LOCUS       sequence-338465-        5200 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a specific DNA-RNA hybrid sensor protein with the HBD 
            domain of RNase H1 in bacterial cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5200)
  AUTHORS   Wang K, Wang H, Li C, Yin Z, Xiao R, Li Q, Xiang Y, Wang W, Huang J,
            Chen L, Fang P, Liang K
  TITLE     Genomic profiling of native R loops with a DNA-RNA hybrid 
            recognition sensor.
  JOURNAL   Sci Adv. 2021 Feb 17;7(8). pii: 7/8/eabe3516. doi: 
            10.1126/sciadv.abe3516. Print 2021 Feb.
  PUBMED    33597247
REFERENCE   2  (bases 1 to 5200)
  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: "Sci Adv."; 
            date: "2021-02-17"; volume: "7(8). pii: 7/8/eabe3516. doi"; pages: "
            10.1126/sciadv.abe3516. Print 2021 Feb"
COMMENT     Sequence Label: pGEX-GST-His6-1_HBD
FEATURES             Location/Qualifiers
     source          1..5200
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(293..298,299..316,317..323)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    331..347
                     /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     355..371
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     355..371
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             367..894
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(387..404)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(387..403)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(396..418)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     promoter        join(996..1001,1002..1017,1018..1024)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1032..1048
                     /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             1071..1724
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     1682..1704
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             1731..1748
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1779..1808
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             1821..1832
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(2063..2085)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2230..2248)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2316..2420
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2421..2489,2490..3281)
                     /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(2639..2658)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3452..4040
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3941..3960
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4194..4211
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4284..4361
                     /gene="lacI"
                     /label=lacI promoter
     CDS             join(4362..5200,1..244)
                     /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(4381..4400)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 atgttatatc ccgccgttaa ccaccatcaa acaggatttt cgcctgctgg ggcaaaccag
       61 cgtggaccgc ttgctgcaac tctctcaggg ccaggcggtg aagggcaatc agctgttgcc
      121 cgtctcactg gtgaaaagaa aaaccaccct ggcgcccaat acgcaaaccg cctctccccg
      181 cgcgttggcc gattcattaa tgcagctggc acgacaggtt tcccgactgg aaagcgggca
      241 gtgagcgcaa cgcaattaat gtaagttagc tcactcatta ggcaccccag gctttacact
      301 ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa
      361 acagctatga ccatgattac ggattcactg gccgtcgttt tacaacgtcg tgactgggaa
      421 aaccctggcg ttacccaact taatcgcctt gcagcacatc cccctttcgc cagctggcgt
      481 aatagcgaag aggcccgcac cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa
      541 tggcgctttg cctggtttcc ggcaccagaa gcggtgccgg aaagctggct ggagtgcgat
      601 cttcctgagg ccgatactgt cgtcgtcccc tcaaactggc agatgcacgg ttacgatgcg
      661 cccatctaca ccaacgtgac ctatcccatt acggtcaatc cgccgtttgt tcccacggag
      721 aatccgacgg gttgttactc gctcacattt aatgttgatg aaagctggct acaggaaggc
      781 cagacgcgaa ttatttttga tggcgttgga attagcttat cgactgcacg gtgcaccaat
      841 gcttctggcg tcaggcagcc atcggaagct gtggtatggc tgtgcaggtc gtaaatcact
      901 gcataattcg tgtcgctcaa ggcgcactcc cgttctggat aatgtttttt gcgccgacat
      961 cataacggtt ctggcaaata ttctgaaatg agctgttgac aattaatcat cggctcgtat
     1021 aatgtgtgga attgtgagcg gataacaatt tcacacagga aacagtattc atgtccccta
     1081 tactaggtta ttggaaaatt aagggccttg tgcaacccac tcgacttctt ttggaatatc
     1141 ttgaagaaaa atatgaagag catttgtatg agcgcgatga aggtgataaa tggcgaaaca
     1201 aaaagtttga attgggtttg gagtttccca atcttcctta ttatattgat ggtgatgtta
