggtca gagacatcaa gaaataacgc cggaacatta
     5581 gtgcaggcag cttccacagc aatggcatcc tggtcatcca gcggatagtt aatgatcagc
     5641 ccactgacgc gttgcgcgag aagattgtgc accgccgctt tacaggcttc gacgccgctt
     5701 cgttctacca tcgacaccac cacgctggca cccagttgat cggcgcgaga tttaatcgcc
     5761 gcgacaattt gcgacggcgc gtgcagggcc agactggagg tggcaacgcc aatcagcaac
     5821 gactgtttgc ccgccagttg ttgtgccacg cggttgggaa tgtaattcag ctccgccatc
     5881 gccgcttcca ctttttcccg cgttttcgca gaaacgtggc tggcctggtt caccacgcgg
     5941 gaaacggtct gataagagac accggcatac tctgcgacat cgtataacgt tactggtttc
     6001 acattcacca ccctgaattg actctcttcc gggcgctatc atgccatacc gcgaaaggtt
     6061 ttgcgccatt caatggtgtc cgggatctcg acgctctccc ttatgcgact cctgcattag
     6121 gaagcagccc agtagtaggt tgaggccgtt gagcaccgcc gccgcaagga atggtgcatg
     6181 caaggagatg gcgcccaaca gtcccccggc cacggggcct gccaccatac ccacgccgaa
     6241 acaagcgctc atgagcccga agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat
     6301 ataggcgcca gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta
     6361 gaggatcgag atctcgatcc cgcgaaatta atacgactca ctatagggga attgtgagcg
     6421 gataacaatt cccctctaga aataattttg tttaacttta agaaggagat ataccatggg
     6481 cagcagccat catcatcatc atcacagcag cggcagagaa aacttgtatt tccagggcca
//
