LOCUS       sequence-460500-        8822 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Recombinant Ag43 protein fused to silaffin-R5 and mCherry. pelB as 
            the periplasmic signal. His-tag for purification. TEV enzyme 
            recognition site between Ag43 alpha subunit and the fluorescent 
            protein..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8822)
  AUTHORS   Kirpat Konak BM, Bakar ME, Ahan RE, Ozyurek EU, Dokmeci S, Safak 
            Seker UO
  TITLE     A living material platform for the biomineralization of biosilica.
  JOURNAL   Mater Today Bio. 2022 Oct 10;17:100461. doi: 
            10.1016/j.mtbio.2022.100461. eCollection 2022 Dec 15.
  PUBMED    36278145
REFERENCE   2  (bases 1 to 8822)
  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: "Mater Today
            Bio."; date: "2022-10-10"; volume: "17:100461. doi"; pages: " 
            10.1016/j.mtbio.2022.100461. eCollection 2022 Dec 15"
COMMENT     Sequence Label: pET22b Ag43 160N 6H mCherry-R5 AmpR
FEATURES             Location/Qualifiers
     source          1..8822
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(26..73)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     69..87
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(140..157)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(2672..2692)
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             complement(3533..3550)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     sig_peptide     complement(3557..3622)
                     /codon_start=1
                     /gene="pelB (fragment)"
                     /product="leader peptide for secretion"
                     /label=pelB signal sequence
                     /translation="MKYLLPTAAAGLLLLAAQPAMA"
     RBS             3630..3652
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    3667..3691
                     /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)."
     primer_bind     complement(3691..3710)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3692..3710)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3794..3813)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        4019..4096
                     /gene="lacI"
                     /label=lacI promoter
     CDS             4097..5179
                     /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     complement(4116..4135)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             5988..6179
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(6173..6195)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    6281..6420
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     6336..6355
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(6435..6452)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6606..7194)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6686..6705)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7365..8156,8157..8225))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     7988..8007
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8226..8330)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(8356..8811)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(8493..8514)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     8705..8724
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
       61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
      121 tgttagcagc cggatctcag tggtggtggt ggtggtgctc gagtcatcag aaggtcacat
      181 tcagtgtggc ctgaccgtta tacccttcag cgctgctgcc gctgacgctg tgggcataac
      241 cggcctgaac gcccagggtg atattttccc ggacacgggc ttccagtccg gcctgcaggt
      301 ccagtgatgt gccattctgt gagggagaga acgtcatccc gctgcctgca gtggaagtcc
      361 ccacacgcat atctccccgg gagctgaagg tgcggataac agaaggctgt acccaccagt
      421 tcaccggtaa ttcactcaca ctgtgttttg cactgtcacg caggggggca cgggatgagg
      481 tgccttcgcc aaaggtcata tcgttgtggc tgcccagacg gaaaccggca cgcacatgtt
      541 gtgcactgcc atgcccgaac ttcacataac cggcgttgtc cttaccgtca tccagggaaa
      601 gtccctgcca ggtatactgc agttgtggct ccagcatcag gttgtcagtg atactgaagg
      661 gcagaccggt ttccagtgag cccagccagc cccagccccg ggcgcggaag tcgttattgt
      721 ccgatgacgc tttcatgctg tggcgggttc cctgtgccac aatgtcagcc cacaggccgg
      781 aggacgtgtg taccagattc aggtatccgc ccaggctgcc ggcatcatcc cggaccgtgc
      841 cggcacggga gccgtcatca tccttaacat caacggaaga atggccagca gcaccatata
      901 cccccgcggt cacagacata ccggcaacct ctgttctcat caggtcaccc tccagacgga
      961 cgaatccata gctgccgctg ctttccggcg tggccccacg ggcaataccg ccattgttat
     1021 cgtgaccgag atgaccgccc tgaatgctga gacggacgct gttgttttca ccatttacac
     1081 cggtctgatg gctgcgggag cctgccacaa tccggtcata gtccattgcc tgtgtcagca
