LOCUS       sequence-455994-        8050 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression in BL21(DE3) of T4 recombinase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8050)
  TITLE     Escherichia phage T4 Expression Constructs
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 8050)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pTYB1-T4uvsX
FEATURES             Location/Qualifiers
     source          1..8050
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(32..545)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(227..248)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     439..458
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     rep_origin      656..1244
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1145..1164
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1398..1415
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(1437..1456)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1597..1619
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(1613..1804)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(2361..3443)
                     /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     3405..3424
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3444..3521)
                     /gene="lacI"
                     /label=lacI promoter
     terminator      3674..3760
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3857..3943
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     4078..4097
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4078..4096
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    4097..4121
                     /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             4136..4158
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             5339..6157
                     /codon_start=1
                     /gene="Saccharomyces cerevisiae VMA1"
                     /product="5' region of the intein from the yeast Vma1 
                     subunit of the vacuolar ATPase (Chong et al., 1998)"
                     /label=Sce VMA intein 5' region
                     /translation="CFAKGTNVLMADGSIECIENIEVGNKVMGKDGRPREVIKLPRGRE
                     TMYSVVQKSQHRAHKSDSSREVPELLKFTCNATHELVVRTPRSVRRLSRTIKGVEYFEV
                     ITFEMGQKKAPDGRIVELVKEVSKSYPISEGPERANELVESYRKASNKAYFEWTIEARD
                     LSLLGSHVRKATYQTYAPILYENDHFFDYMQKSKFHLTIEGPKVLAYLLGLWIGDGLSD
                     RATFSVDSRDTSLMERVTEYAEKLNLCAEYKDRKEPQVAKTVNLYSKVVRG"
     CDS             6613..6624
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             6734..6889
                     /codon_start=1
                     /gene="Bacillus circulans chiA"
                     /product="chitin binding domain from chitinase A1 (Watanabe
                     et al., 1994)"
                     /label=CBD
                     /translation="TTNPGVSAWQVNTAYTAGQLVTYNGKTYKCLQPHTSLAGWEPSNV
                     PALWQLQ"
     primer_bind     complement(6954..6972)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      6968..7015
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        7075..7179
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7180..7248,7249..8040)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(7398..7417)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 aagtttactc atatatactt tagattgatt taccccggtt gataatcaga aaagccccaa
       61 aaacaggaag attgtataag caaatattta aattgtaaac gttaatattt tgttaaaatt
      121 cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat
      181 cccttataaa tcaaaagaat agcccgagat agggttgagt gttgttccag tttggaacaa
      241 gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg
      301 cgatggccca ctacgtgaac catcacccaa atcaagtttt ttggggtcga ggtgccgtaa
      361 agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc
      421 gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag
      481 tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg
      541 cgcgtaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac
      601 gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag
      661 atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg
      721 tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca
      781 gagcgcagat accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga
      841 actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca
