LOCUS       sequence-455998-        6840 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression in BL21(DE3) of T4 3' to 5' helicase/single-strand DNA 
            annealase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6840)
  AUTHORS   Nelson SW, Benkovic SJ
  TITLE     The T4 phage UvsW protein contains both DNA unwinding and strand 
            annealing activities.
  JOURNAL   J Biol Chem. 2007 Jan 5;282(1):407-16. doi: 10.1074/jbc.M608153200. 
            Epub 2006 Nov 8.
  PUBMED    17092935
REFERENCE   2  (bases 1 to 6840)
  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: 
            "10.1074/jbc.M608153200"; journalName: "J Biol Chem"; date: 
            "2007-01-5- 5"; volume: "282"; issue: "1"; pages: "407-16"
COMMENT     Sequence Label: pET28a-T4uvsW
FEATURES             Location/Qualifiers
     source          1..6840
                     /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(1718..1735)
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             complement(1745..1762)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             1781..1803
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    1818..1842
                     /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(1842..1861)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1843..1861)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1945..1964)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        2170..2247
                     /gene="lacI"
                     /label=lacI promoter
     CDS             2248..3330
                     /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(2267..2286)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             4139..4330
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(4324..4346)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4432..4571
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4487..4506
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4586..4603)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4757..5345)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4837..4856)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(5456..5475)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             5467..6282
                     /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             5467..6282
                     /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     complement(5540..5559)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(6374..6829)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6511..6532)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6723..6742
                     /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 gagtgcggcc gcaagcttgt
      181 cgacggagct cgaattcgga tccggttata aattaactgt tttcattacg taattaaatt
      241 tttcatctgc gtagcgctga atacgatcaa tgccgtgttt taaaagatag ttcaaatgaa
      301 catatttctt tttcgtatta gcagattttg gcttgacgcc tgcgtcatct atgaggtccc
      361 agactgttgc tattgtctta gaaccatgct tacgtaatac acgaccgatt gtttgtaata
      421 caatgatttt tgatttaaca ccgtgcgcta aaacaacgtg atgcagattt ttaactgaaa
      481 taccagtaga aaatacacca taactagcta ctataattat tcctttacca ttttcagcta
      541 aggttttcat tatattgcgg gtttcggtat caacttcccc tgatacgtaa taaactttat
      601 cgtattcatt tttaattaaa tcgaaaatag ctttaccatg cgatacatgt ttaaacatga
      661 caaaagcgtt ttcatctttt tgcgcaagct taatagctaa tttagcgatc catttatttc
      721 ttttactaag cccagtaata atttttattt cttcttggta agtttttccc tttaatttag
      781 tagtgaactc atcgggatag cgaagaaaaa tactattaat ttttagctca gttacttgtc
      841 catcttccat taatttagaa gtcgttactg gtttaaatat ttcaccaaac attccaacat
