LOCUS       sequence-86504-e        6588 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses beta-glucosidase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6588)
  AUTHORS   Carlin DA, Caster RW, Wang X, Betzenderfer SA, Chen CX, Duong VM, 
            Ryklansky CV, Alpekin A, Beaumont N, Kapoor H, Kim N, Mohabbot H, 
            Pang B, Teel R, Whithaus L, Tagkopoulos I, Siegel JB
  TITLE     Kinetic Characterization of 100 Glycoside Hydrolase Mutants Enables 
            the Discovery of Structural Features Correlated with Kinetic 
            Constants.
  JOURNAL   PLoS One. 2016 Jan 27;11(1):e0147596. doi: 
            10.1371/journal.pone.0147596. eCollection 2016.
  PUBMED    26815142
REFERENCE   2  (bases 1 to 6588)
  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: "PLoS One.";
            date: "2016-01-27"; volume: "11(1):e0147596. doi"; pages: " 
            10.1371/journal.pone.0147596. eCollection 2016"
COMMENT     Sequence Label: BglB in pET29b(+)
FEATURES             Location/Qualifiers
     source          1..6588
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             2..19
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(72..90)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      86..133
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      170..625
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(257..276)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     467..488
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(718..1533)
                     /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(718..1533)
                     /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     1441..1460
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     1525..1544
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      1655..2243
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2144..2163
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2397..2414
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2429..2571
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2497..2516)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2657..2679
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2673..2864)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(3673..4755)
                     /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     4717..4736
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4756..4833)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5039..5058
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     5146..5165
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5146..5164
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5165..5189
                     /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             5204..5226
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
ORIGIN
        1 gcaccaccac caccaccact gagatccggc tgctaacaaa gcccgaaagg aagctgagtt
       61 ggctgctgcc accgctgagc aataactagc ataacccctt ggggcctcta aacgggtctt
      121 gaggggtttt ttgctgaaag gaggaactat atccggattg gcgaatggga cgcgccctgt
      181 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
      241 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
      301 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
      361 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
      421 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
      481 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
      541 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt
      601 aacaaaatat taacgtttac aatttcaggt ggcacttttc ggggaaatgt gcgcggaacc
      661 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgaa ttaattctta
