LOCUS       sequence-431794-        6401 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of a His-tagged Chitin Binding fusion of 
            Cordyceps militaris EndoCOM.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6401)
  TITLE     Deglycosylation Toolkit
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6401)
  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            "Unpublished"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: pET-29b(+)_notufA-CmEndoCOM_3C-CBD-His
FEATURES             Location/Qualifiers
     source          1..6401
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             299..322
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             335..490
                     /codon_start=1
                     /gene="Bacillus circulans chiA"
                     /product="chitin binding domain from chitinase A1 (Watanabe
                     et al., 1994)"
                     /label=CBD
                     /translation="TTNPGVSAWQVNTAYTAGQLVTYNGKTYKCLQPHTSLAGWEPSNV
                     PALWQLQ"
     CDS             503..532
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             545..562
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(615..633)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      629..676
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      713..1168
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(800..819)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1010..1031
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1260..2075)
                     /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(1260..2075)
                     /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     1983..2002
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2067..2086
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2197..2785
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2686..2705
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2939..2956
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2971..3110
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3036..3055)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3196..3218
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3212..3403)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4212..5294)
                     /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     5256..5275
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5295..5372)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5578..5597
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     5685..5704
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5685..5703
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5704..5728
                     /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             5743..5765
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
ORIGIN
        1 ctataattcc ttcaaaagtt acatctcgtc gaaacagtat cttatcgggt ttagtttcta
       61 cgaggaaaac ggcacaaatt ggggagacac tacaacacca atgacaagca gccgcgcctg
      121 gcaatacgca aaatggcaac catccggggc tacaaaggga gggatttttt catacgctat
      181 cgaccgcgac ggagttgcca tcggcgataa tactctgaaa acaacagatt ttacgtggac
      241 ccgtcaactt atcggggcga tgaatccggg atccgactac gatattccga ctaccggatt
      301 ggaggtattg ttccaaggtc cagccaactc aggcaccacg aatccaggcg tgtcagcctg
      361 gcaagtgaac accgcctaca cagcagggca gttggtcacg tataacggca aaacgtataa
      421 atgccttcaa cctcacacaa gtcttgccgg ttgggagcca tccaacgtac ccgccttatg
      481 gcaactgcaa tctggtaacg gacaccatca tcatcatcat caccatcacc attaatagct
      541 cgagcaccac caccaccacc actgagatcc ggctgctaac aaagcccgaa aggaagctga
      601 gttggctgct gccaccgctg agcaataact agcataaccc cttggggcct ctaaacgggt
      661 cttgaggggt tttttgctga aaggaggaac tatatccgga ttggcgaatg ggacgcgccc
      721 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
      781 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
      841 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
      901 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc
      961 tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg
     1021 ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt
     1081 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     1141 tttaacaaaa tattaacgct tacaatttag gtggcacttt tcggggaaat gtgcgcggaa
     1201 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg aattaattct
     1261 tagaaaaact catcgagcat caaatgaaac tgcaatttat tcatatcagg attatcaata
     1321 ccatattttt gaaaaagccg tttctgtaat gaaggagaaa actcaccgag gcagttccat
     1381 aggatggcaa gatcctggta tcggtctgcg attccgactc gtccaacatc aatacaacct
     1441 attaatttcc cctcgtcaaa aataaggtta tcaagtgaga aatcaccatg agtgacgact
     1501 gaatccggtg agaatggcaa aagtttatgc atttctttcc agacttgttc aacaggccag
     1561 ccattacgct cgtcatcaaa atcactcgca tcaaccaaac cgttattcat tcgtgattgc
     1621 gcctgagcga gacgaaatac gcgatcgctg ttaaaaggac aattacaaac aggaatcgaa
     1681 tgcaaccggc gcaggaacac tgccagcgca tcaacaatat tttcacctga atcaggatat
     1741 tcttctaata cctggaatgc tgttttcccg gggatcgcag tggtgagtaa ccatgcatca
     1801 tcaggagtac ggataaaatg cttgatggtc ggaagaggca taaattccgt cagccagttt
     1861 agtctgacca tctcatctgt aacatcattg gcaacgctac ctttgccatg tttcagaaac
     1921 aactctggcg catcgggctt cccatacaat cgatagattg tcgcacctga ttgcccgaca
     1981 ttatcgcgag cccatttata cccatataaa tcagcatcca tgttggaatt taatcgcggc
