LOCUS       sequence-193697-        6522 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of the Beta-1,3-Endo-Laminarinase FbGH17A in E.coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6522)
  TITLE     FormB
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6522)
  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            "Unpublished"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(+)/FbGH17A
FEATURES             Location/Qualifiers
     source          1..6522
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     960..979
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(999..1076)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1282..1301
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1385..1404
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1385..1403
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1404..1428
                     /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             1443..1465
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1484..1501
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1511..1528
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             2771..2788
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2841..2859)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2855..2902
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2939..3394
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3026..3045)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3236..3257
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3486..4301)
                     /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(3486..4301)
                     /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     4209..4228
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4293..4312
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      4423..5011
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4912..4931
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5165..5182
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5197..5336
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5262..5281)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5422..5444
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5438..5629)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(6438..6522,1..998))
                     /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 ttgggcgcca gggtggtttt tcttttcacc agtgagacgg gcaacagctg attgcccttc
       61 accgcctggc cctgagagag ttgcagcaag cggtccacgc tggtttgccc cagcaggcga
      121 aaatcctgtt tgatggtggt taacggcggg atataacatg agctgtcttc ggtatcgtcg
      181 tatcccacta ccgagatatc cgcaccaacg cgcagcccgg actcggtaat ggcgcgcatt
      241 gcgcccagcg ccatctgatc gttggcaacc agcatcgcag tgggaacgat gccctcattc
      301 agcatttgca tggtttgttg aaaaccggac atggcactcc agtcgccttc ccgttccgct
      361 atcggctgaa tttgattgcg agtgagatat ttatgccagc cagccagacg cagacgcgcc
      421 gagacagaac ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa tgcgaccaga
      481 tgctccacgc ccagtcgcgt accgtcttca tgggagaaaa taatactgtt gatgggtgtc
      541 tggtcagaga catcaagaaa taacgccgga acattagtgc aggcagcttc cacagcaatg
      601 gcatcctggt catccagcgg atagttaatg atcagcccac tgacgcgttg cgcgagaaga
      661 ttgtgcaccg ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga caccaccacg
      721 ctggcaccca gttgatcggc gcgagattta atcgccgcga caatttgcga cggcgcgtgc
      781 agggccagac tggaggtggc aacgccaatc agcaacgact gtttgcccgc cagttgttgt
      841 gccacgcggt tgggaatgta attcagctcc gccatcgccg cttccacttt ttcccgcgtt
      901 ttcgcagaaa cgtggctggc ctggttcacc acgcgggaaa cggtctgata agagacaccg
      961 gcatactctg cgacatcgta taacgttact ggtttcacat tcaccaccct gaattgactc
     1021 tcttccgggc gctatcatgc cataccgcga aaggttttgc gccattcgat ggtgtccggg
     1081 atctcgacgc tctcccttat gcgactcctg cattaggaag cagcccagta gtaggttgag
     1141 gccgttgagc accgccgccg caaggaatgg tgcatgcaag gagatggcgc ccaacagtcc
     1201 cccggccacg gggcctgcca ccatacccac gccgaaacaa gcgctcatga gcccgaagtg
     1261 gcgagcccga tcttccccat cggtgatgtc ggcgatatag gcgccagcaa ccgcacctgt
     1321 ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg atcgagatct cgatcccgcg
     1381 aaattaatac gactcactat aggggaattg tgagcggata acaattcccc tctagaaata
     1441 attttgttta actttaagaa ggagatatac catgggcagc agccatcatc atcatcatca
     1501 cagcagcggc ctggtgccgc gcggcagcca taaaaataag caatcaaata cgcagaaccc
     1561 gaaccaacaa aatcaaaacg aagtgacagc taaagacatc ttaggaaatt caaaatattt
     1621 agcgatctct tacggaggat atcgaaaaaa atctcgagat tttcaaccct caattgaaga
     1681 gttaaaagaa gatatgaaaa ttcttcatgc gatgaatatt agaatattgc gcacctataa
     1741 tgtgcgtttg gcgcatacgt ctaatatttt aaaagccatt cgggagttga aaaatgaaga
     1801 tgccaatttc gaaatgtata tgatggttgg tgcttggatc gattgtaaaa atgcttggac
     1861 cgatcaaccg ttgaatcacc atgaagaaag cgagaacaat gcctctgaga ttgatcgtgc
     1921 agtcgctttg gcacaagaat ttccagatat tgtaaaagtc attgccgttg gaaatgaagc
     1981 catggtcaaa tgggccgcct cttattttgt gcaacccgct gtgattttaa aatgggtaaa
     2041 ccatttacaa gctttaaaga aaaaaggaga cctatccaaa gacttatgga tcaccagttc
     2101 cgataatttt gcttcttggg gtgggggaga tcctcaatac catgtggaag atctcacgaa
     2161 attaatcgag gctgtggatt atctatctgt acatacctat ccaatgcacg atacacatta
