LOCUS       sequence-267723-        6280 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Yeast STE2 coding sequence in vector for in vitro transcription and 
            protein synthesis, with T7 promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6280)
  TITLE     Wagner Lab plasmids
REFERENCE   2  (bases 1 to 6280)
  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"
COMMENT     Sequence Label: pET-24a-STE2
FEATURES             Location/Qualifiers
     source          1..6280
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     31..50
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        31..49
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    50..74
                     /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             89..111
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             119..151
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             167..187
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1109..1129
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1130..1153
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     CDS             1175..1192
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1245..1263)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1259..1306
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1343..1798
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1430..1449)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1640..1661
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1890..2705)
                     /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(1890..2705)
                     /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     2613..2632
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2697..2716
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2827..3415
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3316..3335
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3569..3586
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3601..3740
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3666..3685)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3826..3848
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3842..4033)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4842..5924)
                     /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     5886..5905
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5925..6002)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6208..6227
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 tagaggatcg agatctcgat cccgcgaaat taatacgact cactataggg gaattgtgag
       61 cggataacaa ttcccctcta gaaataattt tgtttaactt taagaaggag atatacatat
      121 ggctagcatg actggtggac agcaaatggg tcgcggatcc gaattcgaaa acctgtattt
      181 tcagggcccg atgtctgatg ctgctccttc tctttccaac cttttttatg accctacgta
      241 caatccaggt cagtctacta tcaactacac ttccatctat ggaaatggtt ccactattac
      301 ctttgatgaa cttcaaggtc ttgttaacag tactgttact caagccatta tgttcggagt
      361 ccgttgtggt gctgcagccc ttactcttat tgttatgtgg atgacctcta gatctagaaa
      421 gactccaata ttcattatca accaggtctc cttgttcctt atcattttgc actctgcctt
      481 gtatttcaag tatcttcttt ctaactactc ctctgttaca tatgccttga ctggtttccc
      541 tcaatttatt tctagaggtg acgttcacgt gtacggtgcc actaacatca ttcaggtttt
      601 gcttgttgct tctattgaga cctcactagt ttttcaaatt aaggtcattt tcaccggtga
      661 caattttaag agaattggac ttatgttgac ttccatctcc tttaccctag gtatcgctac
      721 tgtcactatg tattttgttt ccgccgttaa gggtatgatc gtcacttaca atgacgtctc
      781 cgctacccag gacaaatact ttaacgcttc cactattttg ctagcctctt ccattaattt
      841 catgtctttt gttttggttg tcaaattgat tcttgccatt cgttcgcgac gttttcttgg
      901 tttgaaacaa tttgactcct ttcacattct tcttatcatg tcctgtcaat ctcttcttgt
      961 tccatcgatc atctttatct tggcctattc tcttaagcca aatcaaggta ccgacgtttt
     1021 gactactgtc gccactcttt tggctgtcct ttctcttcct ttgtcttcta tgtgggctac
     1081 cgctgccaat aatgcctcct tgggcccaga aaacctgtac ttccagggtc accatcatca
     1141 ccaccatcat cactaataag cggccgcact cgagcaccac caccaccacc actgagatcc
     1201 ggctgctaac aaagcccgaa aggaagctga gttggctgct gccaccgctg agcaataact
     1261 agcataaccc cttggggcct ctaaacgggt cttgaggggt tttttgctga aaggaggaac
     1321 tatatccgga ttggcgaatg ggacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg
     1381 gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct
     1441 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
     1501 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag
     1561 ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg
     1621 gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc
     1681 tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat
     1741 gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgct tacaatttag
     1801 gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt
     1861 caaatatgta tccgctcatg aattaattct tagaaaaact catcgagcat caaatgaaac
     1921 tgcaatttat tcatatcagg attatcaata ccatattttt gaaaaagccg tttctgtaat
     1981 gaaggagaaa actcaccgag gcagttccat aggatggcaa gatcctggta tcggtctgcg
     2041 attccgactc gtccaacatc aatacaacct attaatttcc cctcgtcaaa aataaggtta