     2041 ctagagcaag acgtttcccg ttgaatatgg ctcataacac cccttgtatt actgtttatg
     2101 taagcagaca gttttattgt tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     2161 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     2221 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     2281 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     2341 tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     2401 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     2461 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     2521 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     2581 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     2641 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     2701 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     2761 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     2821 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
     2881 ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag
     2941 cgagtcagtg agcgaggaag cggaagagcg cctgatgcgg tattttctcc ttacgcatct
     3001 gtgcggtatt tcacaccgca atggtgcact ctcagtacaa tctgctctga tgccgcatag
     3061 ttaagccagt atacactccg ctatcgctac gtgactgggt catggctgcg ccccgacacc
     3121 cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac
     3181 aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac
     3241 gcgcgaggca gctgcggtaa agctcatcag cgtggtcgtg aagcgattca cagatgtctg
     3301 cctgttcatc cgcgtccagc tcgttgagtt tctccagaag cgttaatgtc tggcttctga
     3361 taaagcgggc catgttaagg gcggtttttt cctgtttggt cactgatgcc tccgtgtaag
     3421 ggggatttct gttcatgggg gtaatgatac cgatgaaacg agagaggatg ctcacgatac
     3481 gggttactga tgatgaacat gcccggttac tggaacgttg tgagggtaaa caactggcgg
     3541 tatggatgcg gcgggaccag agaaaaatca ctcagggtca atgccagcgc ttcgttaata
     3601 cagatgtagg tgttccacag ggtagccagc agcatcctgc gatgcagatc cggaacataa
     3661 tggtgcaggg cgctgacttc cgcgtttcca gactttacga aacacggaaa ccgaagacca
     3721 ttcatgttgt tgctcaggtc gcagacgttt tgcagcagca gtcgcttcac gttcgctcgc
     3781 gtatcggtga ttcattctgc taaccagtaa ggcaaccccg ccagcctagc cgggtcctca
     3841 acgacaggag cacgatcatg cgcacccgtg gggccgccat gccggcgata atggcctgct
     3901 tctcgccgaa acgtttggtg gcgggaccag tgacgaaggc ttgagcgagg gcgtgcaaga
     3961 ttccgaatac cgcaagcgac aggccgatca tcgtcgcgct ccagcgaaag cggtcctcgc
     4021 cgaaaatgac ccagagcgct gccggcacct gtcctacgag ttgcatgata aagaagacag
     4081 tcataagtgc ggcgacgata gtcatgcccc gcgcccaccg gaaggagctg actgggttga
     4141 aggctctcaa gggcatcggt cgagatcccg gtgcctaatg agtgagctaa cttacattaa
     4201 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
     4261 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgccagggt ggtttttctt
     4321 ttcaccagtg agacgggcaa cagctgattg cccttcaccg cctggccctg agagagttgc
     4381 agcaagcggt ccacgctggt ttgccccagc aggcgaaaat cctgtttgat ggtggttaac
     4441 ggcgggatat aacatgagct gtcttcggta tcgtcgtatc ccactaccga gatgtccgca
     4501 ccaacgcgca gcccggactc ggtaatggcg cgcattgcgc ccagcgccat ctgatcgttg
     4561 gcaaccagca tcgcagtggg aacgatgccc tcattcagca tttgcatggt ttgttgaaaa
     4621 ccggacatgg cactccagtc gccttcccgt tccgctatcg gctgaatttg attgcgagtg
     4681 agatatttat gccagccagc cagacgcaga cgcgccgaga cagaacttaa tgggcccgct
     4741 aacagcgcga tttgctggtg acccaatgcg accagatgct ccacgcccag tcgcgtaccg
     4801 tcttcatggg agaaaataat actgttgatg ggtgtctggt cagagacatc aagaaataac
     4861 gccggaacat tagtgcaggc agcttccaca gcaatggcat cctggtcatc cagcggatag
     4921 ttaatgatca gcccactgac gcgttgcgcg agaagattgt gcaccgccgc tttacaggct
     4981 tcgacgccgc ttcgttctac catcgacacc accacgctgg cacccagttg atcggcgcga
     5041 gatttaatcg ccgcgacaat ttgcgacggc gcgtgcaggg ccagactgga ggtggcaacg
     5101 ccaatcagca acgactgttt gcccgccagt tgttgtgcca cgcggttggg aatgtaattc
     5161 agctccgcca tcgccgcttc cactttttcc cgcgttttcg cagaaacgtg gctggcctgg
     5221 ttcaccacgc gggaaacggt ctgataagag acaccggcat actctgcgac atcgtataac
     5281 gttactggtt tcacattcac caccctgaat tgactctctt ccgggcgcta tcatgccata
     5341 ccgcgaaagg ttttgcgcca ttcgatggtg tccgggatct cgacgctctc ccttatgcga
     5401 ctcctgcatt aggaagcagc ccagtagtag gttgaggccg ttgagcaccg ccgccgcaag
     5461 gaatggtgca tgcaaggaga tggcgcccaa cagtcccccg gccacggggc ctgccaccat
     5521 acccacgccg aaacaagcgc tcatgagccc gaagtggcga gcccgatctt ccccatcggt
     5581 gatgtcggcg atataggcgc cagcaaccgc acctgtggcg ccggtgatgc cggccacgat
     5641 gcgtccggcg tagaggatcg agatcgatct cgatcccgcg aaattaatac gactcactat
     5701 aggggaattg tgagcggata acaattcccc tctagaaata attttgttta actttaagaa
     5761 ggagatatac atatgaaagg ttcttcttcc gtgtgcccta gtgataatac ccatgcgaca
     5821 ggggcggcat tacaaaagat cttagattac aagaaagggg atcatcaaat catggccggt
     5881 tattttcgtt cgtggcgtga cacagcctcg ggtacgggta ataaggtgtc catgttggac
     5941 ttgcccgact gtctggacat cgcgtttgtg ttccccgagg gagatgagac cgcctccttc
     6001 tggacaacac ttaaagatac ctatgtccct gcgcttcacg gtcgtggtat caaagtggta
     6061 cgcagtgtag gtattgcaca attgattaac acagcctggg acaatacacc ggcaggttgg
     6121 caaggtttag ctgacgctct tatgaagacc gtagacgatt atggtttaga tggattggac
     6181 atcgatgtag agcagagtct taacgccaat cagctgaagc aagcgacagg cgtatttaac
     6241 gctcttgcaa aaaaacttgg tccgaagagt ggaacgggca agcttctgat tttcgatacg
     6301 aacatggatg gcactcaacc gttgtggcgc aacgtgtatc cgacaatctc ttacgttttg
     6361 attcaatcat acggccgttc catttctggc ttacagacga c
//
