LOCUS       sequence-464132-        5987 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Heterologous protein expression of cellulase B in Escherichia coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5987)
  TITLE     AI Proteins, Inc. plasmids
REFERENCE   2  (bases 1 to 5987)
  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"
COMMENT     Sequence Label: pAIP4_1UXZ
FEATURES             Location/Qualifiers
     source          1..5987
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(345..1427)
                     /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     1389..1408
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1428..1505)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1711..1730
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1818..1837
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1818..1836
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1837..1861
                     /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             1876..1898
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1921..1950
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             2665..2682
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2735..2753)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2749..2796
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2833..3288
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2920..2939)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3130..3151
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3380..4195)
                     /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(3380..4195)
                     /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     4103..4122
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4187..4206
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      4317..4905
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4806..4825
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5059..5076
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5091..5230
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5156..5175)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5316..5338
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5332..5523)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 gtggggccgc catgccggcg ataatggcct gcttctcgcc gaaacgtttg gtggcgggac
       61 cagtgacgaa ggcttgagcg agggcgtgca agattccgaa taccgcaagc gacaggccga
      121 tcatcgtcgc gctccagcga aagcggtcct cgccgaaaat gacccagagc gctgccggca
      181 cctgtcctac gagttgcatg ataaagaaga cagtcataag tgcggcgacg atagtcatgc
      241 cccgcgccca ccggaaggag ctgactgggt tgaaggctct caagggcatc ggtcgagatc
      301 ccggtgccta atgagtgagc taacttacat taattgcgtt gcgctcactg cccgctttcc
      361 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
      421 gtttgcgtat tgggcgccag ggtggttttt cttttcacca gtgagacggg caacagctga
      481 ttgcccttca ccgcctggcc ctgagagagt tgcagcaagc ggtccacgct ggtttgcccc
      541 agcaggcgaa aatcctgttt gatggtggtt aacggcggga tataacatga gctgtcttcg
      601 gtatcgtcgt atcccactac cgagatgtcc gcaccaacgc gcagcccgga ctcggtaatg
      661 gcgcgcattg cgcccagcgc catctgatcg ttggcaacca gcatcgcagt gggaacgatg
      721 ccctcattca gcatttgcat ggtttgttga aaaccggaca tggcactcca gtcgccttcc
      781 cgttccgcta tcggctgaat ttgattgcga gtgagatatt tatgccagcc agccagacgc
      841 agacgcgccg agacagaact taatgggccc gctaacagcg cgatttgctg gtgacccaat
      901 gcgaccagat gctccacgcc cagtcgcgta ccgtcttcat gggagaaaat aatactgttg
      961 atgggtgtct ggtcagagac atcaagaaat aacgccggaa cattagtgca ggcagcttcc
     1021 acagcaatgg catcctggtc atccagcgga tagttaatga tcagcccact gacgcgttgc
     1081 gcgagaagat tgtgcaccgc cgctttacag gcttcgacgc cgcttcgttc taccatcgac
     1141 accaccacgc tggcacccag ttgatcggcg cgagatttaa tcgccgcgac aatttgcgac
     1201 ggcgcgtgca gggccagact ggaggtggca acgccaatca gcaacgactg tttgcccgcc
     1261 agttgttgtg ccacgcggtt gggaatgtaa ttcagctccg ccatcgccgc ttccactttt
     1321 tcccgcgttt tcgcagaaac gtggctggcc tggttcacca cgcgggaaac ggtctgataa
     1381 gagacaccgg catactctgc gacatcgtat aacgttactg gtttcacatt caccaccctg
     1441 aattgactct cttccgggcg ctatcatgcc ataccgcgaa aggttttgcg ccattcgatg
     1501 gtgtccggga tctcgacgct ctcccttatg cgactcctgc attaggaagc agcccagtag
     1561 taggttgagg ccgttgagca ccgccgccgc aaggaatggt gcatgcaagg agatggcgcc
     1621 caacagtccc ccggccacgg ggcctgccac catacccacg ccgaaacaag cgctcatgag
     1681 cccgaagtgg cgagcccgat cttccccatc ggtgatgtcg gcgatatagg cgccagcaac
     1741 cgcacctgtg gcgccggtga tgccggccac gatgcgtccg gcgtagagga tcgagatcga
     1801 tctcgatccc gcgaaattaa tacgactcac tataggggaa ttgtgagcgg ataacaattc
     1861 ccctctagaa ataattttgt ttaactttaa gaaggagata tacatatgat ggcgaagatt
     1921 catcaccatc atcaccatca ccatcaccac ggtagcctgc aagattcgga agtcaaccaa
