LOCUS       pYSD085                 3829 bp ds-DNA     circular     12-NOV-2023
DEFINITION  .
ACCESSION   <unknown id>
VERSION     <unknown id>
COMMENT     Encodes the HA-tagged 649 Stalk yeast surface display anchor
            protein as a Type 4a part to be used in the yeast secretion/display
            (YSD) extension of the MoClo yeast toolkit (YTK) Vector backbone is
            the Yeast MoClo toolkit entry vector pYTK001 Reference for 649
            Stalk: McMahon, C., Baier, A.S., Pascolutti, R., Wegrecki, M.,
            Zheng, S., Ong, J.X., et al. (2018). Yeast surface display platform
            for rapid discovery of conformationally selective nanobodies. Nat
            Struct Mol Biol 25(3), 289-296. doi: 10.1038/s41594-018-0028-6.
FEATURES             Location/Qualifiers
     misc_feature    3..8
                     /label="BsaI"
                     /ApEinfo_revcolor="#f8d3a9"
                     /ApEinfo_fwdcolor="#f8d3a9"
     CDS             11..94
                     /label="linker- HA-tag TEV site"
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /translation="SGGGGSGGGGSYPYDVPDYAKENLYFQS"
     misc_feature    14..43
                     /label="GGGGSGGGGS Linker"
                     /ApEinfo_revcolor="#b7e6d7"
                     /ApEinfo_fwdcolor="#b7e6d7"
     misc_feature    44..70
                     /label="HA tag"
                     /ApEinfo_revcolor="#c6c9d1"
                     /ApEinfo_fwdcolor="#c6c9d1"
     misc_feature    74..94
                     /label="TEV protease site"
                     /ApEinfo_revcolor="#d6b295"
                     /ApEinfo_fwdcolor="#d6b295"
     misc_feature    95..2041
                     /label="649 Stalk sequence"
                     /ApEinfo_revcolor="#ff9ccd"
                     /ApEinfo_fwdcolor="#ff9ccd"
     CDS             95..2041
                     /label="649 Stalk"
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /translation="QTSSPSTEASVSTSSTSSSASQSSDPTTTSSSSSSSSPSSQSEEISSSPTVSTTPSTSSSSSSMTSTTTTKSISTSTTSSAPVTDVTVSSSPSKSTSTSTSTETSKTPTSMTEYTSSTSIISTPVSHSQTGLSASSSSSSTTSGSSSTKSESSTTSGSSQSVESTSSHATVLANSAEMVTTSSSSSSTSEMSLTSTATSVPVSSSSSTTYSTSASTQAVTTTSSSTVSTTSSSTTLTSAFTHSSTTSSDQPPSDTTSPSTTHEPHVTTQTSSETSSSKSSSTSSSSTSQTSESATPSDSVSPGSSTSTSSSSTSTSTSISSGETTTSSSSSSATTTSNSATLSVSTTQTSIEASSSTTSTSSSTITTSSSSAHISSKSQSSITYPSSSTSSSTSSSISSESESFESTSAEDAPSTAPSSSVSSKSSTSTTSSTSTSSSTPSPSPSSVSSSSTSSLTTSAVSTPATSHSQSTVVTTTTITTSTGPVMSTTTAYSSSSTSSSESSEVQSVMSSTPSSTSTTTSSESTSSSSTASTSPSTSQTFETSPTIGGVPSTTSFVSTPTTKLSHTTSTMTAQSDSKSTHSSSTSTEDKSSTASAVDESTTTSTSTESTTSVTSGTSHSAKESSSNSKVYSSQTAHSSISVASSPSTK"
     misc_feature    244..247
                     /label="BsmBI overhang"
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
     misc_feature    244..249
                     /label="BsaI site destroyed"
                     /ApEinfo_revcolor="#ffef86"
                     /ApEinfo_fwdcolor="#ffef86"
     misc_feature    2042..2161
                     /label="GPI Anchor Sequence"
                     /ApEinfo_revcolor="#b7e6d7"
                     /ApEinfo_fwdcolor="#b7e6d7"
     CDS             2042..2161
                     /label="GPI Anchor"
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /translation="GAQIQSSMVEISTYAGSANSVNAGAGAGALFLLLSLAII*"
     misc_feature    2159..2161
                     /label="Stop codon"