      901 gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc
      961 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca
     1021 ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa
     1081 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc
     1141 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc
     1201 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg
     1261 cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat
     1321 cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca
     1381 gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc tatggtgcac tctcagtaca
     1441 atctgctctg atgccgcata gttaagccag tatacactcc gctatcgcta cgtgactggg
     1501 tcatggctgc gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc
     1561 tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt
     1621 tttcaccgtc atcaccgaaa cgcgcgaggc agctgcggta aagctcatca gcgtggtcgt
     1681 gcagcgattc acagatgtct gcctgttcat ccgcgtccag ctcgttgagt ttctccagaa
     1741 gcgttaatgt ctggcttctg ataaagcggg ccatgttaag ggcggttttt tcctgtttgg
     1801 tcactgatgc ctccgtgtaa gggggatttc tgttcatggg ggtaatgata ccgatgaaac
     1861 gagagaggat gctcacgata cgggttactg atgatgaaca tgcccggtta ctggaacgtt
     1921 gtgagggtaa acaactggcg gtatggatgc ggcgggacca gagaaaaatc actcagggtc
     1981 aatgccagcc gaacgccagc aagacgtagc ccagcgcgtc ggccgccatg ccggcgataa
     2041 tggcctgctt ctcgccgaaa cgtttggtgg cgggaccagt gacgaaggct tgagcgaggg
     2101 cgtgcaagat tccgaatacc gcaagcgaca ggccgatcat cgtcgcgctc cagcgaaagc
     2161 ggtcctcgcc gaaaatgacc cagagcgctg ccggcacctg tcctacgagt tgcatgataa
     2221 agaagacagt cataagtgcg gcgacgatag tcatgccccg cgcccaccgg aaggagctga
     2281 ctgggttgaa ggctctcaag ggcatcggtc gagatcccgg tgcctaatga gtgagctaac
     2341 ttacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     2401 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgccagggtg
     2461 gtttttcttt tcaccagtga gacgggcaac agctgattgc ccttcaccgc ctggccctga
     2521 gagagttgca gcaagcggtc cacgctggtt tgccccagca ggcgaaaatc ctgtttgatg
     2581 gtggttaacg gcgggatata acatgagctg tcttcggtat cgtcgtatcc cactaccgag
     2641 atatccgcac caacgcgcag cccggactcg gtaatggcgc gcattgcgcc cagcgccatc
     2701 tgatcgttgg caaccagcat cgcagtggga acgatgccct cattcagcat ttgcatggtt
     2761 tgttgaaaac cggacatggc actccagtcg ccttcccgtt ccgctatcgg ctgaatttga
     2821 ttgcgagtga gatatttatg ccagccagcc agacgcagac gcgccgagac agaacttaat
     2881 gggcccgcta acagcgcgat ttgctggtga cccaatgcga ccagatgctc cacgcccagt
     2941 cgcgtaccgt cttcatggga gaaaataata ctgttgatgg gtgtctggtc agagacatca
     3001 agaaataacg ccggaacatt agtgcaggca gcttccacag caatggcatc ctggtcatcc
     3061 agcggatagt taatgatcag cccactgacg cgttgcgcga gaagattgtg caccgccgct
     3121 ttacaggctt cgacgccgct tcgttctacc atcgacacca ccacgctggc acccagttga
     3181 tcggcgcgag atttaatcgc cgcgacaatt tgcgacggcg cgtgcagggc cagactggag
     3241 gtggcaacgc caatcagcaa cgactgtttg cccgccagtt gttgtgccac gcggttggga
     3301 atgtaattca gctccgccat cgccgcttcc actttttccc gcgttttcgc agaaacgtgg
     3361 ctggcctggt tcaccacgcg ggaaacggtc tgataagaga caccggcata ctctgcgaca
     3421 tcgtataacg ttactggttt cacattcacc accctgaatt gactctcttc cgggcgctat
     3481 catgccatac cgcgaaaggt tttgcgccat tcgatggtgt ccgggatctc gacgctctcc
     3541 cttatgcgac tcctgcatta ggaagcagcc cagtagtagg ttgaggccgt tgagcaccgc
     3601 cgccgcaagg aatggtgcat gccggcatgc cgccctttcg tcttcaagaa ttaattccca
     3661 attccccagg catcaaataa aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat
     3721 ctgttgtttg tcggtgaacg ctctcctgag taggacaaat ccgccgggag cggatttgaa
     3781 cgttgcgaag caacggcccg gagggtggcg ggcaggacgc ccgccataaa ctgccaggaa
     3841 ttaattcccc aggcatcaaa taaaacgaaa ggctcagtcg aaagactggg cctttcgttt
     3901 tatctgttgt ttgtcggtga acgctctcct gagtaggaca aatccgccgg gagcggattt
     3961 gaacgttgcg aagcaacggc ccggagggtg gcgggcagga cgcccgccat aaactgccag
     4021 gaattgggga tcggaattaa ttcccggttt aaaccgggga tctcgatccc gcgaaattaa
     4081 tacgactcac tataggggaa ttgtgagcgg ataacaattc ccctctagaa ataattttgt
     4141 ttaactttaa gaaggagata tacatatgtc tgatttaaaa tctcgtttga ttaaagcttc
     4201 tacttctaaa ttgactgcag aattaacagc atctaaattc tttaatgaaa aagatgtagt
     4261 tcgaacaaaa attcctatga tgaatattgc actttctggt gaaattactg gtggtatgca
     4321 atctggtctg ttaattttgg caggtccatc aaaatcattt aaatcaaact ttggattaac
     4381 aatggtgtca tcttacatgc gtcaatatcc tgatgcagta tgtttgtttt atgatagtga
     4441 atttggtatt actcctgctt atttgcggtc tatgggagtc gatccagaac gagtaattca