     1141 tggaggcata cagggggact tctgcacgat aagcattttc actgcgcaga taccagctct
     1201 catcactgtc ccggttgagg gagtagttaa aggcaccggc ctgcagcctg ttcccctgga
     1261 caaaggcccc ttcctccgtg gtggcaccgt taatggcttc caccacctga atacccttac
     1321 cgctggtcgc cagccccgac gcactgttgc cggcgttcac caggttcagg atggtttttc
     1381 cggttgccct gccgccgtca atgaccagtc tgtcagcatt gttctgtgcc atatccgggc
     1441 gtacacgcag gctgatggtg ccattctgtc cgttcaggtt tttcactttc agggttgccg
     1501 gtacgaactt ccctgtgcgg gtggaggtga aatgaatctg tccggcatgg ctgaggtcat
     1561 ccaccaccga ctgcaccgtg gcgttatcgg ggatattcca ggtggcaccg gaggcgagag
     1621 tgacattcgt ggggtcaatg gcaccgttca gcacagtgct gccggtcagc ttcagggctg
     1681 taccgcgctg agcaatgaca tccgtggtga cggtactgtc gttcagcgtc agcgtgcctt
     1741 cattcaggtt gacggctttc tgggtgagtg tggtgttgct gacagtgagt gtgccacttc
     1801 ctgatttttc cacgctgccg ttaccggtga gagagcctcc ctgcaggagt gcatcgcctt
     1861 cacggatggt cagggtatcg ttgttcaccg tcttcccgga aagggtaagt atggcgccgt
     1921 tattcagcgt ggtggtgccc gccagtgtgc cgcccctggc ggtgaacagt ccgccattta
     1981 ccgtggtatc cgtggccgta tcaccggcgt tcagcgtgaa tgaactgcct ttttccagca
     2041 tcagggcatc cgcctgaccg cctccgatac tgaatgcctt tccgtcgctc cgggtaccac
     2101 tgacagcggc accggaatcg gccagcagta caccgccatt tcccatggat acggtggtgg
     2161 cggtgccacc gttgcggaca tccagcgttc cgccatcatc cacgcgggta ttagtggctg
     2221 tgtgtccggt cagcacatcc aggcgtccgc cattttccag tacgacatta tcagctttac
     2281 cctccacaac agagaatgct cccaggcggt ttatgccggt aacggttgca gccgtactgg
     2341 taaccagtgc gccgccctgc ttcagggtga cattcgtggc tgtaccaccg gcgtccacct
     2401 gcagtctgcc cttctgatta acggtggtat tcccggccac acccccgttt ttgacaatct
     2461 gccagccgtc actgttcaca acagtgtcag acgctgtacc gccgttgtgc acatactggt
     2521 atcccccatt cagcgtggta tccagtgcgt gaccatgtac agtctggtcg ccgccggcat
     2581 aaaccacagt ggtatttgcc gttccttcag ctctcacaat ctggcgaccg tttttattga
     2641 tggttgtgcg cttaaggcta ccaccgccac cgccctgaaa gtacaggttt tcgctaccac
     2701 cgccacccag aatgcggcgt ttgctgcctt tgctgccgct atagctgccg ctttttttgc
     2761 tgctagatcc tcccccaccg ctaccgccgc cgccactgcc accgccacct ttgtacagct
     2821 catccatgcc accggtagaa tgacgaccct ccgcgcgctc atattgctct acgatcgtat
     2881 aatcttcatt atgagaggtg atgtccagtt taatattcac attgtacgcg ccaggcagct
     2941 gcacaggttt cttggctttg tacgtggttt tcacttcagc gtcataatgg ccgccatctt
     3001 tcagtttcag gcgctgttta atttcgcctt tcagcgcacc atcttccgga tacatgcgtt
     3061 cgctagacgc ctcccaaccc atcgtctttt tctgcatcac cgggccatca gatggaaaat
     3121 tagtaccacg cagtttaact ttatagatga actcgccatc ctgcagggag gagtcctgag
     3181 tgacggtcac gacaccacca tcttcaaaat tcattacgcg ttcccacttg aaaccttccg
     3241 gaaaagacag cttcagatag tccgggatat ccgctgggtg tttaacatac gctttagaac
     3301 cgtacataaa ttgcgggctc aggatgtccc acgcaaaagg cagcgggccg cctttagtca
     3361 ctttcagttt ggcggtctgg gtgccttcat acggacggcc ctcgccttcg ccttcgatct
     3421 cgaactcgtg accgttaaca gaaccctcca tgtgaacttt gaagcgcatg aactctttaa
     3481 tgatagccat gttatcctcc tcgcccttgg aaaccatgga tccaccgcca ccgtggtggt
     3541 ggtggtggtg gcccatggcc atcgccggct gggcagcgag gagcagcaga ccagcagcag
     3601 cggtcggcag caggtatttc atatgtatat ctccttctta aagttaaaca aaattatttc
     3661 tagaggggaa ttgttatccg ctcacaattc ccctatagtg agtcgtatta atttcgcggg
     3721 atcgagatct cgatcctcta cgccggacgc atcgtggccg gcatcaccgg cgccacaggt
     3781 gcggttgctg gcgcctatat cgccgacatc accgatgggg aagatcgggc tcgccacttc
     3841 gggctcatga gcgcttgttt cggcgtgggt atggtggcag gccccgtggc cgggggactg
     3901 ttgggcgcca tctccttgca tgcaccattc cttgcggcgg cggtgctcaa cggcctcaac
     3961 ctactactgg gctgcttcct aatgcaggag tcgcataagg gagagcgtcg agatcccgga
     4021 caccatcgaa tggcgcaaaa cctttcgcgg tatggcatga tagcgcccgg aagagagtca
     4081 attcagggtg gtgaatgtga aaccagtaac gttatacgat gtcgcagagt atgccggtgt
     4141 ctcttatcag accgtttccc gcgtggtgaa ccaggccagc cacgtttctg cgaaaacgcg
     4201 ggaaaaagtg gaagcggcga tggcggagct gaattacatt cccaaccgcg tggcacaaca
     4261 actggcgggc aaacagtcgt tgctgattgg cgttgccacc tccagtctgg ccctgcacgc
     4321 gccgtcgcaa attgtcgcgg cgattaaatc tcgcgccgat caactgggtg ccagcgtggt
     4381 ggtgtcgatg gtagaacgaa gcggcgtcga agcctgtaaa gcggcggtgc acaatcttct
     4441 cgcgcaacgc gtcagtgggc tgatcattaa ctatccgctg gatgaccagg atgccattgc
     4501 tgtggaagct gcctgcacta atgttccggc gttatttctt gatgtctctg accagacacc
     4561 catcaacagt attattttct cccatgaaga cggtacgcga ctgggcgtgg agcatctggt
     4621 cgcattgggt caccagcaaa tcgcgctgtt agcgggccca ttaagttctg tctcggcgcg
     4681 tctgcgtctg gctggctggc ataaatatct cactcgcaat caaattcagc cgatagcgga
     4741 acgggaaggc gactggagtg ccatgtccgg ttttcaacaa accatgcaaa tgctgaatga
