LOCUS       sequence-193695-        6096 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of the Beta-1,3-Exo-Laminarinase FbGH17B in E.coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6096)
  TITLE     FormB
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6096)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pET28a(+)/FbGH17B
FEATURES             Location/Qualifiers
     source          1..6096
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     940..959
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(979..1056)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1262..1281
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1365..1384
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1365..1383
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1384..1408
                     /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             1423..1445
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1464..1481
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1491..1508
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             2325..2342
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2395..2413)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2409..2456
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2493..2948
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2580..2599)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2790..2811
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3040..3855)
                     /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(3040..3855)
                     /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     3763..3782
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3847..3866
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3977..4565
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4466..4485
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4719..4736
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4751..4890
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4816..4835)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4976..4998
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4992..5183)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(5992..6096,1..978))
                     /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"
ORIGIN
        1 tcttttcacc agtgagacgg gcaacagctg attgcccttc accgcctggc cctgagagag
       61 ttgcagcaag cggtccacgc tggtttgccc cagcaggcga aaatcctgtt tgatggtggt
      121 taacggcggg atataacatg agctgtcttc ggtatcgtcg tatcccacta ccgagatatc
      181 cgcaccaacg cgcagcccgg actcggtaat ggcgcgcatt gcgcccagcg ccatctgatc
      241 gttggcaacc agcatcgcag tgggaacgat gccctcattc agcatttgca tggtttgttg
      301 aaaaccggac atggcactcc agtcgccttc ccgttccgct atcggctgaa tttgattgcg
      361 agtgagatat ttatgccagc cagccagacg cagacgcgcc gagacagaac ttaatgggcc
      421 cgctaacagc gcgatttgct ggtgacccaa tgcgaccaga tgctccacgc ccagtcgcgt
      481 accgtcttca tgggagaaaa taatactgtt gatgggtgtc tggtcagaga catcaagaaa
      541 taacgccgga acattagtgc aggcagcttc cacagcaatg gcatcctggt catccagcgg
      601 atagttaatg atcagcccac tgacgcgttg cgcgagaaga ttgtgcaccg ccgctttaca
      661 ggcttcgacg ccgcttcgtt ctaccatcga caccaccacg ctggcaccca gttgatcggc
      721 gcgagattta atcgccgcga caatttgcga cggcgcgtgc agggccagac tggaggtggc
      781 aacgccaatc agcaacgact gtttgcccgc cagttgttgt gccacgcggt tgggaatgta
      841 attcagctcc gccatcgccg cttccacttt ttcccgcgtt ttcgcagaaa cgtggctggc
      901 ctggttcacc acgcgggaaa cggtctgata agagacaccg gcatactctg cgacatcgta
      961 taacgttact ggtttcacat tcaccaccct gaattgactc tcttccgggc gctatcatgc
     1021 cataccgcga aaggttttgc gccattcgat ggtgtccggg atctcgacgc tctcccttat
     1081 gcgactcctg cattaggaag cagcccagta gtaggttgag gccgttgagc accgccgccg
     1141 caaggaatgg tgcatgcaag gagatggcgc ccaacagtcc cccggccacg gggcctgcca
     1201 ccatacccac gccgaaacaa gcgctcatga gcccgaagtg gcgagcccga tcttccccat
     1261 cggtgatgtc ggcgatatag gcgccagcaa ccgcacctgt ggcgccggtg atgccggcca
     1321 cgatgcgtcc ggcgtagagg atcgagatct cgatcccgcg aaattaatac gactcactat
     1381 aggggaattg tgagcggata acaattcccc tctagaaata attttgttta actttaagaa
     1441 ggagatatac catgggcagc agccatcatc atcatcatca cagcagcggc ctggtgccgc
     1501 gcggcagcca tatggctagc accctgaata agggtctgca cggcatgtgc tttagcccgt
     1561 acctggaagg tcaaaacatc ggtaatcaac tgagcgagag ccaaatcagc cagcgtatgg
     1621 acgtgatcgc gccgtacacc caatgggttc gtagcttcag ctgcaccgag ggtaacgagc
     1681 acaccccgaa gatcgcgcac gaaaagcacc tgaaaaccat ggtgggtgcg tggattggca
     1741 gcgacaaggc gcagaacgaa aaagagctga acaccctgat caagctgggt aaaagcggct
     1801 ttgttgatat tgcggtggtt ggcaacgaaa ccctgatgcg tgaggaactg accgaggaag
     1861 agctgatcgc gtacattaac cgtgtgaagc gtagcctgcc gggtatcccg gtgggctatg
     1921 ttgatgcgta ctatcaattc gtggagcgtc cgaaactgat cgacgcgtgc gatgttattc
     1981 tggcgaactg ctatccgttt tgggaaggtt gcagcattga gcagagcgcg acctacctga