     1981 gaagcaaagc cggaagtcaa gccggaagtg aaaccggaaa cccatattaa cctgaaagtt
     2041 agtgacggca gctctgaaat tttctttaag atcaaaaaga ccacgccgct gcgtcgcctg
     2101 atggaagcgt ttgccaaacg tcagggcaag gaaatggata gcctgcgttt cctgtatgac
     2161 ggtattcgca tccaggcaga tcaagcgccg gaagacctgg acatggaaga caacgacatc
     2221 attgaagccc accgtgaaca gattggcggc tctggaatgg taatagcaac aattcaagct
     2281 gaagatcaca gccaacaaag cggtacgcag caagaaacca ccacggatac cggtggcggt
     2341 aaaaacgttg gttatatcga cgctggcgac tggctgtcgt acgcgggtac tccggtgaac
     2401 atcccatcca gcggttctta cctgattgaa tatcgtgttg cgagccagaa tggtggaggt
     2461 tctctgacct tcgaggaggc gggcggcgca ccggtgcacg gcactatcgc catcccggcg
     2521 accggtggct ggcagacctg gaccacgatc cagcataccg tcaacttaag cgcaggctcc
     2581 catcagtttg gtattaaggc taatgcgggc ggctggaact tgaattggat tcgcattaac
     2641 aaaaccggtt cgtaatgact cgagcaccac caccaccacc actgagatcc ggctgctaac
     2701 aaagcccgaa aggaagctga gttggctgct gccaccgctg agcaataact agcataaccc
     2761 cttggggcct ctaaacgggt cttgaggggt tttttgctga aaggaggaac tatatccgga
     2821 ttggcgaatg ggacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
     2881 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt
     2941 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag
     3001 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt
     3061 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
     3121 tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt
     3181 cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt
     3241 aacaaaaatt taacgcgaat tttaacaaaa tattaacgct tacaatttag gtggcacttt
     3301 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta
     3361 tccgctcatg aattaattct tagaaaaact catcgagcat caaatgaaac tgcaatttat
     3421 tcatatcagg attatcaata ccatattttt gaaaaagccg tttctgtaat gaaggagaaa
     3481 actcaccgag gcagttccat aggatggcaa gatcctggta tcggtctgcg attccgactc
     3541 gtccaacatc aatacaacct attaatttcc cctcgtcaaa aataaggtta tcaagtgaga
     3601 aatcaccatg agtgacgact gaatccggtg agaatggcaa aagtttatgc atttctttcc
     3661 agacttgttc aacaggccag ccattacgct cgtcatcaaa atcactcgca tcaaccaaac
     3721 cgttattcat tcgtgattgc gcctgagcga gacgaaatac gcgatcgctg ttaaaaggac
     3781 aattacaaac aggaatcgaa tgcaaccggc gcaggaacac tgccagcgca tcaacaatat
     3841 tttcacctga atcaggatat tcttctaata cctggaatgc tgttttcccg gggatcgcag
     3901 tggtgagtaa ccatgcatca tcaggagtac ggataaaatg cttgatggtc ggaagaggca
     3961 taaattccgt cagccagttt agtctgacca tctcatctgt aacatcattg gcaacgctac
     4021 ctttgccatg tttcagaaac aactctggcg catcgggctt cccatacaat cgatagattg
     4081 tcgcacctga ttgcccgaca ttatcgcgag cccatttata cccatataaa tcagcatcca
     4141 tgttggaatt taatcgcggc ctagagcaag acgtttcccg ttgaatatgg ctcataacac
     4201 cccttgtatt actgtttatg taagcagaca gttttattgt tcatgaccaa aatcccttaa
     4261 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     4321 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     4381 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     4441 agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag
     4501 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     4561 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     4621 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     4681 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     4741 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     4801 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     4861 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     4921 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     4981 tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc
     5041 agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cctgatgcgg
     5101 tattttctcc ttacgcatct gtgcggtatt tcacaccgca atggtgcact ctcagtacaa
     5161 tctgctctga tgccgcatag ttaagccagt atacactccg ctatcgctac gtgactgggt
     5221 catggctgcg ccccgacacc cgccaacacc cgctgacgcg ccctgacggg cttgtctgct
     5281 cccggcatcc gcttacagac aagctgtgac cgtctccggg agctgcatgt gtcagaggtt
     5341 ttcaccgtca tcaccgaaac gcgcgaggca gctgcggtaa agctcatcag cgtggtcgtg
     5401 aagcgattca cagatgtctg cctgttcatc cgcgtccagc tcgttgagtt tctccagaag
     5461 cgttaatgtc tggcttctga taaagcgggc catgttaagg gcggtttttt cctgtttggt
     5521 cactgatgcc tccgtgtaag ggggatttct gttcatgggg gtaatgatac cgatgaaacg
     5581 agagaggatg ctcacgatac gggttactga tgatgaacat gcccggttac tggaacgttg
     5641 tgagggtaaa caactggcgg tatggatgcg gcgggaccag agaaaaatca ctcagggtca
     5701 atgccagcgc ttcgttaata cagatgtagg tgttccacag ggtagccagc agcatcctgc
     5761 gatgcagatc cggaacataa tggtgcaggg cgctgacttc cgcgtttcca gactttacga
     5821 aacacggaaa ccgaagacca ttcatgttgt tgctcaggtc gcagacgttt tgcagcagca
     5881 gtcgcttcac gttcgctcgc gtatcggtga ttcattctgc taaccagtaa ggcaaccccg
     5941 ccagcctagc cgggtcctca acgacaggag cacgatcatg cgcaccc
//