     1261 aattaacaca gtctatggcc atcatacgtt atatagctga caagcacaac atgttgggtg
     1321 gttgtccaaa agagcgtgca gagatttcaa tgcttgaagg agcggttttg gatattagat
     1381 acggtgtttc gagaattgca tatagtaaag actttgaaac tctcaaagtt gattttctta
     1441 gcaagctacc tgaaatgctg aaaatgttcg aagatcgttt atgtcataaa acatatttaa
     1501 atggtgatca tgtaacccat cctgacttca tgttgtatga cgctcttgat gttgttttat
     1561 acatggaccc aatgtgcctg gatgcgttcc caaaattagt ttgttttaaa aaacgtattg
     1621 aagctatccc acaaattgat aagtacttga aatccagcaa gtatatagca tggcctttgc
     1681 agggctggca agccacgttt ggtggtggcg accatcctcc aaaatcggat ctggttccgc
     1741 gtggatctcg tcgtgcatct gttggatata ccatgggcca tcatcatcat catcatcatc
     1801 atcatcacag cagcggccat atcgaaggtc gtcatatgat gttctatgcc gtgaggaggg
     1861 gccgcaagac cggggtcttt ctgacctgga atgagtgcag agcacaggtg gaccggtttc
     1921 ctgctgccag atttaagaag tttgccacag aggatgaggc ctgggccttt gtcaggaaat
     1981 ctgcaagcgg aggcggaggg ggaggaattc atcgtgactg actgacgatc tgcctcgcgc
     2041 gtttcggtga tgacggtgaa aacctctgac acatgcagct cccggagacg gtcacagctt
     2101 gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg
     2161 ggtgtcgggg cgcagccatg acccagtcac gtagcgatag cggagtgtat aattcttgaa
     2221 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
     2281 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     2341 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     2401 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
     2461 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     2521 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     2581 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
     2641 tatgtggcgc ggtattatcc cgtgttgacg ccgggcaaga gcaactcggt cgccgcatac
     2701 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
     2761 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     2821 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
     2881 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     2941 acgagcgtga caccacgatg cctgcagcaa tggcaacaac gttgcgcaaa ctattaactg
     3001 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     3061 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     3121 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
     3181 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     3241 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     3301 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     3361 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     3421 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     3481 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     3541 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc
     3601 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     3661 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     3721 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     3781 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     3841 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     3901 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     3961 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     4021 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     4081 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     4141 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     4201 cagtgagcga ggaagcggaa gagcgcctga tgcggtattt tctccttacg catctgtgcg
     4261 gtatttcaca ccgcataaat tccgacacca tcgaatggcg caaaaccttt cgcggtatgg
     4321 catgatagcg cccggaagag agtcaattca gggtggtgaa tgtgaaacca gtaacgttat
     4381 acgatgtcgc agagtatgcc ggtgtctctt atcagaccgt ttcccgcgtg gtgaaccagg
     4441 ccagccacgt ttctgcgaaa acgcgggaaa aagtggaagc ggcgatggcg gagctgaatt
     4501 acattcccaa ccgcgtggca caacaactgg cgggcaaaca gtcgttgctg attggcgttg
     4561 ccacctccag tctggccctg cacgcgccgt cgcaaattgt cgcggcgatt aaatctcgcg
     4621 ccgatcaact gggtgccagc gtggtggtgt cgatggtaga acgaagcggc gtcgaagcct
     4681 gtaaagcggc ggtgcacaat cttctcgcgc aacgcgtcag tgggctgatc attaactatc
     4741 cgctggatga ccaggatgcc attgctgtgg aagctgcctg cactaatgtt ccggcgttat
     4801 ttcttgatgt ctctgaccag acacccatca acagtattat tttctcccat gaagacggta
     4861 cgcgactggg cgtggagcat ctggtcgcat tgggtcacca gcaaatcgcg ctgttagcgg
     4921 gcccattaag ttctgtctcg gcgcgtctgc gtctggctgg ctggcataaa tatctcactc
     4981 gcaatcaaat tcagccgata gcggaacggg aaggcgactg gagtgccatg tccggttttc
     5041 aacaaaccat gcaaatgctg aatgagggca tcgttcccac tgcgatgctg gttgccaacg
     5101 atcagatggc gctgggcgca atgcgcgcca ttaccgagtc cgggctgcgc gttggtgcgg
     5161 atatctcggt agtgggatac gacgataccg aagacagctc
//
