LOCUS       sequence-338473-        7655 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses protein A and Tn5 Transposase fusion protein in bacterial 
            cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7655)
  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 7655)
  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: pA-His6-Tn5
FEATURES             Location/Qualifiers
     source          1..7655
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(479..1561)
                     /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     1523..1542
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1562..1639)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1845..1864
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1948..1967
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1948..1966
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1967..1991
                     /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             2006..2028
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2041..2070
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             2083..2094
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             2116..2541
                     /codon_start=1
                     /product="immunoglobulin binding domains of Staphylocuccus 
                     aureus protein A"
                     /label=protein A
                     /translation="SLKDDPSQSANLLSEAKKLNESQAPKADNKFNKEQQNAFYEILHL
                     PNLNEEQRNGFIQSLKDDPSQSANLLAEAKKLNDAQAPKADNKFNKEQQNAFYEILHLP
                     NLTEEQRNGFIQSLKDDPSVSKEILAEAKKLNDAQAPK"
     CDS             2623..4050
                     /codon_start=1
                     /product="transposase from the bacterial Tn5 transposon 
                     (Reznikoff, 1993)"
                     /label=Tn5 transposase
                     /translation="MITSALHRAADWAKSVFSSAALGDPRRTARLVNVAAQLAKYSGKS
                     ITISSEGSKAMQEGAYRFIRNPNVSAEAIRKAGAMQTVKLAQEFPELLAIEDTTSLSYR
                     HQVAEELGKLGSIQDKSRGWWVHSVLLLEATTFRTVGLLHQEWWMRPDDPADADEKESG
                     KWLAAAATSRLRMGSMMSNVIAVCDREADIHAYLQDKLAHNERFVVRSKHPRKDVESGL
                     YLYDHLKNQPELGGYQISIPQKGVVDKRGKRKNRPARKASLSLRSGRITLKQGNITLNA
                     VLAEEINPPKGETPLKWLLLTSEPVESLAQALRVIDIYTHRWRIEEFHKAWKTGAGAER
                     QRMEEPDNLERMVSILSFVAVRLLQLRESFTPPQALRAQGLLKEAEHVESQSAETVLTP
                     DECQLLGYLDKGKRKRKEKAGSLQWAYMAIARLGGFMDSKRTGIASWGALWEGWEALQS
                     KLDGFLAAKDLMAQGIKI"
     primer_bind     complement(4103..4121)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      4117..4164
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(4339..4344,4345..4361,4362..4367))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(4394..4412)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4480..4584
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4585..4653,4654..5445)
                     /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(4803..4822)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5616..6204
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6105..6124
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6358..6375
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    6390..6529
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(6455..6474)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6615..6637
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(6631..6822)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 atggtcgtca tctacctgcc tggacagcat ggcctgcaac gcgggcatcc cgatgccgcc
       61 ggaagcgaga agaatcataa tggggaaggc catccagcct cgcgtcgcga acgccagcaa
      121 gacgtagccc agcgcgtcgg ccgccatgcc ggcgataatg gcctgcttct cgccgaaacg
      181 tttggtggcg ggaccagtga cgaaggcttg agcgagggcg tgcaagattc cgaataccgc
      241 aagcgacagg ccgatcatcg tcgcgctcca gcgaaagcgg tcctcgccga aaatgaccca
      301 gagcgctgcc ggcacctgtc ctacgagttg catgataaag aagacagtca taagtgcggc
      361 gacgatagtc atgccccgcg cccaccggaa ggagctgact gggttgaagg ctctcaaggg
      421 catcggtcga gatcccggtg cctaatgagt gagctaactt acattaattg cgttgcgctc
      481 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
      541 cgcggggaga ggcggtttgc gtattgggcg ccagggtggt ttttcttttc accagtgaga
