LOCUS       sequence-74806-e        5480 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Galactose-inducible expression of ER-luminal S. pyogenes SrtA in 
            yeast.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5480)
  AUTHORS   Strijbis K, Spooner E, Ploegh HL
  TITLE     Protein ligation in living cells using sortase.
  JOURNAL   Traffic. 2012 Jun;13(6):780-9. doi: 
            10.1111/j.1600-0854.2012.01345.x. Epub 2012 Mar 23.
  PUBMED    22348280
REFERENCE   2  (bases 1 to 5480)
  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"; doi: 
            "10.1111/j.1600-0854.2012.01345.x"; journalName: "Traffic"; date: 
            "2012-06"; volume: "13"; issue: "6"; pages: "780-9"
COMMENT     Sequence Label: pRS303-pGAL-SPPHO5-G-SrtAstrep-HA-HDEL (pKS82)
FEATURES             Location/Qualifiers
     source          1..5480
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        315..502
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     CDS             503..1162
                     /codon_start=1
                     /gene="S. cerevisiae HIS3"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=HIS3
                     /note="yeast auxotrophic marker"
                     /translation="MTEQKALVKRITNETKIQIAISLKGGPLAIEHSIFPEKEAEAVAE
                     QATQSQVINVHTGIGFLDHMIHALAKHSGWSLIVECIGDLHIDDHHTTEDCGIALGQAF
                     KEALLARGVKRFGSGFAPLDEALSRAVVDLSNRPYAVVELGLQREKVGDLSCEMIPHFL
                     ESFAEASRITLHVDCLRGKNDHHRSESAFKALAVAIREATSPNGTNDVPSTKGVLM"
     rep_origin      complement(1423..1878)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1560..1581)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1772..1791
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2008..2030
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2022..2039
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2023..2039
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2046..2065
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2046..2064
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        2084..2525
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    2084..2201
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     2496..2519
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     CDS             3140..3166
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     promoter        complement(3232..3250)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3269..3285)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3269..3285)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3282..3304)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3293..3309
                     /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)."
     promoter        complement(join(3317..3323,3324..3341,3342..3347))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(3500..3517)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3671..4259)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(4430..5221,5222..5290))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFFHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5053..5072
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5291..5395)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5453..5471
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accataattc cgttttaaga gcttggtgag cgctaggagt cactgccagg tatcgtttga
      241 acacggcatt agtcagggaa gtcataacac agtcctttcc cgcaattttc tttttctatt
      301 actcttggcc tcctctagta cactctatat ttttttatgc ctcggtaatg attttcattt
      361 ttttttttcc acctagcgga tgactctttt tttttcttag cgattggcat tatcacataa
      421 tgaattatac attatataaa gtaatgtgat ttcttcgaag aatatactaa aaaatgagca
      481 ggcaagataa acgaaggcaa agatgacaga gcagaaagcc ctagtaaagc gtattacaaa
      541 tgaaaccaag attcagattg cgatctcttt aaagggtggt cccctagcga tagagcactc
      601 gatcttccca gaaaaagagg cagaagcagt agcagaacag gccacacaat cgcaagtgat
      661 taacgtccac acaggtatag ggtttctgga ccatatgata catgctctgg ccaagcattc
      721 cggctggtcg ctaatcgttg agtgcattgg tgacttacac atagacgacc atcacaccac
      781 tgaagactgc gggattgctc tcggtcaagc ttttaaagag gccctactgg cgcgtggagt
      841 aaaaaggttt ggatcaggat ttgcgccttt ggatgaggca ctttccagag cggtggtaga
      901 tctttcgaac aggccgtacg cagttgtcga acttggtttg caaagggaga aagtaggaga
      961 tctctcttgc gagatgatcc cgcattttct tgaaagcttt gcagaggcta gcagaattac
     1021 cctccacgtt gattgtctgc gaggcaagaa tgatcatcac cgtagtgaga gtgcgttcaa
     1081 ggctcttgcg gttgccataa gagaagccac ctcgcccaat ggtaccaacg atgttccctc
     1141 caccaaaggt gttcttatgt agtgacaccg attatttaaa gctgcagcat acgatatata
     1201 tacatgtgta tatatgtata cctatgaatg tcagtaagta tgtatacgaa cagtatgata
     1261 ctgaagatga caaggtaatg catcattcta tacgtgtcat tctgaacgag gcgcgctttc
     1321 cttttttctt tttgcttttt cttttttttt ctcttgaact cgacggatca tatgcggtgt