                     /ApEinfo_revcolor="#9eafd2"
                     /ApEinfo_fwdcolor="#9eafd2"
     misc_feature    2162..2167
                     /label="XhoI"
                     /ApEinfo_revcolor="#b4abac"
                     /ApEinfo_fwdcolor="#b4abac"
     misc_feature    complement(2168..2171)
                     /label="tggc 4a BsaI overhang"
                     /ApEinfo_revcolor="#b7e6d7"
                     /ApEinfo_fwdcolor="#b7e6d7"
     misc_feature    complement(2173..2174)
                     /label="BsaI"
                     /ApEinfo_revcolor="#b7e6d7"
                     /ApEinfo_fwdcolor="#b7e6d7"
     protein_bind    2175..2178
                     /label="BsmBI"
                     /ApEinfo_revcolor="#b1ff67"
                     /ApEinfo_fwdcolor="#b1ff67"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}width 5 offset 0"
     misc_feature    complement(2190..2298)
                     /label="CamR Terminator"
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}width 5 offset 0"
     CDS             complement(2299..2958)
                     /label="CamR"
                     /ApEinfo_revcolor="#0000ff"
                     /ApEinfo_fwdcolor="#0000ff"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}width 5 offset 0"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFLKTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSSLWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANMDNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA*"
     promoter        complement(2959..3063)
                     /label="CamR Promoter"
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}width 5 offset 0"
     rep_origin      complement(3064..3827)
                     /label="ColE1"
                     /ApEinfo_revcolor="#7f7f7f"
                     /ApEinfo_fwdcolor="#7f7f7f"
                     /ApEinfo_graphicformat="arrow_data {{0 1 2 0 0 -1} {} 0}width 5 offset 0"
ORIGIN
        1 TCGGTCTCAA TCCGGTGGAG GGGGTTCTGG TGGTGGCGGA TCATACCCAT ACGATGTTCC
       61 AGATTACGCT Aaggaaaatt tgtattttca atctCAAACA TCCTCACCCT CTACAGAAGC
      121 ATCAGTATCA ACCTCTAGTA CCTCTTCCTC TGCTTCACAA TCATCTGACC CAACTACAAC
      181 ATCTTCGTCC AGTTCATCGT CTTCCCCATC TTCCCAATCT GAGGAAATTT CATCATCACC
      241 GACcGTaTCA ACTACACCAT CAACTTCTTC ATCATCTTCC TCAATGACTT CAACCACCAC
      301 AACAAAGTCA ATCTCAACTT CCACTACAAG TTCAGCTCCA GTTACAGATG TGACAGTTTC
      361 CTCATCGCCT AGTAAATCTA CCTCTACTTC GACAAGTACA GAAACATCTA AAACACCTAC
      421 TTCAATGACA GAGTATACAT CTAGTACATC GATAATTTCG ACTCCAGTTA GTCACTCGCA
      481 GACAGGTTTG TCGGCTTCAT CAAGTTCATC ATCTACAACA TCCGGTTCTT CGTCCACTAA
      541 ATCAGAAAGT TCGACAACAT CTGGCTCTTC CCAGTCCGTG GAATCAACCT CCAGCCACGC
      601 CACTGTTCTT GCTAATTCCG CAGAAATGGT CACAACATCC TCTAGTTCAT CCTCAACATC
      661 CGAAATGTCA TTAACTAGTA CTGCTACCAG TGTACCAGTC TCATCTAGTA GCAGTACGAC
      721 ATATTCTACT AGCGCATCTA CACAAGCCGT CACTACAACA TCTTCTTCCA CTGTATCTAC
      781 AACTTCTTCT AGTACAACGT TAACAAGCGC ATTCACACAT TCTTCAACCA CATCGTCCGA
      841 CCAGCCACCC AGCGACACTA CAAGTCCATC TACGACACAC GAACCTCATG TAACCACTCA
      901 GACGTCATCA GAAACATCTT CTTCTAAGTC ATCTTCTACT TCTTCTTCAA GTACATCTCA
      961 AACCTCTGAG TCTGCAACAC CATCCGATTC CGTATCACCT GGAAGTTCTA CATCAACATC
     1021 TTCTAGTAGC ACTTCTACTT CCACTTCTAT TTCCAGTGGA GAAACGACAA CTTCTTCTTC
