LOCUS       pSUMO-CBWDpep-Me        5809 bp DNA     circular SYN 24-JAN-2023
DEFINITION  synthetic circular DNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5809)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jan 24, 2023 from SnapGene 6.1.1
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5809
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        21..39
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    40..64
                     /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             79..101
                     /note="efficient ribosome binding site from bacteriophage
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             114..131
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             138..425
                     /codon_start=1
                     /gene="S. cerevisiae SMT3 (truncated and modified)"
                     /product="cleavable ubiquitin-like protein tag"
                     /label=SUMO
                     /note="modified to prevent cleavage by eukaryotic SUMO 
                     proteases"
                     /translation="DSEVNQEAKPEVKPEVKPETHINLKVSDGSSEIFFKIKKTTPLRR
                     LMEAFAKRQGKEMDSLRFLYDGIRIQADQAPEDLDMEDNDIIEAHREQIGG"
     primer_bind     366..390
                     /label=SeqSUMO_for
     primer_bind     complement(403..462)
                     /label=KE102
     CDS             426..668
                     /codon_start=1
                     /label=CBWD-metap1-fusion
                     /translation="AEEEYAEDCPELVPIETKNQEMAAVETRVCETDGCSSEAKLQCPT
                     CIKLGIQGSYFCSQECFKGSWATHKLLHKKAKDEK"
     CDS             426..476
                     /codon_start=1
                     /label=peptide
                     /translation="AEEEYAEDCPELVPIET"
     primer_bind     444..503
                     /label=KE103
     CDS             690..707
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(718..741)
                     /label=SeqSUMO_rev
     terminator      774..821
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      858..1313
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     CDS             complement(1406..2221)
                     /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"
     rep_origin      2343..2931
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    3117..3259
                     /label=bom
                     /note="basis of mobility region from pBR322"
     CDS             complement(3361..3552)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4361..5443)
                     /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"
     promoter        complement(5444..5521)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 agatctcgat cccgcgaaat taatacgact cactataggg gaattgtgag cggataacaa
       61 ttcccctcta gaaataattt tgtttaactt taagaaggag atataccatg ggtcatcacc
      121 atcatcatca cgggtcggac tcagaagtca atcaagaagc taagccagag gtcaagccag
      181 aagtcaagcc tgagactcac atcaatttaa aggtgtccga tggatcttca gagatcttct
      241 tcaagatcaa aaagaccact cctttaagaa ggctgatgga agcgttcgct aaaagacagg
      301 gtaaggaaat ggactcctta agattcttgt acgacggtat tagaattcaa gctgatcagg
      361 cccctgaaga tttggacatg gaggataacg atattattga ggctcaccgc gaacagattg
      421 gaggagcgga agaagagtac gcggaggatt gtcctgaatt ggtccccatt gagaccaaga
      481 accaagaaat ggcggcggtg gagacgcggg tgtgcgagac ggacggctgc agcagcgagg
      541 ccaagctcca gtgtcccacc tgcatcaagc tgggcatcca gggctcgtac ttctgctcgc
      601 aggaatgctt taaaggaagc tgggccactc acaagttact acataagaaa gcaaaagatg
      661 aaaagtaaaa gcttgcggcc gcactcgagc accaccacca ccaccactga gatccggctg
      721 ctaacaaagc ccgaaaggaa gctgagttgg ctgctgccac cgctgagcaa taactagcat
      781 aaccccttgg ggcctctaaa cgggtcttga ggggtttttt gctgaaagga ggaactatat
      841 ccggattggc gaatgggacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt
      901 tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt
      961 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc
     1021 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga
     1081 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc
     1141 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt
     1201 ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct
     1261 gatttaacaa aaatttaacg cgaattttaa caaaatatta acgtttacaa tttcaggtgg
     1321 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
     1381 tatgtatccg ctcatgaatt aattcttaga aaaactcatc gagcatcaaa tgaaactgca
     1441 atttattcat atcaggatta tcaataccat atttttgaaa aagccgtttc tgtaatgaag
     1501 gagaaaactc accgaggcag ttccatagga tggcaagatc ctggtatcgg tctgcgattc
     1561 cgactcgtcc aacatcaata caacctatta atttcccctc gtcaaaaata aggttatcaa
     1621 gtgagaaatc accatgagtg acgactgaat ccggtgagaa tggcaaaagt ttatgcattt
     1681 ctttccagac ttgttcaaca ggccagccat tacgctcgtc atcaaaatca ctcgcatcaa
     1741 ccaaaccgtt attcattcgt gattgcgcct gagcgagacg aaatacgcga tcgctgttaa