     1381 gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggaaattgta aacgttaata
     1441 ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg
     1501 aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc
     1561 cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa
     1621 ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt
     1681 cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac
     1741 ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta
     1801 gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg
     1861 cgccgctaca gggcgcgtcg cgccattcgc cattcaggct gcgcaactgt tgggaagggc
     1921 gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa ggggggatgt gctgcaaggc
     1981 gattaagttg ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg acggccagtg
     2041 aattgtaata cgactcacta tagggcgaat tggagctcta gtacggatta gaagccgccg
     2101 agcgggtgac agccctccga aggaagactc tcctccgtgc gtcctcgtct tcaccggtcg
     2161 cgttcctgaa acgcagatgt gcctcgcgcc gcactgctcc gaacaataaa gattctacaa
     2221 tactagcttt tatggttatg aagaggaaaa attggcagta acctggcccc acaaaccttc
     2281 aaatgaacga atcaaattaa caaccatagg atgataatgc gattagtttt ttagccttat
     2341 ttctggggta attaatcagc gaagcgatga tttttgatct attaacagat atataaatgc
     2401 aaaaactgca taaccacttt aactaatact ttcaacattt tcggtttgta ttacttctta
     2461 ttcaaatgta ataaaagtat caacaaaaaa ttgttaatat acctctatac tttaacgtca
     2521 aggagaaaaa accccggatt ctagaatgtt taaatctgtt gtttattcaa ttttagccgc
     2581 ttctttggcc aatgcaggta ccattccctt aggatccagt gtcttgcaag cacaaatggc
     2641 ggctcagcaa cttcctgtta tagggggcat tgccatacca gagcttggca ttaatttacc
     2701 aatttttaaa ggtttaggaa atactgagct tatttatggc gcaggaacga tgaaagaaga
     2761 acaagttatg ggaggagaaa ataattattc tcttgccagt catcatattt ttggaattac
     2821 aggttcatct caaatgctct tttcgccgct tgaaagagca caaaatggga tgtccatcta
     2881 tttaacagat aaagaaaaaa tttacgaata catcataaaa gatgttttca cggtagctcc
     2941 tgaacgcgtt gatgttatcg atgatacagc tggtctcaaa gaagtgactt tagtgacttg
     3001 tacagatatc gaagcaacag aacgtattat tgtcaaagga gaactaaaaa cagaatacga
     3061 ctttgataaa gcgcccgccg atgtattgaa agcttttaat cattcttata accaagtatc
     3121 taccgtcgac gcggccgcat acccatacga cgtaccagac tacgcaaatg gggccgcaag
     3181 cgagcatgac gagctgtagc tcgagggggg gcccggtacc agcttttgtt ccctttagtg
     3241 agggttaatt cgagctggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
     3301 cgctcacaat tccacacaac ataggagccg gaagcataaa gtgtaaagcc tggggtgcct
     3361 aatgagtgag gtaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     3421 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     3481 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     3541 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     3601 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     3661 tgctggcgtt tttccatagg ctcggccccc ctgacgagca tcacaaaaat cgacgctcaa
     3721 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgttcccc cctggaagct
     3781 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     3841 cttcgggaag cgtggcgctt tctcaatgct cacgctgtag gtatctcagt tcggtgtagg
     3901 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     3961 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     4021 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     4081 agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc gctctgctga
     4141 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     4201 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     4261 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     4321 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     4381 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     4441 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     4501 tgcccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     4561 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     4621 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     4681 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     4741 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     4801 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg aaaaaaagcg gttagctcct
     4861 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     4921 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     4981 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     5041 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     5101 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     5161 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     5221 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     5281 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     5341 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     5401 ttccccgaaa agtgccacct gacgtcttat tatcatgaca ttaacctata aaaataggcg
     5461 tatcacgagg ccctttcgtc
//