     2041 agcaaatgta tgcggtgacc cagcaagttg cgaacggcaa accggttatc attaccgaaa
     2101 ccggttggcc gaaccagggc gaccacacca aggatgcgga accgagcgag aacaacgcga
     2161 tgaaatactt cattgacacc gcgaactggg cgggtcaaaa cgaggtgccg ctgttctatt
     2221 ttagcagctt tgatgaaagc tggaaagttc accacgaggg cgacgttggc gcgcgttggg
     2281 gtctgtggga caaaaatgag aacctgaagt ttcaataact cgagcaccac caccaccacc
     2341 actgagatcc ggctgctaac aaagcccgaa aggaagctga gttggctgct gccaccgctg
     2401 agcaataact agcataaccc cttggggcct ctaaacgggt cttgaggggt tttttgctga
     2461 aaggaggaac tatatccgga ttggcgaatg ggacgcgccc tgtagcggcg cattaagcgc
     2521 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
     2581 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     2641 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     2701 acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc
     2761 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact
     2821 caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg
     2881 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgct
     2941 tacaatttag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc
     3001 taaatacatt caaatatgta tccgctcatg aattaattct tagaaaaact catcgagcat
     3061 caaatgaaac tgcaatttat tcatatcagg attatcaata ccatattttt gaaaaagccg
     3121 tttctgtaat gaaggagaaa actcaccgag gcagttccat aggatggcaa gatcctggta
     3181 tcggtctgcg attccgactc gtccaacatc aatacaacct attaatttcc cctcgtcaaa
     3241 aataaggtta tcaagtgaga aatcaccatg agtgacgact gaatccggtg agaatggcaa
     3301 aagtttatgc atttctttcc agacttgttc aacaggccag ccattacgct cgtcatcaaa
     3361 atcactcgca tcaaccaaac cgttattcat tcgtgattgc gcctgagcga gacgaaatac
     3421 gcgatcgctg ttaaaaggac aattacaaac aggaatcgaa tgcaaccggc gcaggaacac
     3481 tgccagcgca tcaacaatat tttcacctga atcaggatat tcttctaata cctggaatgc
     3541 tgttttcccg gggatcgcag tggtgagtaa ccatgcatca tcaggagtac ggataaaatg
     3601 cttgatggtc ggaagaggca taaattccgt cagccagttt agtctgacca tctcatctgt
     3661 aacatcattg gcaacgctac ctttgccatg tttcagaaac aactctggcg catcgggctt
     3721 cccatacaat cgatagattg tcgcacctga ttgcccgaca ttatcgcgag cccatttata
     3781 cccatataaa tcagcatcca tgttggaatt taatcgcggc ctagagcaag acgtttcccg
     3841 ttgaatatgg ctcataacac cccttgtatt actgtttatg taagcagaca gttttattgt
     3901 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
     3961 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
     4021 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
     4081 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgtccttcta gtgtagccgt
     4141 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     4201 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     4261 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     4321 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     4381 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     4441 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
     4501 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
     4561 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
     4621 acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt
     4681 gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag
     4741 cggaagagcg cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca
     4801 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt atacactccg
     4861 ctatcgctac gtgactgggt catggctgcg ccccgacacc cgccaacacc cgctgacgcg
     4921 ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg
     4981 agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgaggca gctgcggtaa
     5041 agctcatcag cgtggtcgtg aagcgattca cagatgtctg cctgttcatc cgcgtccagc
     5101 tcgttgagtt tctccagaag cgttaatgtc tggcttctga taaagcgggc catgttaagg
     5161 gcggtttttt cctgtttggt cactgatgcc tccgtgtaag ggggatttct gttcatgggg
     5221 gtaatgatac cgatgaaacg agagaggatg ctcacgatac gggttactga tgatgaacat
     5281 gcccggttac tggaacgttg tgagggtaaa caactggcgg tatggatgcg gcgggaccag
     5341 agaaaaatca ctcagggtca atgccagcgc ttcgttaata cagatgtagg tgttccacag
     5401 ggtagccagc agcatcctgc gatgcagatc cggaacataa tggtgcaggg cgctgacttc
     5461 cgcgtttcca gactttacga aacacggaaa ccgaagacca ttcatgttgt tgctcaggtc
     5521 gcagacgttt tgcagcagca gtcgcttcac gttcgctcgc gtatcggtga ttcattctgc
     5581 taaccagtaa ggcaaccccg ccagcctagc cgggtcctca acgacaggag cacgatcatg
     5641 cgcacccgtg gggccgccat gccggcgata atggcctgct tctcgccgaa acgtttggtg
     5701 gcgggaccag tgacgaaggc ttgagcgagg gcgtgcaaga ttccgaatac cgcaagcgac
     5761 aggccgatca tcgtcgcgct ccagcgaaag cggtcctcgc cgaaaatgac ccagagcgct
     5821 gccggcacct gtcctacgag ttgcatgata aagaagacag tcataagtgc ggcgacgata
     5881 gtcatgcccc gcgcccaccg gaaggagctg actgggttga aggctctcaa gggcatcggt
     5941 cgagatcccg gtgcctaatg agtgagctaa cttacattaa ttgcgttgcg ctcactgccc
     6001 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     6061 agaggcggtt tgcgtattgg gcgccagggt ggtttt
//