     1801 aaggacaatt acaaacagga atcgaatgca accggcgcag gaacactgcc agcgcatcaa
     1861 caatattttc acctgaatca ggatattctt ctaatacctg gaatgctgtt ttcccgggga
     1921 tcgcagtggt gagtaaccat gcatcatcag gagtacggat aaaatgcttg atggtcggaa
     1981 gaggcataaa ttccgtcagc cagtttagtc tgaccatctc atctgtaaca tcattggcaa
     2041 cgctaccttt gccatgtttc agaaacaact ctggcgcatc gggcttccca tacaatcgat
     2101 agattgtcgc acctgattgc ccgacattat cgcgagccca tttataccca tataaatcag
     2161 catccatgtt ggaatttaat cgcggcctag agcaagacgt ttcccgttga atatggctca
     2221 taacacccct tgtattactg tttatgtaag cagacagttt tattgttcat gaccaaaatc
     2281 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     2341 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     2401 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     2461 ttcagcagag cgcagatacc aaatactgtc cttctagtgt agccgtagtt aggccaccac
     2521 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     2581 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     2641 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     2701 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     2761 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
     2821 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
     2881 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
     2941 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct
     3001 gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct
     3061 cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgcctg
     3121 atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatata tggtgcactc
     3181 tcagtacaat ctgctctgat gccgcatagt taagccagta tacactccgc tatcgctacg
     3241 tgactgggtc atggctgcgc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc
     3301 ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg
     3361 tcagaggttt tcaccgtcat caccgaaacg cgcgaggcag ctgcggtaaa gctcatcagc
     3421 gtggtcgtga agcgattcac agatgtctgc ctgttcatcc gcgtccagct cgttgagttt
     3481 ctccagaagc gttaatgtct ggcttctgat aaagcgggcc atgttaaggg cggttttttc
     3541 ctgtttggtc actgatgcct ccgtgtaagg gggatttctg ttcatggggg taatgatacc
     3601 gatgaaacga gagaggatgc tcacgatacg ggttactgat gatgaacatg cccggttact
     3661 ggaacgttgt gagggtaaac aactggcggt atggatgcgg cgggaccaga gaaaaatcac
     3721 tcagggtcaa tgccagcgct tcgttaatac agatgtaggt gttccacagg gtagccagca
     3781 gcatcctgcg atgcagatcc ggaacataat ggtgcagggc gctgacttcc gcgtttccag
     3841 actttacgaa acacggaaac cgaagaccat tcatgttgtt gctcaggtcg cagacgtttt
     3901 gcagcagcag tcgcttcacg ttcgctcgcg tatcggtgat tcattctgct aaccagtaag
     3961 gcaaccccgc cagcctagcc gggtcctcaa cgacaggagc acgatcatgc gcacccgtgg
     4021 ggccgccatg ccggcgataa tggcctgctt ctcgccgaaa cgtttggtgg cgggaccagt
     4081 gacgaaggct tgagcgaggg cgtgcaagat tccgaatacc gcaagcgaca ggccgatcat
     4141 cgtcgcgctc cagcgaaagc ggtcctcgcc gaaaatgacc cagagcgctg ccggcacctg
     4201 tcctacgagt tgcatgataa agaagacagt cataagtgcg gcgacgatag tcatgccccg
     4261 cgcccaccgg aaggagctga ctgggttgaa ggctctcaag ggcatcggtc gagatcccgg
     4321 tgcctaatga gtgagctaac ttacattaat tgcgttgcgc tcactgcccg ctttccagtc
     4381 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     4441 gcgtattggg cgccagggtg gtttttcttt tcaccagtga gacgggcaac agctgattgc
     4501 ccttcaccgc ctggccctga gagagttgca gcaagcggtc cacgctggtt tgccccagca
     4561 ggcgaaaatc ctgtttgatg gtggttaacg gcgggatata acatgagctg tcttcggtat
     4621 cgtcgtatcc cactaccgag atatccgcac caacgcgcag cccggactcg gtaatggcgc
     4681 gcattgcgcc cagcgccatc tgatcgttgg caaccagcat cgcagtggga acgatgccct
     4741 cattcagcat ttgcatggtt tgttgaaaac cggacatggc actccagtcg ccttcccgtt
     4801 ccgctatcgg ctgaatttga ttgcgagtga gatatttatg ccagccagcc agacgcagac
     4861 gcgccgagac agaacttaat gggcccgcta acagcgcgat ttgctggtga cccaatgcga
     4921 ccagatgctc cacgcccagt cgcgtaccgt cttcatggga gaaaataata ctgttgatgg
     4981 gtgtctggtc agagacatca agaaataacg ccggaacatt agtgcaggca gcttccacag
     5041 caatggcatc ctggtcatcc agcggatagt taatgatcag cccactgacg cgttgcgcga
     5101 gaagattgtg caccgccgct ttacaggctt cgacgccgct tcgttctacc atcgacacca
     5161 ccacgctggc acccagttga tcggcgcgag atttaatcgc cgcgacaatt tgcgacggcg
     5221 cgtgcagggc cagactggag gtggcaacgc caatcagcaa cgactgtttg cccgccagtt
     5281 gttgtgccac gcggttggga atgtaattca gctccgccat cgccgcttcc actttttccc
     5341 gcgttttcgc agaaacgtgg ctggcctggt tcaccacgcg ggaaacggtc tgataagaga
     5401 caccggcata ctctgcgaca tcgtataacg ttactggttt cacattcacc accctgaatt
     5461 gactctcttc cgggcgctat catgccatac cgcgaaaggt tttgcgccat tcgatggtgt
     5521 ccgggatctc gacgctctcc cttatgcgac tcctgcatta ggaagcagcc cagtagtagg
     5581 ttgaggccgt tgagcaccgc cgccgcaagg aatggtgcat gcaaggagat ggcgcccaac
     5641 agtcccccgg ccacggggcc tgccaccata cccacgccga aacaagcgct catgagcccg
     5701 aagtggcgag cccgatcttc cccatcggtg atgtcggcga tataggcgcc agcaaccgca
     5761 cctgtggcgc cggtgatgcc ggccacgatg cgtccggcgt agaggatcg
//