      601 cgggcaacag ctgattgccc ttcaccgcct ggccctgaga gagttgcagc aagcggtcca
      661 cgctggtttg ccccagcagg cgaaaatcct gtttgatggt ggttaacggc gggatataac
      721 atgagctgtc ttcggtatcg tcgtatccca ctaccgagat atccgcacca acgcgcagcc
      781 cggactcggt aatggcgcgc attgcgccca gcgccatctg atcgttggca accagcatcg
      841 cagtgggaac gatgccctca ttcagcattt gcatggtttg ttgaaaaccg gacatggcac
      901 tccagtcgcc ttcccgttcc gctatcggct gaatttgatt gcgagtgaga tatttatgcc
      961 agccagccag acgcagacgc gccgagacag aacttaatgg gcccgctaac agcgcgattt
     1021 gctggtgacc caatgcgacc agatgctcca cgcccagtcg cgtaccgtct tcatgggaga
     1081 aaataatact gttgatgggt gtctggtcag agacatcaag aaataacgcc ggaacattag
     1141 tgcaggcagc ttccacagca atggcatcct ggtcatccag cggatagtta atgatcagcc
     1201 cactgacgcg ttgcgcgaga agattgtgca ccgccgcttt acaggcttcg acgccgcttc
     1261 gttctaccat cgacaccacc acgctggcac ccagttgatc ggcgcgagat ttaatcgccg
     1321 cgacaatttg cgacggcgcg tgcagggcca gactggaggt ggcaacgcca atcagcaacg
     1381 actgtttgcc cgccagttgt tgtgccacgc ggttgggaat gtaattcagc tccgccatcg
     1441 ccgcttccac tttttcccgc gttttcgcag aaacgtggct ggcctggttc accacgcggg
     1501 aaacggtctg ataagagaca ccggcatact ctgcgacatc gtataacgtt actggtttca
     1561 cattcaccac cctgaattga ctctcttccg ggcgctatca tgccataccg cgaaaggttt
     1621 tgcgccattc gatggtgtcc gggatctcga cgctctccct tatgcgactc ctgcattagg
     1681 aagcagccca gtagtaggtt gaggccgttg agcaccgccg ccgcaaggaa tggtgcatgc
     1741 aaggagatgg cgcccaacag tcccccggcc acggggcctg ccaccatacc cacgccgaaa
     1801 caagcgctca tgagcccgaa gtggcgagcc cgatcttccc catcggtgat gtcggcgata
     1861 taggcgccag caaccgcacc tgtggcgccg gtgatgccgg ccacgatgcg tccggcgtag
     1921 aggatcgaga tctcgatccc gcgaaattaa tacgactcac tataggggaa ttgtgagcgg
     1981 ataacaattc ccctctagaa ataattttgt ttaactttaa gaaggagata taccatgggc
     2041 catcatcatc atcatcatca tcatcatcac agcagcggcc atatcgaagg tcgtcatatg
     2101 accatgatta cgccaagctt aaaagatgac ccaagccaaa gtgctaacct attgtcagaa
     2161 gctaaaaagt taaatgaatc tcaagcaccg aaagcggata acaaattcaa caaagaacaa
     2221 caaaatgctt tctatgaaat cttacattta cctaacttaa acgaagaaca acgcaatggt
     2281 ttcatccaaa gcctaaaaga tgacccaagc caaagcgcta accttttagc agaagctaaa
     2341 aagctaaatg atgctcaagc accaaaagct gacaacaaat tcaacaaaga acaacaaaat
     2401 gctttctatg aaattttaca tttacctaac ttaactgaag aacaacgtaa cggcttcatc
     2461 caaagcctta aagacgatcc ttcagtgagc aaagaaattt tagcagaagc taaaaagcta
     2521 aacgatgctc aagcaccaaa agatgacgat aaagaattcg gtggcggtgg ctctggcggt
     2581 ggtgggagtg gaggtggggg atcaggagga ggcggttccc atatgattac cagtgcactg
     2641 catcgtgcgg cggattgggc gaaaagcgtg ttttctagtg ctgcgctggg tgatccgcgt
     2701 cgtaccgcgc gtctggtgaa tgttgcggcg caactggcca aatatagcgg caaaagcatt
     2761 accattagca gcgaaggcag caaagccatg caggaaggcg cgtatcgttt tattcgtaat
     2821 ccgaacgtga gcgcggaagc gattcgtaaa gcgggtgcca tgcagaccgt gaaactggcc
     2881 caggaatttc cggaactgct ggcaattgaa gataccacct ctctgagcta tcgtcatcag
     2941 gtggcggaag aactgggcaa actgggtagc attcaggata aaagccgtgg ttggtgggtg
     3001 catagcgtgc tgctgctgga agcgaccacc tttcgtaccg tgggcctgct gcatcaagaa
     3061 tggtggatgc gtccggatga tccggcggat gcggatgaaa aagaaagcgg caaatggctg
     3121 gccgctgctg caacttcgcg tctgagaatg ggcagcatga tgagcaacgt gattgcggtg
     3181 tgcgatcgtg aagcggatat tcatgcgtat ctgcaagata aactggccca taacgaacgt
     3241 tttgtggtgc gtagcaaaca tccgcgtaaa gatgtggaaa gcggcctgta tctgtatgat
     3301 cacctgaaaa accagccgga actgggcggc tatcagatta gcattccgca gaaaggcgtg
     3361 gtggataaac gtggcaaacg taaaaaccgt ccggcgcgta aagcgagcct gagcctgcgt
     3421 agcggccgta ttaccctgaa acagggcaac attaccctga acgcggtgct ggccgaagaa
     3481 attaatccgc cgaaaggcga aaccccgctg aaatggctgc tgctgaccag cgagccggtg
     3541 gaaagtctgg cccaagcgct gcgtgtgatt gatatttata cccatcgttg gcgcattgaa
     3601 gaatttcaca aagcgtggaa aacgggtgcg ggtgcggaac gtcagcgtat ggaagaaccg
     3661 gataacctgg aacgtatggt gagcattctg agctttgtgg cggtgcgtct gctgcaactg
     3721 cgtgaatctt ttactccgcc gcaagcactg cgtgcgcagg gcctgctgaa agaagcggaa
     3781 cacgttgaaa gccagagcgc ggaaaccgtg ctgaccccgg atgaatgcca actgctgggc
     3841 tatctggata aaggcaaacg caaacgcaaa gaaaaagcgg gcagcctgca atgggcgtat
     3901 atggcgattg cgcgtctggg cggctttatg gatagcaaac gtaccggcat tgcgagctgg