      901 actgcatgat attggctttg ccatcacgta atgaaccaga caaaccgaat ttgaacatgc
      961 agttatttaa acctgatatg atagatgaaa tactttttcc tgtagcaaga tggcattcat
     1021 cattcatcat cattccaaac tgtgagaacc attctttcgg ttgttttact acagtttgcc
     1081 atgtaccaac aacgactggt gcatcatttt tatatttatc atcttttgat gctccgccac
     1141 caattttctt tatcattgca tggctgaata aacgatagtc gacgaagtca tcagccatct
     1201 gagttgtcag agcagttgtt ggaacaatga taagaatttt accttcataa ttttccaaat
     1261 aatatcgcgc aagcaaagct tgaattaaag atttacctgc agatgttgga agattaagaa
     1321 ttctacgacg attaactaat ccttcgaaca ctgcatcttt ttgataccag tgcggttcaa
     1381 ttcttttatt tcctgaatag atttctaatt tagaaagcca ttcatcaaaa tcttttcttg
     1441 ataattcttc tttttcgtta atttgtgggt caatccaggc tttatagcca aaattatcac
     1501 agaatttttt aatttgcccg actaagccga atggaagaag acgattataa tctaaaagac
     1561 ggattcgtcc atcccagttg ccatatctga agcgaggatt aaatctatat ccatcggcct
     1621 caaacgaaaa gaaatctctt aattcgtgga acgtgctctc ttcacaatcg atgcgtacat
     1681 gactgaagtc gtgaaaatgt actttaatat ccatatggct gccgcgcggc accaggccgc
     1741 tgctgtgatg atgatgatga tggctgctgc ccatggtata tctccttctt aaagttaaac
     1801 aaaattattt ctagagggga attgttatcc gctcacaatt cccctatagt gagtcgtatt
     1861 aatttcgcgg gatcgagatc tcgatcctct acgccggacg catcgtggcc ggcatcaccg
     1921 gcgccacagg tgcggttgct ggcgcctata tcgccgacat caccgatggg gaagatcggg
     1981 ctcgccactt cgggctcatg agcgcttgtt tcggcgtggg tatggtggca ggccccgtgg
     2041 ccgggggact gttgggcgcc atctccttgc atgcaccatt ccttgcggcg gcggtgctca
     2101 acggcctcaa cctactactg ggctgcttcc taatgcagga gtcgcataag ggagagcgtc
     2161 gagatcccgg acaccatcga atggcgcaaa acctttcgcg gtatggcatg atagcgcccg
     2221 gaagagagtc aattcagggt ggtgaatgtg aaaccagtaa cgttatacga tgtcgcagag
     2281 tatgccggtg tctcttatca gaccgtttcc cgcgtggtga accaggccag ccacgtttct
     2341 gcgaaaacgc gggaaaaagt ggaagcggcg atggcggagc tgaattacat tcccaaccgc
     2401 gtggcacaac aactggcggg caaacagtcg ttgctgattg gcgttgccac ctccagtctg
     2461 gccctgcacg cgccgtcgca aattgtcgcg gcgattaaat ctcgcgccga tcaactgggt
     2521 gccagcgtgg tggtgtcgat ggtagaacga agcggcgtcg aagcctgtaa agcggcggtg
     2581 cacaatcttc tcgcgcaacg cgtcagtggg ctgatcatta actatccgct ggatgaccag
     2641 gatgccattg ctgtggaagc tgcctgcact aatgttccgg cgttatttct tgatgtctct
     2701 gaccagacac ccatcaacag tattattttc tcccatgaag acggtacgcg actgggcgtg
     2761 gagcatctgg tcgcattggg tcaccagcaa atcgcgctgt tagcgggccc attaagttct
     2821 gtctcggcgc gtctgcgtct ggctggctgg cataaatatc tcactcgcaa tcaaattcag
     2881 ccgatagcgg aacgggaagg cgactggagt gccatgtccg gttttcaaca aaccatgcaa
     2941 atgctgaatg agggcatcgt tcccactgcg atgctggttg ccaacgatca gatggcgctg
     3001 ggcgcaatgc gcgccattac cgagtccggg ctgcgcgttg gtgcggatat ctcggtagtg
     3061 ggatacgacg ataccgaaga cagctcatgt tatatcccgc cgttaaccac catcaaacag
     3121 gattttcgcc tgctggggca aaccagcgtg gaccgcttgc tgcaactctc tcagggccag
     3181 gcggtgaagg gcaatcagct gttgcccgtc tcactggtga aaagaaaaac caccctggcg
     3241 cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gctggcacga
     3301 caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtaa gttagctcac
     3361 tcattaggca ccgggatctc gaccgatgcc cttgagagcc ttcaacccag tcagctcctt
     3421 ccggtgggcg cggggcatga ctatcgtcgc cgcacttatg actgtcttct ttatcatgca
     3481 actcgtagga caggtgccgg cagcgctctg ggtcattttc ggcgaggacc gctttcgctg
     3541 gagcgcgacg atgatcggcc tgtcgcttgc ggtattcgga atcttgcacg ccctcgctca
     3601 agccttcgtc actggtcccg ccaccaaacg tttcggcgag aagcaggcca ttatcgccgg
     3661 catggcggcc ccacgggtgc gcatgatcgt gctcctgtcg ttgaggaccc ggctaggctg
     3721 gcggggttgc cttactggtt agcagaatga atcaccgata cgcgagcgaa cgtgaagcga