      721 gaaaaactca tcgagcatca aatgaaactg caatttattc atatcaggat tatcaatacc
      781 atatttttga aaaagccgtt tctgtaatga aggagaaaac tcaccgaggc agttccatag
      841 gatggcaaga tcctggtatc ggtctgcgat tccgactcgt ccaacatcaa tacaacctat
      901 taatttcccc tcgtcaaaaa taaggttatc aagtgagaaa tcaccatgag tgacgactga
      961 atccggtgag aatggcaaaa gtttatgcat ttctttccag acttgttcaa caggccagcc
     1021 attacgctcg tcatcaaaat cactcgcatc aaccaaaccg ttattcattc gtgattgcgc
     1081 ctgagcgaga cgaaatacgc gatcgctgtt aaaaggacaa ttacaaacag gaatcgaatg
     1141 caaccggcgc aggaacactg ccagcgcatc aacaatattt tcacctgaat caggatattc
     1201 ttctaatacc tggaatgctg ttttcccggg gatcgcagtg gtgagtaacc atgcatcatc
     1261 aggagtacgg ataaaatgct tgatggtcgg aagaggcata aattccgtca gccagtttag
     1321 tctgaccatc tcatctgtaa catcattggc aacgctacct ttgccatgtt tcagaaacaa
     1381 ctctggcgca tcgggcttcc catacaatcg atagattgtc gcacctgatt gcccgacatt
     1441 atcgcgagcc catttatacc catataaatc agcatccatg ttggaattta atcgcggcct
     1501 agagcaagac gtttcccgtt gaatatggct cataacaccc cttgtattac tgtttatgta
     1561 agcagacagt tttattgttc atgaccaaaa tcccttaacg tgagttttcg ttccactgag
     1621 cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa
     1681 tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag
     1741 agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg
     1801 tccttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat
     1861 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta
     1921 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg
     1981 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc
     2041 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa
     2101 gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc
     2161 tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt
     2221 caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct
     2281 tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc
     2341 gtattaccgc ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg
     2401 agtcagtgag cgaggaagcg gaagagcgcc tgatgcggta ttttctcctt acgcatctgt
     2461 gcggtatttc acaccgcata tatggtgcac tctcagtaca atctgctctg atgccgcata
     2521 gttaagccag tatacactcc gctatcgcta cgtgactggg tcatggctgc gccccgacac
     2581 ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga
     2641 caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa
     2701 cgcgcgaggc agctgcggta aagctcatca gcgtggtcgt gaagcgattc acagatgtct
     2761 gcctgttcat ccgcgtccag ctcgttgagt ttctccagaa gcgttaatgt ctggcttctg
     2821 ataaagcggg ccatgttaag ggcggttttt tcctgtttgg tcactgatgc ctccgtgtaa
     2881 gggggatttc tgttcatggg ggtaatgata ccgatgaaac gagagaggat gctcacgata
     2941 cgggttactg atgatgaaca tgcccggtta ctggaacgtt gtgagggtaa acaactggcg
     3001 gtatggatgc ggcgggacca gagaaaaatc actcagggtc aatgccagcg cttcgttaat
     3061 acagatgtag gtgttccaca gggtagccag cagcatcctg cgatgcagat ccggaacata
     3121 atggtgcagg gcgctgactt ccgcgtttcc agactttacg aaacacggaa accgaagacc
     3181 attcatgttg ttgctcaggt cgcagacgtt ttgcagcagc agtcgcttca cgttcgctcg
     3241 cgtatcggtg attcattctg ctaaccagta aggcaacccc gccagcctag ccgggtcctc
     3301 aacgacagga gcacgatcat gcgcacccgt ggggccgcca tgccggcgat aatggcctgc
     3361 ttctcgccga aacgtttggt ggcgggacca gtgacgaagg cttgagcgag ggcgtgcaag
     3421 attccgaata ccgcaagcga caggccgatc atcgtcgcgc tccagcgaaa gcggtcctcg
     3481 ccgaaaatga cccagagcgc tgccggcacc tgtcctacga gttgcatgat aaagaagaca
     3541 gtcataagtg cggcgacgat agtcatgccc cgcgcccacc ggaaggagct gactgggttg
     3601 aaggctctca agggcatcgg tcgagatccc ggtgcctaat gagtgagcta acttacatta
     3661 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