     3961 ggtgcgctgt gggaaggttg ggaagcgctg caaagcaaac tggatggctt tctggccgcg
     4021 aaagacctga tggcgcaggg cattaaaatc taagatccgg ctgctaacaa agcccgaaag
     4081 gaagctgagt tggctgctgc caccgctgag caataactag cataacccct tggggcctct
     4141 aaacgggtct tgaggggttt tttgctgaaa ggaggaacta tatccggata tcccgcaaga
     4201 ggcccggcag taccggcata accaagccta tgcctacagc atccagggtg acggtgccga
     4261 ggatgacgat gagcgcattg ttagatttca tacacggtgc ctgactgcgt tagcaattta
     4321 actgtgataa actaccgcat taaagcttat cgatgataag ctgtcaaaca tgagaattct
     4381 tgaagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg
     4441 gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     4501 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     4561 caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
     4621 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
     4681 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     4741 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     4801 ctgctatgtg gcgcggtatt atcccgtgtt gacgccgggc aagagcaact cggtcgccgc
     4861 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     4921 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     4981 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     5041 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     5101 aacgacgagc gtgacaccac gatgcctgca gcaatggcaa caacgttgcg caaactatta
     5161 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     5221 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     5281 tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag
     5341 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     5401 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     5461 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     5521 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     5581 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     5641 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     5701 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     5761 gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     5821 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     5881 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     5941 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     6001 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     6061 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     6121 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     6181 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     6241 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     6301 cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc
     6361 gagtcagtga gcgaggaagc ggaagagcgc ctgatgcggt attttctcct tacgcatctg
     6421 tgcggtattt cacaccgcaa tggtgcactc tcagtacaat ctgctctgat gccgcatagt
     6481 taagccagta tacactccgc tatcgctacg tgactgggtc atggctgcgc cccgacaccc
     6541 gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca
     6601 agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg
     6661 cgcgaggcag ctgcggtaaa gctcatcagc gtggtcgtga agcgattcac agatgtctgc
     6721 ctgttcatcc gcgtccagct cgttgagttt ctccagaagc gttaatgtct ggcttctgat
     6781 aaagcgggcc atgttaaggg cggttttttc ctgtttggtc actgatgcct ccgtgtaagg
     6841 gggatttctg ttcatggggg taatgatacc gatgaaacga gagaggatgc tcacgatacg
     6901 ggttactgat gatgaacatg cccggttact ggaacgttgt gagggtaaac aactggcggt
     6961 atggatgcgg cgggaccaga gaaaaatcac tcagggtcaa tgccagcgct tcgttaatac
     7021 agatgtaggt gttccacagg gtagccagca gcatcctgcg atgcagatcc ggaacataat
     7081 ggtgcagggc gctgacttcc gcgtttccag actttacgaa acacggaaac cgaagaccat
     7141 tcatgttgtt gctcaggtcg cagacgtttt gcagcagcag tcgcttcacg ttcgctcgcg
     7201 tatcggtgat tcattctgct aaccagtaag gcaaccccgc cagcctagcc gggtcctcaa
     7261 cgacaggagc acgatcatgc gcacccgtgg ccaggaccca acgctgcccg agatgcgccg
     7321 cgtgcggctg ctggagatgg cggacgcgat ggatatgttc tgccaagggt tggtttgcgc
     7381 attcacagtt ctccgcaaga attgattggc tccaattctt ggagtggtga atccgttagc
     7441 gaggtgccgc cggcttccat tcaggtcgag gtggcccggc tccatgcacc gcgacgcaac
     7501 gcggggaggc agacaaggta tagggcggcg cctacaatcc atgccaaccc gttccatgtg
     7561 ctcgccgagg cggcataaat cgccgtgacg atcagcggtc caatgatcga agttaggctg
     7621 gtaagagccg cgagcgatcc ttgaagctgt ccctg
//