     4501 tactccggtt caatcacttg agcaattacg tattgacatg gttaatcaat tggatgcaat
     4561 tgaacgcggc gaaaaggtag tcgtttttat cgattcactt ggtaacttag cttctaagaa
     4621 agaaactgaa gatgctttaa atgaaaaagt tgttagtgat atgactagag ctaaaacaat
     4681 gaaaagctta tttcgtatcg taactcccta ttttagtact aaaaatattc catgtattgc
     4741 tattaaccat acatacgaaa cacaagaaat gtttagtaaa acagttatgg gaggtggtac
     4801 tggaccgatg tattcggctg ataccgtatt cattatcggt aaacgccaga ttaaagatgg
     4861 ttctgatctt caggggtatc aatttgttct aaatgtagaa aaatctcgta ccgttaaaga
     4921 aaaaagtaaa ttctttattg atgttaaatt tgacggtggt atcgatcctt attctggatt
     4981 gttagatatg gctctagaat taggattcgt ggtaaaacct aaaaatggct ggtatgctcg
     5041 tgaatttctt gacgaagaaa ctggcgagat gattcgcgaa gaaaaatctt ggcgtgcaaa
     5101 agataccaac tgcactacat tctggggtcc tttatttaag catcaaccat tccgagatgc
     5161 tattaaacgt gcttatcagt taggtgctat tgatagtaat gaaattgttg aagctgaagt
     5221 tgatgaattg attaactcaa aggttgaaaa atttaaatct ccagaaagta aaagtaaatc
     5281 agctgctgat ttagaaactg acctcgaaca gctaagtgat atggaagaat ttaatgcgtg
     5341 ctttgccaag ggtaccaatg ttttaatggc ggatgggtct attgaatgta ttgaaaacat
     5401 tgaggttggt aataaggtca tgggtaaaga tggcagacct cgtgaggtaa ttaaattgcc
     5461 cagaggaaga gaaactatgt acagcgtcgt gcagaaaagt cagcacagag cccacaaaag
     5521 tgactcaagt cgtgaagtgc cagaattact caagtttacg tgtaatgcga cccatgagtt
     5581 ggttgttaga acacctcgta gtgtccgccg tttgtctcgt accattaagg gtgtcgaata
     5641 ttttgaagtt attacttttg agatgggcca aaagaaagcc cccgacggta gaattgttga
     5701 gcttgtcaag gaagtttcaa agagctaccc aatatctgag gggcctgaga gagccaacga
     5761 attagtagaa tcctatagaa aggcttcaaa taaagcttat tttgagtgga ctattgaggc
     5821 cagagatctt tctctgttgg gttcccatgt tcgtaaagct acctaccaga cttacgctcc
     5881 aattctttat gagaatgacc actttttcga ctacatgcaa aaaagtaagt ttcatctcac
     5941 cattgaaggt ccaaaagtac ttgcttattt acttggttta tggattggtg atggattgtc
     6001 tgacagggca actttttcgg ttgattccag agatacttct ttgatggaac gtgttactga
     6061 atatgctgaa aagttgaatt tgtgcgccga gtataaggac agaaaagaac cacaagttgc
     6121 caaaactgtt aatttgtact ctaaagttgt cagaggtaat ggtattcgca ataatcttaa
     6181 tactgagaat ccattatggg acgctattgt tggcttagga ttcttgaagg acggtgtcaa
     6241 aaatattcct tctttcttgt ctacggacaa tatcggtact cgtgaaacat ttcttgctgg
     6301 tctaattgat tctgatggct atgttactga tgagcatggt attaaagcaa caataaagac
     6361 aattcatact tctgtcagag atggtttggt ttcccttgct cgttctttag gcttagtagt
     6421 ctcggttaac gcagaacctg ctaaggttga catgaatgtc accaaacata aaattagtta
     6481 tgctatttat atgtctggtg gagatgtttt gcttaacgtt ctttcgaagt gtgccggctc
     6541 taaaaaattc aggcctgctc ccgccgctgc ttttgcacgt gagtgccgcg gattttattt
     6601 cgagttacaa gaattgaagg aagacgatta ttatgggatt actttatctg atgattctga
     6661 tcatcagttt ttgcttggat cccaggttgt cgtccatgca tgcggtggcc tgaccggtct
     6721 gaactcaggc ctcacgacaa atcctggtgt atccgcttgg caggtcaaca cagcttatac
     6781 tgcgggacaa ttggtcacat ataacggcaa gacgtataaa tgtttgcagc cccacacctc
     6841 cttggcagga tgggaaccat ccaacgttcc tgccttgtgg cagcttcaat gactgcagga
     6901 aggggatccg gctgctaaca aagcccgaaa ggaagctgag ttggctgctg ccaccgctga
     6961 gcaataacta gcataacccc ttggggcctc taaacgggtc ttgaggggtt ttttgctgaa
     7021 aggaggaact atatccggat aactacgtca ggtggcactt ttcggggaaa tgtgcgcgga
     7081 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     7141 ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt
     7201 gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg
     7261 ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg
     7321 gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt cccaatgatg
     7381 agcactttta aagttctgct atgtggcgcg gtattatccc gtgttgacgc cgggcaagag
     7441 caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca
     7501 gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg
     7561 agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc
     7621 gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg
     7681 aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg
     7741 ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca attaatagac
     7801 tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg
     7861 tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg
     7921 gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact
     7981 atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa
     8041 ctgtcagacc
//