     4801 gggcatcgtt cccactgcga tgctggttgc caacgatcag atggcgctgg gcgcaatgcg
     4861 cgccattacc gagtccgggc tgcgcgttgg tgcggatatc tcggtagtgg gatacgacga
     4921 taccgaagac agctcatgtt atatcccgcc gttaaccacc atcaaacagg attttcgcct
     4981 gctggggcaa accagcgtgg accgcttgct gcaactctct cagggccagg cggtgaaggg
     5041 caatcagctg ttgcccgtct cactggtgaa aagaaaaacc accctggcgc ccaatacgca
     5101 aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg
     5161 actggaaagc gggcagtgag cgcaacgcaa ttaatgtaag ttagctcact cattaggcac
     5221 cgggatctcg accgatgccc ttgagagcct tcaacccagt cagctccttc cggtgggcgc
     5281 ggggcatgac tatcgtcgcc gcacttatga ctgtcttctt tatcatgcaa ctcgtaggac
     5341 aggtgccggc agcgctctgg gtcattttcg gcgaggaccg ctttcgctgg agcgcgacga
     5401 tgatcggcct gtcgcttgcg gtattcggaa tcttgcacgc cctcgctcaa gccttcgtca
     5461 ctggtcccgc caccaaacgt ttcggcgaga agcaggccat tatcgccggc atggcggccc
     5521 cacgggtgcg catgatcgtg ctcctgtcgt tgaggacccg gctaggctgg cggggttgcc
     5581 ttactggtta gcagaatgaa tcaccgatac gcgagcgaac gtgaagcgac tgctgctgca
     5641 aaacgtctgc gacctgagca acaacatgaa tggtcttcgg tttccgtgtt tcgtaaagtc
     5701 tggaaacgcg gaagtcagcg ccctgcacca ttatgttccg gatctgcatc gcaggatgct
     5761 gctggctacc ctgtggaaca cctacatctg tattaacgaa gcgctggcat tgaccctgag
     5821 tgatttttct ctggtcccgc cgcatccata ccgccagttg tttaccctca caacgttcca
     5881 gtaaccgggc atgttcatca tcagtaaccc gtatcgtgag catcctctct cgtttcatcg
     5941 gtatcattac ccccatgaac agaaatcccc cttacacgga ggcatcagtg accaaacagg
     6001 aaaaaaccgc ccttaacatg gcccgcttta tcagaagcca gacattaacg cttctggaga
     6061 aactcaacga gctggacgcg gatgaacagg cagacatctg tgaatcgctt cacgaccacg
     6121 ctgatgagct ttaccgcagc tgcctcgcgc gtttcggtga tgacggtgaa aacctctgac
     6181 acatgcagct cccggagacg gtcacagctt gtctgtaagc ggatgccggg agcagacaag
     6241 cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg cgcagccatg acccagtcac
     6301 gtagcgatag cggagtgtat actggcttaa ctatgcggca tcagagcaga ttgtactgag
     6361 agtgcaccat tgcggtgtga aataccgcac agatgcgtaa ggagaaaata ccgcatcagg
     6421 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
     6481 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     6541 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     6601 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     6661 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     6721 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     6781 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     6841 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     6901 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     6961 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     7021 tggcctaact acggctacac tagaaggaca gtatttggta tctgcgctct gctgaagcca
     7081 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     7141 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     7201 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     7261 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     7321 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     7381 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     7441 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     7501 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     7561 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     7621 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     7681 gcaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     7741 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     7801 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     7861 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     7921 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     7981 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     8041 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     8101 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     8161 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     8221 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     8281 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     8341 cgaaaagtgc cacctaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt
     8401 gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa
     8461 aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa
     8521 agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac
     8581 gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga
     8641 accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa
     8701 aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc
     8761 tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg tcccattcgc
     8821 ca
//
