LOCUS       sequence-208996-        5606 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For expression in E coli, and affinity purification of N-terminally 
            GST-tagged Archaeoglobus fulgidus RNase HII.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5606)
  AUTHORS   Reijns MA, Bubeck D, Gibson LC, Graham SC, Baillie GS, Jones EY, 
            Jackson AP
  TITLE     The structure of the human RNase H2 complex defines key interaction 
            interfaces relevant to enzyme function and human disease.
  JOURNAL   J Biol Chem. 2011 Mar 25;286(12):10530-9. doi: 
            10.1074/jbc.M110.177394. Epub 2010 Dec 22.
  PUBMED    21177854
REFERENCE   2  (bases 1 to 5606)
  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.M110.177394"; journalName: "J Biol Chem"; date: 
            "2011-03-25- 25"; volume: "286"; issue: "12"; pages: "10530-9"
COMMENT     Sequence Label: pGEX6P1‐Afu rnhB
FEATURES             Location/Qualifiers
     source          1..5606
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(106..111,112..127,128..134)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    142..158
                     /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             181..834
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     792..814
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             841..864
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(1579..1601)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1746..1764)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1832..1936
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1937..2005,2006..2797)
                     /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(2155..2174)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2968..3556
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3457..3476
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3710..3727
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3800..3877
                     /gene="lacI"
                     /label=lacI promoter
     CDS             3878..4960
                     /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(3897..3916)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(5009..5014,5015..5032,5033..5039)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5047..5063
                     /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     5071..5087
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5071..5087
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(5083..5606,1..4)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5103..5120)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5103..5119)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5112..5134)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 gtaaatcact gcataattcg tgtcgctcaa ggcgcactcc cgttctggat aatgtttttt
       61 gcgccgacat cataacggtt ctggcaaata ttctgaaatg agctgttgac aattaatcat
      121 cggctcgtat aatgtgtgga attgtgagcg gataacaatt tcacacagga aacagtattc
      181 atgtccccta tactaggtta ttggaaaatt aagggccttg tgcaacccac tcgacttctt
      241 ttggaatatc ttgaagaaaa atatgaagag catttgtatg agcgcgatga aggtgataaa
      301 tggcgaaaca aaaagtttga attgggtttg gagtttccca atcttcctta ttatattgat
      361 ggtgatgtta aattaacaca gtctatggcc atcatacgtt atatagctga caagcacaac
      421 atgttgggtg gttgtccaaa agagcgtgca gagatttcaa tgcttgaagg agcggttttg
      481 gatattagat acggtgtttc gagaattgca tatagtaaag actttgaaac tctcaaagtt
      541 gattttctta gcaagctacc tgaaatgctg aaaatgttcg aagatcgttt atgtcataaa
      601 acatatttaa atggtgatca tgtaacccat cctgacttca tgttgtatga cgctcttgat
      661 gttgttttat acatggaccc aatgtgcctg gatgcgttcc caaaattagt ttgttttaaa
      721 aaacgtattg aagctatccc acaaattgat aagtacttga aatccagcaa gtatatagca
      781 tggcctttgc agggctggca agccacgttt ggtggtggcg accatcctcc aaaatcggat
      841 ctggaagttc tgttccaggg gcccctggga tccccggaat tcccgggtcg actcatgaag
      901 gcaggcatcg atgaggctgg aaagggctgc gtcatcggcc cactggttgt tgcaggagtg
      961 gcttgcagcg atgaggatag gctgagaaag cttggtgtga aagactccaa aaagctaagt
     1021 caggggagga gagaggaact agccgaggaa ataaggaaaa tctgcagaac ggaggttttg
     1081 aaagtttctc ccgaaaatct cgacgaaagg atggctgcta aaaccataaa cgagattttg
     1141 aaggagtgct acgctgaaat aattctcagg ctgaagccgg aaattgctta tgttgacagt
     1201 cctgatgtga ttcccgagag actttcgagg gagcttgagg agattacggg gttgagagtt
     1261 gtggccgagc acaaggcgga cgagaagtat cccctggtag ctgcggcttc aatcatcgca
     1321 aaggtggaaa gggagcggga gattgagagg ctgaaagaaa aattcgggga tttcggcagc
     1381 ggctatgcga gcgatccgag gacaagagaa gtgctgaagg agtggatagc ttcaggcaga
     1441 attccgagct gcgtgagaat gcgctggaag acggtgtcaa atctgaggca gaagacgctt
     1501 gacgatttct aagagctcgc ggccgcatcg tgactgactg acgatctgcc tcgcgcgttt