     3721 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgccaggg tggtttttct
     3781 tttcaccagt gagacgggca acagctgatt gcccttcacc gcctggccct gagagagttg
     3841 cagcaagcgg tccacgctgg tttgccccag caggcgaaaa tcctgtttga tggtggttaa
     3901 cggcgggata taacatgagc tgtcttcggt atcgtcgtat cccactaccg agatgtccgc
     3961 accaacgcgc agcccggact cggtaatggc gcgcattgcg cccagcgcca tctgatcgtt
     4021 ggcaaccagc atcgcagtgg gaacgatgcc ctcattcagc atttgcatgg tttgttgaaa
     4081 accggacatg gcactccagt cgccttcccg ttccgctatc ggctgaattt gattgcgagt
     4141 gagatattta tgccagccag ccagacgcag acgcgccgag acagaactta atgggcccgc
     4201 taacagcgcg atttgctggt gacccaatgc gaccagatgc tccacgccca gtcgcgtacc
     4261 gtcttcatgg gagaaaataa tactgttgat gggtgtctgg tcagagacat caagaaataa
     4321 cgccggaaca ttagtgcagg cagcttccac agcaatggca tcctggtcat ccagcggata
     4381 gttaatgatc agcccactga cgcgttgcgc gagaagattg tgcaccgccg ctttacaggc
     4441 ttcgacgccg cttcgttcta ccatcgacac caccacgctg gcacccagtt gatcggcgcg
     4501 agatttaatc gccgcgacaa tttgcgacgg cgcgtgcagg gccagactgg aggtggcaac
     4561 gccaatcagc aacgactgtt tgcccgccag ttgttgtgcc acgcggttgg gaatgtaatt
     4621 cagctccgcc atcgccgctt ccactttttc ccgcgttttc gcagaaacgt ggctggcctg
     4681 gttcaccacg cgggaaacgg tctgataaga gacaccggca tactctgcga catcgtataa
     4741 cgttactggt ttcacattca ccaccctgaa ttgactctct tccgggcgct atcatgccat
     4801 accgcgaaag gttttgcgcc attcgatggt gtccgggatc tcgacgctct cccttatgcg
     4861 actcctgcat taggaagcag cccagtagta ggttgaggcc gttgagcacc gccgccgcaa
     4921 ggaatggtgc atgcaaggag atggcgccca acagtccccc ggccacgggg cctgccacca
     4981 tacccacgcc gaaacaagcg ctcatgagcc cgaagtggcg agcccgatct tccccatcgg
     5041 tgatgtcggc gatataggcg ccagcaaccg cacctgtggc gccggtgatg ccggccacga
     5101 tgcgtccggc gtagaggatc gagatcgatc tcgatcccgc gaaattaata cgactcacta
     5161 taggggaatt gtgagcggat aacaattccc ctctagaaat aattttgttt aactttaaga
     5221 aggagatata catatgagcg agaacacctt tatctttccg gcaaccttta tgtggggcac
     5281 cagcaccagc agctatcaga ttgaaggtgg caccgatgaa ggtggtcgta ccccgagcat
     5341 ttgggatacc ttttgtcaga ttccgggtaa agttattggt ggtgattgtg gtgatgttgc
     5401 ctgtgatcat tttcaccact ttaaagaaga tgtgcagctg atgaaacagc tgggttttct
     5461 gcattatcgt tttagcgttg catggcctcg tattatgcct gcagcaggta ttatcaatga
     5521 agagggtctg ctgttttatg agcatctgct ggatgaaatt gaactggcag gtctgattcc
     5581 gatgctgacc ctgtatcatt gggatctgcc gcagtggatt gaagatgaag gcggttggac
     5641 ccagcgtgaa accattcagc atttcaaaac ctatgccagc gttatcatgg atcgttttgg
     5701 tgaacgtatt aattggtgga acaccatcaa tgaaccgtat tgtgcaagca ttctgggtta
     5761 tggcaccggt gaacatgcac cgggtcatga aaattggcgt gaagcattta ccgcagcaca
     5821 tcatattctg atgtgtcatg gtattgcaag caacctgcat aaagaaaaag gtctgaccgg
     5881 taaaattggc attaccctga atatggaaca tgttgatgca gcaagcgaac gtccggaaga
     5941 tgttgccgca gcaattcgtc gtgatggttt tatcaatcgt tggtttgcag aaccgctgtt
     6001 caatggtaaa tatcctgaag atatggtgga atggtatggc acctatctga atggtctgga
     6061 ttttgttcag cctggtgata tggaactgat tcagcagcca ggtgattttc tgggcattaa
     6121 ctattatacc cgtagcatta ttcgcagcac caatgatgca agcctgctgc aagttgaaca
     6181 ggttcacatg gaagaaccgg ttaccgatat gggttgggaa attcatccgg aaagcttcta
     6241 taaactgctg acccgcattg aaaaagattt tagcaaaggt ctgccgatcc tgattaccga
     6301 aaatggtgca gcaatgcgtg atgaactggt taatggtcag atcgaagata ccggtcgtca
     6361 tggttatatt gaagaacatc tgaaagcctg ccaccgcttt atcgaagaag gtggccagct
     6421 gaaaggttat tttgtttgga gctttctgga taactttgaa tgggcatggg gttatagcaa
     6481 acgttttggt attgtccaca tcaactatga aacccaagaa cgcaccccga aacagagcgc
     6541 actgtggttt aaacaaatga tggccaaaaa tggtttcggc agcctcga
//