     2101 tcaagtgaga aatcaccatg agtgacgact gaatccggtg agaatggcaa aagtttatgc
     2161 atttctttcc agacttgttc aacaggccag ccattacgct cgtcatcaaa atcactcgca
     2221 tcaaccaaac cgttattcat tcgtgattgc gcctgagcga gacgaaatac gcgatcgctg
     2281 ttaaaaggac aattacaaac aggaatcgaa tgcaaccggc gcaggaacac tgccagcgca
     2341 tcaacaatat tttcacctga atcaggatat tcttctaata cctggaatgc tgttttcccg
     2401 gggatcgcag tggtgagtaa ccatgcatca tcaggagtac ggataaaatg cttgatggtc
     2461 ggaagaggca taaattccgt cagccagttt agtctgacca tctcatctgt aacatcattg
     2521 gcaacgctac ctttgccatg tttcagaaac aactctggcg catcgggctt cccatacaat
     2581 cgatagattg tcgcacctga ttgcccgaca ttatcgcgag cccatttata cccatataaa
     2641 tcagcatcca tgttggaatt taatcgcggc ctagagcaag acgtttcccg ttgaatatgg
     2701 ctcataacac cccttgtatt actgtttatg taagcagaca gttttattgt tcatgaccaa
     2761 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     2821 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     2881 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     2941 tggcttcagc agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca
     3001 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     3061 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     3121 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     3181 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     3241 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     3301 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     3361 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     3421 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
     3481 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac
     3541 cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg
     3601 cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca atggtgcact
     3661 ctcagtacaa tctgctctga tgccgcatag ttaagccagt atacactccg ctatcgctac
     3721 gtgactgggt catggctgcg ccccgacacc cgccaacacc cgctgacgcg ccctgacggg
     3781 cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg agctgcatgt
     3841 gtcagaggtt ttcaccgtca tcaccgaaac gcgcgaggca gctgcggtaa agctcatcag
     3901 cgtggtcgtg aagcgattca cagatgtctg cctgttcatc cgcgtccagc tcgttgagtt
     3961 tctccagaag cgttaatgtc tggcttctga taaagcgggc catgttaagg gcggtttttt
     4021 cctgtttggt cactgatgcc tccgtgtaag ggggatttct gttcatgggg gtaatgatac
     4081 cgatgaaacg agagaggatg ctcacgatac gggttactga tgatgaacat gcccggttac
     4141 tggaacgttg tgagggtaaa caactggcgg tatggatgcg gcgggaccag agaaaaatca
     4201 ctcagggtca atgccagcgc ttcgttaata cagatgtagg tgttccacag ggtagccagc
     4261 agcatcctgc gatgcagatc cggaacataa tggtgcaggg cgctgacttc cgcgtttcca
     4321 gactttacga aacacggaaa ccgaagacca ttcatgttgt tgctcaggtc gcagacgttt
     4381 tgcagcagca gtcgcttcac gttcgctcgc gtatcggtga ttcattctgc taaccagtaa
     4441 ggcaaccccg ccagcctagc cgggtcctca acgacaggag cacgatcatg cgcacccgtg
     4501 gggccgccat gccggcgata atggcctgct tctcgccgaa acgtttggtg gcgggaccag
     4561 tgacgaaggc ttgagcgagg gcgtgcaaga ttccgaatac cgcaagcgac aggccgatca
     4621 tcgtcgcgct ccagcgaaag cggtcctcgc cgaaaatgac ccagagcgct gccggcacct
     4681 gtcctacgag ttgcatgata aagaagacag tcataagtgc ggcgacgata gtcatgcccc
     4741 gcgcccaccg gaaggagctg actgggttga aggctctcaa gggcatcggt cgagatcccg
     4801 gtgcctaatg agtgagctaa cttacattaa ttgcgttgcg ctcactgccc gctttccagt
     4861 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
     4921 tgcgtattgg gcgccagggt ggtttttctt ttcaccagtg agacgggcaa cagctgattg
     4981 cccttcaccg cctggccctg agagagttgc agcaagcggt ccacgctggt ttgccccagc
     5041 aggcgaaaat cctgtttgat ggtggttaac ggcgggatat aacatgagct gtcttcggta
     5101 tcgtcgtatc ccactaccga gatatccgca ccaacgcgca gcccggactc ggtaatggcg
     5161 cgcattgcgc ccagcgccat ctgatcgttg gcaaccagca tcgcagtggg aacgatgccc
     5221 tcattcagca tttgcatggt ttgttgaaaa ccggacatgg cactccagtc gccttcccgt
     5281 tccgctatcg gctgaatttg attgcgagtg agatatttat gccagccagc cagacgcaga
     5341 cgcgccgaga cagaacttaa tgggcccgct aacagcgcga tttgctggtg acccaatgcg
     5401 accagatgct ccacgcccag tcgcgtaccg tcttcatggg agaaaataat actgttgatg
     5461 ggtgtctggt cagagacatc aagaaataac gccggaacat tagtgcaggc agcttccaca
     5521 gcaatggcat cctggtcatc cagcggatag ttaatgatca gcccactgac gcgttgcgcg
     5581 agaagattgt gcaccgccgc tttacaggct tcgacgccgc ttcgttctac catcgacacc
     5641 accacgctgg cacccagttg atcggcgcga gatttaatcg ccgcgacaat ttgcgacggc
     5701 gcgtgcaggg ccagactgga ggtggcaacg ccaatcagca acgactgttt gcccgccagt
     5761 tgttgtgcca cgcggttggg aatgtaattc agctccgcca tcgccgcttc cactttttcc
     5821 cgcgttttcg cagaaacgtg gctggcctgg ttcaccacgc gggaaacggt ctgataagag
     5881 acaccggcat actctgcgac atcgtataac gttactggtt tcacattcac caccctgaat
     5941 tgactctctt ccgggcgcta tcatgccata ccgcgaaagg ttttgcgcca ttcgatggtg
     6001 tccgggatct cgacgctctc ccttatgcga ctcctgcatt aggaagcagc ccagtagtag
     6061 gttgaggccg ttgagcaccg ccgccgcaag gaatggtgca tgcaaggaga tggcgcccaa
     6121 cagtcccccg gccacggggc ctgccaccat acccacgccg aaacaagcgc tcatgagccc
     6181 gaagtggcga gcccgatctt ccccatcggt gatgtcggcg atataggcgc cagcaaccgc
     6241 acctgtggcg ccggtgatgc cggccacgat gcgtccggcg
//
