LOCUS       sequence-139434-        3817 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Vector for SICLOPPS-mediated protein circularization using the 
            Nostoc punctiforme DnaE intein..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3817)
  AUTHORS   Waldhauer MC, Schmitz SN, Ahlmann-Eltze C, Gleixner JG, Schmelas CC,
            Huhn AG, Bunne C, Buscher M, Horn M, Klughammer N, Kreft J, Schafer 
            E, Bayer PA, Kramer SG, Neugebauer J, Wehler P, Mayer MP, Eils R, Di
            Ventura B
  TITLE     Backbone circularization of Bacillus subtilis family 11 xylanase 
            increases its thermostability and its resistance against 
            aggregation.
  JOURNAL   Mol Biosyst. 2015 Nov 10;11(12):3231-43. doi: 10.1039/c5mb00341e.
  PUBMED    26434634
REFERENCE   2  (bases 1 to 3817)
  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"; doi: "10"; journalName: 
            "Mol Biosyst"; date: "2015-11-10- 10"; volume: "11"; issue: "12"; 
            pages: "3231-43"
COMMENT     Sequence Label: pCIRCNpu
FEATURES             Location/Qualifiers
     source          1..3817
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    20..44
                     /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)."
     misc_feature    47..68
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
     RBS             75..97
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    319..340
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(355..360,361..378,379..385)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    393..409
                     /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             427..438
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             445..1122
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     terminator      1167..1238
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1254..1281
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     misc_feature    1631..1651
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     terminator      1652..1709
                     /label=his operon terminator
                     /note="This putative transcriptin terminator from the E. 
                     coli his operon has a 2-bp deletion introduced during 
                     synthesis. Its efficiency has not been determined."
     primer_bind     complement(1733..1750)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1904..2492)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1984..2003)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2725..3540)
                     /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(2725..3540)
                     /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     3448..3467
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3532..3551
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     3745..3763
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     terminator      complement(3767..3810)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttccccgaat tcgcggccgc
       61 ttctagagaa taattttgtt taactttaag aaggagatac tagatgatca aaatagccac
      121 acgtaaatat ttaggcaaac aaaatgtcta tgacattgga gttgagcgcg accataattt
      181 tgcactcaaa aatggcttca tagcttccaa cagagaccca atacgcaaac cgcctctccc
      241 cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg tttcccgact ggaaagcggg
      301 cagtgagcgc aacgcaatta atgtgagtta gctcactcat taggcacccc aggctttaca
      361 ctttatgctt ccggctcgta tgttgtgtgg aattgtgagc ggataacaat ttcacacata
      421 ctagagaaag aggagaaata ctagatggct tcctccgaag acgttatcaa agagttcatg
      481 cgtttcaaag ttcgtatgga aggttccgtt aacggtcacg agttcgaaat cgaaggtgaa
      541 ggtgaaggtc gtccgtacga aggtacccag accgctaaac tgaaagttac caaaggtggt
      601 ccgctgccgt tcgcttggga catcctgtcc ccgcagttcc agtacggttc caaagcttac
      661 gttaaacacc cggctgacat cccggactac ctgaaactgt ccttcccgga aggtttcaaa
      721 tgggaacgtg ttatgaactt cgaagacggt ggtgttgtta ccgttaccca ggactcctcc
      781 ctgcaagacg gtgagttcat ctacaaagtt aaactgcgtg gtaccaactt cccgtccgac
      841 ggtccggtta tgcagaaaaa aaccatgggt tgggaagctt ccaccgaacg tatgtacccg
      901 gaagacggtg ctctgaaagg tgaaatcaaa atgcgtctga aactgaaaga cggtggtcac
      961 tacgacgctg aagttaaaac cacctacatg gctaaaaaac cggttcagct gccgggtgct
     1021 tacaaaaccg acatcaaact ggacatcacc tcccacaacg aagactacac catcgttgaa
     1081 cagtacgaac gtgctgaagg tcgtcactcc accggtgctt aataacgctg atagtgctag
     1141 tgtagatcgc tactagagcc aggcatcaaa taaaacgaaa ggctcagtcg aaagactggg
     1201 cctttcgttt tatctgttgt ttgtcggtga acgctctcta ctagagtcac actggctcac
     1261 cttcgggtgg gcctttctgc gtttataggt ctcatgctta agctatgaaa cggaaatatt
     1321 gacagtagaa tatggattat taccgattgg taaaattgta gaaaagcgca tcgaatgtac
     1381 tgtttatagc gttgataata atggaaatat ttatacacaa cctgtagcac aatggcacga
     1441 tcgcggagaa caagaggtgt ttgagtattg tttggaagat ggttcattga ttcgggcaac
     1501 aaaagaccat aagtttatga ctgttgatgg tcaaatgttg ccaattgatg aaatatttga
     1561 acgtgaattg gatttgatgc gggttgataa tttgccgaat taataaatcg gtgaaatgca
     1621 cgactgatag tactagtagc ggccgctgca gtccggcaaa aaagggcaag gtgtcaccac
     1681 cctgcccttt ttctttaaaa ccgaaaagat tacttcgcgt tatgcaggct tcctcgctca
     1741 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     1801 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     1861 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccac aggctccgcc
     1921 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     1981 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     2041 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     2101 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     2161 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     2221 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     2281 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     2341 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     2401 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     2461 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     2521 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     2581 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     2641 atgagtaaac ttggtctgac agctcgagtc ccgtcaagtc agcgtaatgc tctgccagtg
     2701 ttacaaccaa ttaaccaatt ctgattagaa aaactcatcg agcatcaaat gaaactgcaa
     2761 tttattcata tcaggattat caataccata tttttgaaaa agccgtttct gtaatgaagg
     2821 agaaaactca ccgaggcagt tccataggat ggcaagatcc tggtatcggt ctgcgattcc
     2881 gactcgtcca acatcaatac aacctattaa tttcccctcg tcaaaaataa ggttatcaag
     2941 tgagaaatca ccatgagtga cgactgaatc cggtgagaat ggcaaaagct tatgcatttc
     3001 tttccagact tgttcaacag gccagccatt acgctcgtca tcaaaatcac tcgcatcaac
     3061 caaaccgtta ttcattcgtg attgcgcctg agcgagacga aatacgcgat cgctgttaaa
     3121 aggacaatta caaacaggaa tcgaatgcaa ccggcgcagg aacactgcca gcgcatcaac
     3181 aatattttca cctgaatcag gatattcttc taatacctgg aatgctgttt tcccggggat
     3241 cgcagtggtg agtaaccatg catcatcagg agtacggata aaatgcttga tggtcggaag
     3301 aggcataaat tccgtcagcc agtttagtct gaccatctca tctgtaacat cattggcaac
     3361 gctacctttg ccatgtttca gaaacaactc tggcgcatcg ggcttcccat acaatcgata
     3421 gattgtcgca cctgattgcc cgacattatc gcgagcccat ttatacccat ataaatcagc
     3481 atccatgttg gaatttaatc gcggcctgga gcaagacgtt tcccgttgaa tatggctcat
     3541 aacacccctt gtattactgt ttatgtaagc agacagtttt attgttcatg atgatatatt
     3601 tttatcttgt gcaatgtaac atcagagatt ttgagacaca acgtggcttt gttgaataaa
     3661 tcgaactttt gctgagttga aggatcagct cgagtgccac ctgacgtcta agaaaccatt
     3721 attatcatga cattaaccta taaaaatagg cgtatcacga ggcagaattt cagataaaaa
     3781 aaatccttag ctttcgctaa ggatgatttc tggaatt
//