     2221 caatcctatt ttttggggcg ttttcgggga cgaaaccgaa ttgtcaagtt tgaaacgcat
     2281 tgatattgcc atgaaccgag ccaaaactta tgccgtttca caaagcgata gtgtggcaag
     2341 ttatataaag agcttgggca ttaacaaacc aatccatatt ggtgaaacag gttgggcaag
     2401 tttttctaac ggttattacg gggctaaagg ctctaaagca accgatgagt acaaggaggc
     2461 tattttttac aaccatattc gtgaatggac taatgaagct aatatgtcat gcttctattt
     2521 tgaagccttt gatgaaccat ggaaagatgc gcataactca ggaggttctg aaaaccattt
     2581 tggtttattt acggtggatg gtaaagcaaa atacgttctc tgggatttgg tagataaagg
     2641 cgtttttgaa ggattgacac gtggcgggaa tcccatcacc aaaacctata atggaaataa
     2701 agaagcacta ttccttgagg ttgaattgcc tccggtaaaa aaagaaatca caaagaacca
     2761 ttaactcgag caccaccacc accaccactg agatccggct gctaacaaag cccgaaagga
     2821 agctgagttg gctgctgcca ccgctgagca ataactagca taaccccttg gggcctctaa
     2881 acgggtcttg aggggttttt tgctgaaagg aggaactata tccggattgg cgaatgggac
     2941 gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct
     3001 acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg
     3061 ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt
     3121 gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca
     3181 tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga
     3241 ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa
     3301 gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac
     3361 gcgaatttta acaaaatatt aacgcttaca atttaggtgg cacttttcgg ggaaatgtgc
     3421 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgaatt
     3481 aattcttaga aaaactcatc gagcatcaaa tgaaactgca atttattcat atcaggatta
     3541 tcaataccat atttttgaaa aagccgtttc tgtaatgaag gagaaaactc accgaggcag
     3601 ttccatagga tggcaagatc ctggtatcgg tctgcgattc cgactcgtcc aacatcaata
     3661 caacctatta atttcccctc gtcaaaaata aggttatcaa gtgagaaatc accatgagtg
     3721 acgactgaat ccggtgagaa tggcaaaagt ttatgcattt ctttccagac ttgttcaaca
     3781 ggccagccat tacgctcgtc atcaaaatca ctcgcatcaa ccaaaccgtt attcattcgt
     3841 gattgcgcct gagcgagacg aaatacgcga tcgctgttaa aaggacaatt acaaacagga
     3901 atcgaatgca accggcgcag gaacactgcc agcgcatcaa caatattttc acctgaatca
     3961 ggatattctt ctaatacctg gaatgctgtt ttcccgggga tcgcagtggt gagtaaccat
     4021 gcatcatcag gagtacggat aaaatgcttg atggtcggaa gaggcataaa ttccgtcagc
     4081 cagtttagtc tgaccatctc atctgtaaca tcattggcaa cgctaccttt gccatgtttc
     4141 agaaacaact ctggcgcatc gggcttccca tacaatcgat agattgtcgc acctgattgc
     4201 ccgacattat cgcgagccca tttataccca tataaatcag catccatgtt ggaatttaat
     4261 cgcggcctag agcaagacgt ttcccgttga atatggctca taacacccct tgtattactg
     4321 tttatgtaag cagacagttt tattgttcat gaccaaaatc ccttaacgtg agttttcgtt
     4381 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     4441 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     4501 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     4561 aaatactgtc cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     4621 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     4681 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     4741 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     4801 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     4861 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     4921 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     4981 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     5041 cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt
     5101 ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga
     5161 gcgcagcgag tcagtgagcg aggaagcgga agagcgcctg atgcggtatt ttctccttac
     5221 gcatctgtgc ggtatttcac accgcaatgg tgcactctca gtacaatctg ctctgatgcc
     5281 gcatagttaa gccagtatac actccgctat cgctacgtga ctgggtcatg gctgcgcccc
     5341 gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt
     5401 acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac
     5461 cgaaacgcgc gaggcagctg cggtaaagct catcagcgtg gtcgtgaagc gattcacaga
     5521 tgtctgcctg ttcatccgcg tccagctcgt tgagtttctc cagaagcgtt aatgtctggc
     5581 ttctgataaa gcgggccatg ttaagggcgg ttttttcctg tttggtcact gatgcctccg
     5641 tgtaaggggg atttctgttc atgggggtaa tgataccgat gaaacgagag aggatgctca
     5701 cgatacgggt tactgatgat gaacatgccc ggttactgga acgttgtgag ggtaaacaac
     5761 tggcggtatg gatgcggcgg gaccagagaa aaatcactca gggtcaatgc cagcgcttcg
     5821 ttaatacaga tgtaggtgtt ccacagggta gccagcagca tcctgcgatg cagatccgga
     5881 acataatggt gcagggcgct gacttccgcg tttccagact ttacgaaaca cggaaaccga
     5941 agaccattca tgttgttgct caggtcgcag acgttttgca gcagcagtcg cttcacgttc
     6001 gctcgcgtat cggtgattca ttctgctaac cagtaaggca accccgccag cctagccggg
     6061 tcctcaacga caggagcacg atcatgcgca cccgtggggc cgccatgccg gcgataatgg
     6121 cctgcttctc gccgaaacgt ttggtggcgg gaccagtgac gaaggcttga gcgagggcgt
     6181 gcaagattcc gaataccgca agcgacaggc cgatcatcgt cgcgctccag cgaaagcggt
     6241 cctcgccgaa aatgacccag agcgctgccg gcacctgtcc tacgagttgc atgataaaga
     6301 agacagtcat aagtgcggcg acgatagtca tgccccgcgc ccaccggaag gagctgactg
     6361 ggttgaaggc tctcaagggc atcggtcgag atcccggtgc ctaatgagtg agctaactta
     6421 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
     6481 attaatgaat cggccaacgc gcggggagag gcggtttgcg ta
//