     1081 TTCATCATCT GCCACGACCA CTTCTAACAG CGCAACCTTG TCAGTCTCTA CCACACAAAC
     1141 TTCGATTGAA GCCAGTTCAT CTACTACATC TACATCTAGT TCAACAATTA CAACTTCAAG
     1201 TAGTAGCGCT CACATATCGT CGAAATCTCA ATCTAGTATT ACCTATCCCT CTTCCTCGAC
     1261 ATCTTCATCT ACATCGTCCT CAATTTCTAG CGAATCTGAA AGTTTTGAAT CGACATCAGC
     1321 AGAAGATGCT CCATCAACAG CACCTTCATC AAGTGTCTCT TCTAAGAGTT CTACCTCTAC
     1381 AACATCAAGC ACATCGACAT CTTCAAGCAC TCCATCTCCA TCACCATCTT CCGTGAGTTC
     1441 TTCCTCCACC AGCTCATTGA CAACTTCTGC TGTATCAACA CCAGCTACCT CTCATTCTCA
     1501 AAGTACTGTA GTAACCACCA CTACTATTAC TACATCAACA GGTCCAGTGA TGTCTACGAC
     1561 AACAGCTTAT TCTTCTAGTT CTACTAGCAG CTCGGAATCT TCTGAGGTTC AGTCTGTCAT
     1621 GTCATCTACG CCTAGTTCAA CATCAACAAC AACCAGTTCG GAATCTACTT CATCTAGCTC
     1681 CACAGCTTCT ACCTCACCAT CAACCTCGCA AACTTTCGAA ACTTCTCCTA CTATAGGAGG
     1741 TGTCCCCTCA ACCACTTCAT TTGTCTCTAC GCCAACAACG AAATTGTCGC ACACTACTTC
     1801 CACTATGACA GCACAGTCCG ATAGTAAGTC TACCCACTCC TCAAGCACAT CGACAGAAGA
     1861 TAAATCATCC ACTGCTTCTG CAGTTGACGA AAGCACTACA ACATCCACTT CCACGGAGTC
     1921 TACTACATCA GTAACATCAG GCACCTCCCA TTCCGCTAAA GAATCTTCGT CAAATTCTAA
     1981 GGTGTATAGT TCACAGACAG CACACTCATC CATAAGTGTT GCATCATCAC CTAGTACAAA
     2041 GGGCGCCCAA ATCCAATCTT CTATGGTTGA AATCTCTACC TACGCTGGTT CTGCTAACTC
     2101 TGTTAACGCT GGTGCTGGTG CTGGTGCTTT GTTCTTGTTG TTGTCTTTGG CTATCATCta
     2161 aCTCGAGTGG CTGAgaccgc actgactaga ccaataaaaa acgcccggcg gcaaccgagc
     2221 gttctgaaca aatccagatg gagttctgag gtcattactg gatctatcaa caggagtcca
     2281 agcgagctcg atatcaaatt acgccccgcc ctgccactca tcgcagtact gttgtaattc
     2341 attaagcatt ctgccgacat ggaagccatc acaaacggca tgatgaacct gaatcgccag
     2401 cggcatcagc accttgtcgc cttgcgtata atatttgccc atggtgaaaa cgggggcgaa
     2461 gaagttgtcc atattggcca cgtttaaatc aaaactggtg aaactcaccc agggattggc
     2521 tgaaacgaaa aacatattct caataaaccc tttagggaaa taggccaggt tttcaccgta
     2581 acacgccaca tcttgcgaat atatgtgtag aaactgccgg aaatcgtcgt ggtattcact
     2641 ccagagcgat gaaaacgttt cagtttgctc atggaaaacg gtgtaacaag ggtgaacact
     2701 atcccatatc accagctcac cgtctttcat tgccatacga aattccggat gagcattcat
     2761 caggcgggca agaatgtgaa taaaggccgg ataaaacttg tgcttatttt tctttacggt
     2821 ctttaaaaag gccgtaatat ccagctgaac ggtctggtta taggtacatt gagcaactga
     2881 ctgaaatgcc tcaaaatgtt ctttacgatg ccattgggat atatcaacgg tggtatatcc
     2941 agtgattttt ttctccattt tagcttcctt agctcctgaa aatctcgata actcaaaaaa
     3001 tacgcccggt agtgatctta tttcattatg gtgaaagttg gaacctctta cgtgcccgat
     3061 caatcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     3121 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     3181 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     3241 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc
     3301 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     3361 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     3421 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     3481 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     3541 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     3601 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     3661 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     3721 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     3781 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtag
//