     3781 ctgctgctgc aaaacgtctg cgacctgagc aacaacatga atggtcttcg gtttccgtgt
     3841 ttcgtaaagt ctggaaacgc ggaagtcagc gccctgcacc attatgttcc ggatctgcat
     3901 cgcaggatgc tgctggctac cctgtggaac acctacatct gtattaacga agcgctggca
     3961 ttgaccctga gtgatttttc tctggtcccg ccgcatccat accgccagtt gtttaccctc
     4021 acaacgttcc agtaaccggg catgttcatc atcagtaacc cgtatcgtga gcatcctctc
     4081 tcgtttcatc ggtatcatta cccccatgaa cagaaatccc ccttacacgg aggcatcagt
     4141 gaccaaacag gaaaaaaccg cccttaacat ggcccgcttt atcagaagcc agacattaac
     4201 gcttctggag aaactcaacg agctggacgc ggatgaacag gcagacatct gtgaatcgct
     4261 tcacgaccac gctgatgagc tttaccgcag ctgcctcgcg cgtttcggtg atgacggtga
     4321 aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag cggatgccgg
     4381 gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg gcgcagccat
     4441 gacccagtca cgtagcgata gcggagtgta tactggctta actatgcggc atcagagcag
     4501 attgtactga gagtgcacca ttgcggtgtg aaataccgca cagatgcgta aggagaaaat
     4561 accgcatcag gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     4621 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     4681 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     4741 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     4801 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     4861 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     4921 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     4981 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     5041 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     5101 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     5161 tcttgaagtg gtggcctaac tacggctaca ctagaaggac agtatttggt atctgcgctc
     5221 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     5281 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     5341 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     5401 gttaagggat tttggtcatg aacaataaaa ctgtctgctt acataaacag taatacaagg
     5461 ggtgttatga gccatattca acgggaaacg tcttgctcta ggccgcgatt aaattccaac
     5521 atggatgctg atttatatgg gtataaatgg gctcgcgata atgtcgggca atcaggtgcg
     5581 acaatctatc gattgtatgg gaagcccgat gcgccagagt tgtttctgaa acatggcaaa
     5641 ggtagcgttg ccaatgatgt tacagatgag atggtcagac taaactggct gacggaattt
     5701 atgcctcttc cgaccatcaa gcattttatc cgtactcctg atgatgcatg gttactcacc
     5761 actgcgatcc ccgggaaaac agcattccag gtattagaag aatatcctga ttcaggtgaa
     5821 aatattgttg atgcgctggc agtgttcctg cgccggttgc attcgattcc tgtttgtaat
     5881 tgtcctttta acagcgatcg cgtatttcgt ctcgctcagg cgcaatcacg aatgaataac
     5941 ggtttggttg atgcgagtga ttttgatgac gagcgtaatg gctggcctgt tgaacaagtc
     6001 tggaaagaaa tgcataaact tttgccattc tcaccggatt cagtcgtcac tcatggtgat
     6061 ttctcacttg ataaccttat ttttgacgag gggaaattaa taggttgtat tgatgttgga
     6121 cgagtcggaa tcgcagaccg ataccaggat cttgccatcc tatggaactg cctcggtgag
     6181 ttttctcctt cattacagaa acggcttttt caaaaatatg gtattgataa tcctgatatg
     6241 aataaattgc agtttcattt gatgctcgat gagtttttct aagaattaat tcatgagcgg
     6301 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     6361 aaaagtgcca cctaaattgt aagcgttaat attttgttaa aattcgcgtt aaatttttgt
     6421 taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta taaatcaaaa
     6481 gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc actattaaag
     6541 aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg cccactacgt
     6601 gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac
     6661 cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt ggcgagaaag
     6721 gaagggaaga aagcgaaagg agcgggcgct agggcgctgg caagtgtagc ggtcacgctg
     6781 cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc ccattcgcca
//
