LOCUS       sequence-422888-        5908 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Enzymatic biotinylation of proteins at a specific lysine residue in 
            a recognition sequence. This has multiple applications, especially 
            in the study of protein-protein interactions..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5908)
  AUTHORS   O'callaghan CA, Byford MF, Wyer JR, Willcox BE, Jakobsen BK, 
            McMichael AJ, Bell JI
  TITLE     BirA enzyme: production and application in the study of membrane 
            receptor-ligand interactions by site-specific biotinylation.
  JOURNAL   Anal Biochem. 1999 Jan 1;266(1):9-15. doi: 10.1006/abio.1998.2930.
  PUBMED    9887208
REFERENCE   2  (bases 1 to 5908)
  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: "10.1006/abio.1998";
            journalName: "Anal Biochem"; date: "1999-01-1- 1"; volume: "266"; 
            issue: "1"; pages: "9-15"
COMMENT     Sequence Label: COC053
FEATURES             Location/Qualifiers
     source          1..5908
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(194..199,200..215,216..222)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    230..246
                     /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)."
     CDS             269..922
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     880..902
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             929..946
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(1969..1991)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2136..2154)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2222..2326
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2327..2395,2396..3187)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2545..2564)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3358..3946
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3847..3866
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4100..4117
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4190..4267
                     /gene="lacI"
                     /label=lacI promoter
     CDS             4268..5350
                     /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(4287..4306)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(5399..5404,5405..5422,5423..5429)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5437..5453
                     /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     5461..5477
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5461..5477
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(5473..5908,1..92)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5493..5510)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5493..5509)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5502..5524)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 gcgttggaat tagcttatcg actgcacggt gcaccaatgc ttctggcgtc aggcagccat
       61 cggaagctgt ggtatggctg tgcaggtcgt aaatcactgc ataattcgtg tcgctcaagg
      121 cgcactcccg ttctggataa tgttttttgc gccgacatca taacggttct ggcaaatatt
      181 ctgaaatgag ctgttgacaa ttaatcatcg gctcgtataa tgtgtggaat tgtgagcgga
      241 taacaatttc acacaggaaa cagtattcat gtcccctata ctaggttatt ggaaaattaa
      301 gggccttgtg caacccactc gacttctttt ggaatatctt gaagaaaaat atgaagagca
      361 tttgtatgag cgcgatgaag gtgataaatg gcgaaacaaa aagtttgaat tgggtttgga
      421 gtttcccaat cttccttatt atattgatgg tgatgttaaa ttaacacagt ctatggccat
      481 catacgttat atagctgaca agcacaacat gttgggtggt tgtccaaaag agcgtgcaga
      541 gatttcaatg cttgaaggag cggttttgga tattagatac ggtgtttcga gaattgcata
      601 tagtaaagac tttgaaactc tcaaagttga ttttcttagc aagctacctg aaatgctgaa
      661 aatgttcgaa gatcgtttat gtcataaaac atatttaaat ggtgatcatg taacccatcc
      721 tgacttcatg ttgtatgacg ctcttgatgt tgttttatac atggacccaa tgtgcctgga
      781 tgcgttccca aaattagttt gttttaaaaa acgtattgaa gctatcccac aaattgataa
      841 gtacttgaaa tccagcaagt atatagcatg gcctttgcag ggctggcaag ccacgtttgg
      901 tggtggcgac catcctccaa aatcggatct ggttccgcgt ggatccaagg ataacaccgt
      961 gccactgaaa ttgattgccc tgttagcgaa cggtgaattt cactctggcg agcagttggg
     1021 tgaaacgctg ggaatgagcc gggcggctat taataaacac attcagacac tgcgtgactg
     1081 gggcgttgat gtctttaccg ttccgggtaa aggatacagc ctgcctgagc ctatccagtt
     1141 acttaatgct aaacagatat tgggtcagct ggatggcggt agtgtagccg tgctgccagt
     1201 gattgactcc acgaatcagt accttcttga tcgtatcgga gagcttaaat cgggcgatgc
     1261 ttgcattgca gaataccagc aggctggccg tggtcgccgg ggtcggaaat ggttttcgcc
     1321 ttttggcgca aacttatatt tgtcgatgtt ctggcgtctg gaacaaggcc cggcggcggc
     1381 gattggttta agtctggtta tcggtatcgt gatggcggaa gtattacgca agctgggtgc
     1441 agataaagtt cgtgttaaat ggcctaatga cctctatctg caggatcgca agctggcagg
     1501 cattctggtg gagctgactg gcaaaactgg cgatgcggcg caaatagtca ttggagccgg
     1561 gatcaacatg gcaatgcgcc gtgttgaaga gagtgtcgtt aatcaggggt ggatcacgct
     1621 gcaggaagcg gggatcaatc tcgatcgtaa tacgttggcg gccatgctaa tacgtgaatt
     1681 acgtgctgcg ttggaactct tcgaacaaga aggattggca ccttatctgt cgcgctggga