     1561 cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca cagcttgtct
     1621 gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg
     1681 tcggggcgca gccatgaccc agtcacgtag cgatagcgga gtgtataatt cttgaagacg
     1741 aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta
     1801 gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
     1861 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata
     1921 ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
     1981 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
     2041 agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
     2101 tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg
     2161 tggcgcggta ttatcccgtg ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
     2221 ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat
     2281 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
     2341 acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga
     2401 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
     2461 gcgtgacacc acgatgcctg cagcaatggc aacaacgttg cgcaaactat taactggcga
     2521 actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc
     2581 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
     2641 cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg
     2701 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
     2761 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata
     2821 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct
     2881 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     2941 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     3001 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     3061 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgtccttct
     3121 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
     3181 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     3241 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     3301 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     3361 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     3421 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     3481 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     3541 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
     3601 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac
     3661 cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt
     3721 gagcgaggaa gcggaagagc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat
     3781 ttcacaccgc ataaattccg acaccatcga atggcgcaaa acctttcgcg gtatggcatg
     3841 atagcgcccg gaagagagtc aattcagggt ggtgaatgtg aaaccagtaa cgttatacga
     3901 tgtcgcagag tatgccggtg tctcttatca gaccgtttcc cgcgtggtga accaggccag
     3961 ccacgtttct gcgaaaacgc gggaaaaagt ggaagcggcg atggcggagc tgaattacat
     4021 tcccaaccgc gtggcacaac aactggcggg caaacagtcg ttgctgattg gcgttgccac
     4081 ctccagtctg gccctgcacg cgccgtcgca aattgtcgcg gcgattaaat ctcgcgccga
     4141 tcaactgggt gccagcgtgg tggtgtcgat ggtagaacga agcggcgtcg aagcctgtaa
     4201 agcggcggtg cacaatcttc tcgcgcaacg cgtcagtggg ctgatcatta actatccgct
     4261 ggatgaccag gatgccattg ctgtggaagc tgcctgcact aatgttccgg cgttatttct
     4321 tgatgtctct gaccagacac ccatcaacag tattattttc tcccatgaag acggtacgcg
     4381 actgggcgtg gagcatctgg tcgcattggg tcaccagcaa atcgcgctgt tagcgggccc
     4441 attaagttct gtctcggcgc gtctgcgtct ggctggctgg cataaatatc tcactcgcaa
     4501 tcaaattcag ccgatagcgg aacgggaagg cgactggagt gccatgtccg gttttcaaca
     4561 aaccatgcaa atgctgaatg agggcatcgt tcccactgcg atgctggttg ccaacgatca
     4621 gatggcgctg ggcgcaatgc gcgccattac cgagtccggg ctgcgcgttg gtgcggatat
     4681 ctcggtagtg ggatacgacg ataccgaaga cagctcatgt tatatcccgc cgttaaccac
     4741 catcaaacag gattttcgcc tgctggggca aaccagcgtg gaccgcttgc tgcaactctc
     4801 tcagggccag gcggtgaagg gcaatcagct gttgcccgtc tcactggtga aaagaaaaac
     4861 cacgctagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca
     4921 gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtaa
     4981 gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt
     5041 gtggaattgt gagcggataa caatttcaca caggaaacag ctatgaccat gattacggat
     5101 tcactggccg tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat
     5161 cgccttgcag cacatccccc tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat
     5221 cgcccttccc aacagttgcg cagcctgaat ggcgaatggc gctttgcctg gtttccggca
     5281 ccagaagcgg tgccggaaag ctggctggag tgcgatcttc ctgaggccga tactgtcgtc
     5341 gtcccctcaa actggcagat gcacggttac gatgcgccca tctacaccaa cgtgacctat
     5401 cccattacgg tcaatccgcc gtttgttccc acggagaatc cgacgggttg ttactcgctc
     5461 acatttaatg ttgatgaaag ctggctacag gaaggccaga cgcgaattat ttttgatggc
     5521 gttggaatta gcttatcgac tgcacggtgc accaatgctt ctggcgtcag gcagccatcg
     5581 gaagctgtgg tatggctgtg caggtc
//
