LOCUS       sequence-299953-        4837 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Chimeric protein of inactive T4L lysozyme and an estarase from 
            Bacillus subtilis.TEV cleaving site after the N-terminal Histag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4837)
  AUTHORS   Planchestainer M, Padrosa DR, Contente ML, Paradisi F
  TITLE     Genetically Fused T4L Acts as a Shield in Covalent Enzyme 
            Immobilisation Enhancing the Rescued Activity
  JOURNAL   Catalysts 2018, 8(1), 40
REFERENCE   2  (bases 1 to 4837)
  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: "Catalysts 
            2018, 8(1), 40"
COMMENT     Sequence Label: pRSETB BS2mT4L1
FEATURES             Location/Qualifiers
     source          1..4837
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     20..39
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        20..38
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             70..92
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             112..129
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             133..165
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             172..192
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     primer_bind     complement(2238..2256)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2252..2299
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2396..2851
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2483..2502)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2693..2714
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2877..2981
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2982..3050,3051..3842)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3200..3219)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4013..4601
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4502..4521
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4755..4772
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gatctcgatc ccgcgaaatt aatacgactc actataggga gaccacaacg gtttccctct
       61 agaaataatt ttgtttaact ttaagaagga gatatacata tgcggggttc tcatcatcat
      121 catcatcatg gtatggctag catgactggt ggacagcaaa tgggtcggga tgagaaccta
      181 tatttccagg gggatccgaa catcttcgag atgctgcgca tcgaccaggg cctgcgtctc
      241 aagatttaca aggacaccga aggttattac acgattggca tcggccacct cctgacaaag
      301 agcccatcac tcaacgctgc caagtctgaa ctggacaaag ccattggtcg caacaccaac
      361 ggtgtcatta caaaggacga ggcggagaaa ctcttcaacc aagatgtaga tgcggctgtc
      421 cgtggcatcc tgcgtaatgc caagttgaag cccgtgtatg actcccttga tgctgttcgc
      481 cgtgcagcct tgatcaacat ggttttccaa atgggtgaga ccggagtggc tggttttacg
      541 aactccctgc gcatgctcca gcagaagcgc tgggacgagg ccgcagtgaa tttggctaaa
      601 tctcgctggt acaatcagac acctaaccgt gccaagcgtg tcatcactac cttccgtact
      661 ggaacttggg acgcttacct cgagatctgc agctggtacc atgggatgac tcatcaaata
      721 gtaacgactc aatacggcaa agtaaaaggc acaacggaaa acggcgtaca taagtggaaa
      781 ggcatcccct atgccaagcc gcctgtcgga caatggcgtt ttaaagcacc tgagccgcct
      841 gaagtgtggg aagatgtgct tgatgccaca gcgtacggct ctatttgccc gcagccgtct
      901 gatttgctgt cactttcgta tactgagctg ccccgccagt ccgaggattg cttgtatgtc
      961 aatgtatttg cgcctgacac cccaagtaaa aatcttcctg tcatggtgtg gattcacgga
     1021 ggcgcttttt atctaggagc gggcagtgag ccattgtatg acggatcaaa acttgcggca
     1081 cagggagaag tcattgtcgt tacattgaac tatcggctgg ggccgtttgg ctttttgcac
     1141 ttgtcttcat ttaatgaggc gtattctgat aaccttgggc ttttagacca agccgccgcg
     1201 ctgaaatggg tgcgagagaa tatttcagcg tttggcggtg atcccgataa cgtaacagta
     1261 tttggagaat ccgccggcgg gatgagcatt gccgcgctgc ttgctatgcc tgcggcaaaa
     1321 ggcctgttcc agaaagcaat catggaaagc ggcgcttctc gaacgatgac gaaagaacaa
     1381 gcggcgagca cctcggcagc ctttttacag gtccttggga ttaacgaggg ccaactggat
     1441 aaattgcata cggtttctgc ggaagatttg ctaaaagcgg ctgatcagct tcggattgca
     1501 gaaaaagaaa atttctttca gctgttcttc cagcccgccc ttgatccgaa aacgctgcct
     1561 gaagaaccag aaaaagcgat cgcagaaggg gctgcttccg gtattccgct attaattgga
     1621 acaacccgtg atgaaggata tttatacttc accccggatt cagacgttca ttctcaggaa
     1681 acgcttgatg cagcgctcga gtatttacta gggaagccgc tggcagagaa agttgccgat
     1741 ttgtatccgc gttctctgga aagccaaatt cacatgatga ctgatttatt attttggcgc
     1801 cctgccgtcg cctatgcatc cgcacagtct cattacgccc ctgtctggat gtacaggttc
     1861 gattggcacc cgaagaagcc gccgtacaat aaagcgtttc acgcattaga gcttcctttt
     1921 gtctttggaa atctggacgg attggaacga atggcaaaag cggagattac ggatgaggtg
     1981 aaacagcttt ctcacacgat acaatcagcg tggatcacgt tcgccaaaac aggaaaccca
     2041 agcaccgaag ctgtgaattg gcctgcgtat catgaagaaa cgagagagac gctgatttta
     2101 gactcagaga ttacgatcga aaacgatccc gaatctgaaa aaaggcagaa gctattccct
     2161 tcaaaaggag aaggatcctg aaaagcttga tccggctgct aacaaagccc gaaaggaagc
     2221 tgagttggct gctgccaccg ctgagcaata actagcataa ccccttgggg cctctaaacg
     2281 ggtcttgagg ggttttttgc tgaaaggagg aactatatcc ggatctggcg taatagcgaa
     2341 gaggcccgca ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga atgggacgcg
     2401 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca
     2461 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc
     2521 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct
     2581 ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg
     2641 ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc
     2701 ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg
     2761 attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg
     2821 aattttaaca aaatattaac gcttacaatt taggtggcac ttttcgggga aatgtgcgcg
     2881 gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat
     2941 aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc
     3001 gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa
     3061 cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac
     3121 tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga
     3181 tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag
     3241 agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca
     3301 cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca
     3361 tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa
     3421 ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc
     3481 tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa
     3541 cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag
     3601 actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct
     3661 ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac
     3721 tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa
     3781 ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt
     3841 aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat
     3901 ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg
     3961 agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc
     4021 ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg
     4081 tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag
     4141 cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact
     4201 ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg
     4261 gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc
     4321 ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg
     4381 aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg
     4441 cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag
     4501 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc
     4561 gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct
     4621 ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc
     4681 ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc
     4741 gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgccca atacgcaaac
     4801 cgcctctccc cgcgcgttgg ccgattcatt aatgcag
//