     1741 aaagctggat aattttatta atcgcccagt gaaacttatc attggtgata aagaaatatt
     1801 tggcatttca cgcggaatag acaaacaggg ggctttatta cttgagcagg atggaataat
     1861 aaaaccctgg atgggcggtg aaatatccct gcgtagtgca ggatccccgg gaattcatcg
     1921 tgactgactg acgatctgcc tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat
     1981 gcagctcccg gagacggtca cagcttgtct gtaagcggat gccgggagca gacaagcccg
     2041 tcagggcgcg tcagcgggtg ttggcgggtg tcggggcgca gccatgaccc agtcacgtag
     2101 cgatagcgga gtgtataatt cttgaagacg aaagggcctc gtgatacgcc tatttttata
     2161 ggttaatgtc atgataataa tggtttctta gacgtcaggt ggcacttttc ggggaaatgt
     2221 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag
     2281 acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca
     2341 tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc
     2401 agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat
     2461 cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc
     2521 aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgtg ttgacgccgg
     2581 gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc
     2641 agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat
     2701 aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga
     2761 gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc
     2821 ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg cagcaatggc
     2881 aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt
     2941 aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc
     3001 tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc
     3061 agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca
     3121 ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca
     3181 ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa aacttcattt
     3241 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     3301 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     3361 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     3421 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     3481 cagagcgcag ataccaaata ctgtccttct agtgtagccg tagttaggcc accacttcaa
     3541 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     3601 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     3661 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     3721 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     3781 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     3841 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     3901 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     3961 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt
     4021 atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg
     4081 cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcctgatgcg
     4141 gtattttctc cttacgcatc tgtgcggtat ttcacaccgc ataaattccg acaccatcga
     4201 atggcgcaaa acctttcgcg gtatggcatg atagcgcccg gaagagagtc aattcagggt
     4261 ggtgaatgtg aaaccagtaa cgttatacga tgtcgcagag tatgccggtg tctcttatca
     4321 gaccgtttcc cgcgtggtga accaggccag ccacgtttct gcgaaaacgc gggaaaaagt
     4381 ggaagcggcg atggcggagc tgaattacat tcccaaccgc gtggcacaac aactggcggg
     4441 caaacagtcg ttgctgattg gcgttgccac ctccagtctg gccctgcacg cgccgtcgca
     4501 aattgtcgcg gcgattaaat ctcgcgccga tcaactgggt gccagcgtgg tggtgtcgat
     4561 ggtagaacga agcggcgtcg aagcctgtaa agcggcggtg cacaatcttc tcgcgcaacg
     4621 cgtcagtggg ctgatcatta actatccgct ggatgaccag gatgccattg ctgtggaagc
     4681 tgcctgcact aatgttccgg cgttatttct tgatgtctct gaccagacac ccatcaacag
     4741 tattattttc tcccatgaag acggtacgcg actgggcgtg gagcatctgg tcgcattggg
     4801 tcaccagcaa atcgcgctgt tagcgggccc attaagttct gtctcggcgc gtctgcgtct
     4861 ggctggctgg cataaatatc tcactcgcaa tcaaattcag ccgatagcgg aacgggaagg
     4921 cgactggagt gccatgtccg gttttcaaca aaccatgcaa atgctgaatg agggcatcgt
     4981 tcccactgcg atgctggttg ccaacgatca gatggcgctg ggcgcaatgc gcgccattac
     5041 cgagtccggg ctgcgcgttg gtgcggatat ctcggtagtg ggatacgacg ataccgaaga
     5101 cagctcatgt tatatcccgc cgttaaccac catcaaacag gattttcgcc tgctggggca
     5161 aaccagcgtg gaccgcttgc tgcaactctc tcagggccag gcggtgaagg gcaatcagct
     5221 gttgcccgtc tcactggtga aaagaaaaac caccctggcg cccaatacgc aaaccgcctc
     5281 tccccgcgcg ttggccgatt cattaatgca gctggcacga caggtttccc gactggaaag
     5341 cgggcagtga gcgcaacgca attaatgtaa gttagctcac tcattaggca ccccaggctt
     5401 tacactttat gcttccggct cgtatgttgt gtggaattgt gagcggataa caatttcaca
     5461 caggaaacag ctatgaccat gattacggat tcactggccg tcgttttaca acgtcgtgac
     5521 tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc tttcgccagc
     5581 tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat
     5641 ggcgaatggc gctttgcctg gtttccggca ccagaagcgg tgccggaaag ctggctggag
     5701 tgcgatcttc ctgaggccga tactgtcgtc gtcccctcaa actggcagat gcacggttac
     5761 gatgcgccca tctacaccaa cgtgacctat cccattacgg tcaatccgcc gtttgttccc
     5821 acggagaatc cgacgggttg ttactcgctc acatttaatg ttgatgaaag ctggctacag
     5881 gaaggccaga cgcgaattat ttttgatg
//
