LOCUS       sequence-292227-        6035 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6035)
  AUTHORS   Kobashigawa Y, Kumeta H, Ogura K, Inagaki F
  TITLE     Attachment of an NMR-invisible solubility enhancement tag using a 
            sortase-mediated protein ligation method.
  JOURNAL   J Biomol NMR. 2009 Mar . 43(3):145-50.
  PUBMED    19140010
REFERENCE   2  (bases 1 to 6035)
  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: "J Biomol 
            NMR. 2009 Mar . 43(3):145-50."
COMMENT     Sequence Label: pGBHPS-SortA
FEATURES             Location/Qualifiers
     source          1..6035
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    20..44
                     /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             59..81
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             94..255
                     /codon_start=1
                     /product="B1 domain of Streptococcal protein G   "
                     /label=GB1
                     /note="effective as a solubilizing fusion partner (Cheng 
                     and Patel, 2004)"
                     /translation="YKLILNGKTLKGETTTEAVDAATAEKVFKQYANDNGVDGEWTYDD
                     ATKTFTVTE"
     CDS             262..279
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             280..303
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             767..784
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(837..855)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      851..898
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      935..1390
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1022..1041)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1232..1253
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1416..1520
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1521..1589,1590..2381)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1739..1758)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2552..3140
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3041..3060
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3294..3311
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3326..3465
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3391..3410)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3551..3573
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3567..3758)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4567..5649)
                     /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     5611..5630
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5650..5727)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5933..5952
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atataccatg gagtacaaac tgatcctgaa cggtaaaacc
      121 ctgaaaggtg aaaccaccac cgaagctgtt gacgctgcta ccgcggaaaa agttttcaaa
      181 cagtacgcta acgacaacgg tgttgacggt gaatggacct acgacgacgc taccaaaacc
      241 ttcacggtaa ccgaacatag ccatcatcat catcatcatc tggaagtgct gtttcagggc
      301 ccgcatatgg gatcccaagc taaacctcaa attccgaaag ataaatcaaa agtggcaggc
      361 tatattgaaa ttccagatgc tgatattaaa gaaccagtat atccaggacc agcaacacct
      421 gaacaattaa atagaggtgt aagctttgca gaagaaaatg aatcactaga tgatcaaaat
      481 atttcaattg caggacacac tttcattgac cgtccgaact atcaatttac aaatcttaaa
      541 gcagccaaaa aaggtagtat ggtgtacttt aaagttggta atgaaacacg taagtataaa
      601 atgacaagta taagagatgt taagccaaca gatgtagaag ttctagatga acaaaaaggt
      661 aaagataaac aattaacatt aattacttgt gatgattaca atgaaaagac aggcgtttgg
      721 gaaaaacgta aaatctttgt agctacagaa gtcaaataat ctcgagcacc accaccacca
      781 ccactgagat ccggctgcta acaaagcccg aaaggaagct gagttggctg ctgccaccgc
      841 tgagcaataa ctagcataac cccttggggc ctctaaacgg gtcttgaggg gttttttgct
      901 gaaaggagga actatatccg gattggcgaa tgggacgcgc cctgtagcgg cgcattaagc
      961 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc
     1021 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
     1081 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
     1141 aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc
     1201 cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca
     1261 ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat
     1321 tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg
     1381 cttacaattt aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     1441 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     1501 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
     1561 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     1621 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     1681 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
     1741 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
     1801 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
     1861 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     1921 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
     1981 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     2041 acgagcgtga caccacgatg cctgcagcaa tggcaacaac gttgcgcaaa ctattaactg
     2101 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     2161 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     2221 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
     2281 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     2341 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     2401 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     2461 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     2521 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     2581 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     2641 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc
     2701 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     2761 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     2821 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     2881 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     2941 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     3001 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     3061 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     3121 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     3181 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     3241 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     3301 cagtgagcga ggaagcggaa gagcgcctga tgcggtattt tctccttacg catctgtgcg
     3361 gtatttcaca ccgcaatggt gcactctcag tacaatctgc tctgatgccg catagttaag
     3421 ccagtataca ctccgctatc gctacgtgac tgggtcatgg ctgcgccccg acacccgcca
     3481 acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta cagacaagct
     3541 gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg
     3601 aggcagctgc ggtaaagctc atcagcgtgg tcgtgaagcg attcacagat gtctgcctgt
     3661 tcatccgcgt ccagctcgtt gagtttctcc agaagcgtta atgtctggct tctgataaag
     3721 cgggccatgt taagggcggt tttttcctgt ttggtcactg atgcctccgt gtaaggggga
     3781 tttctgttca tgggggtaat gataccgatg aaacgagaga ggatgctcac gatacgggtt
     3841 actgatgatg aacatgcccg gttactggaa cgttgtgagg gtaaacaact ggcggtatgg
     3901 atgcggcggg accagagaaa aatcactcag ggtcaatgcc agcgcttcgt taatacagat
     3961 gtaggtgttc cacagggtag ccagcagcat cctgcgatgc agatccggaa cataatggtg
     4021 cagggcgctg acttccgcgt ttccagactt tacgaaacac ggaaaccgaa gaccattcat
     4081 gttgttgctc aggtcgcaga cgttttgcag cagcagtcgc ttcacgttcg ctcgcgtatc
     4141 ggtgattcat tctgctaacc agtaaggcaa ccccgccagc ctagccgggt cctcaacgac
     4201 aggagcacga tcatgcgcac ccgtggggcc gccatgccgg cgataatggc ctgcttctcg
     4261 ccgaaacgtt tggtggcggg accagtgacg aaggcttgag cgagggcgtg caagattccg
     4321 aataccgcaa gcgacaggcc gatcatcgtc gcgctccagc gaaagcggtc ctcgccgaaa
     4381 atgacccaga gcgctgccgg cacctgtcct acgagttgca tgataaagaa gacagtcata
     4441 agtgcggcga cgatagtcat gccccgcgcc caccggaagg agctgactgg gttgaaggct
     4501 ctcaagggca tcggtcgaga tcccggtgcc taatgagtga gctaacttac attaattgcg
     4561 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     4621 ggccaacgcg cggggagagg cggtttgcgt attgggcgcc agggtggttt ttcttttcac
     4681 cagtgagacg ggcaacagct gattgccctt caccgcctgg ccctgagaga gttgcagcaa
     4741 gcggtccacg ctggtttgcc ccagcaggcg aaaatcctgt ttgatggtgg ttaacggcgg
     4801 gatataacat gagctgtctt cggtatcgtc gtatcccact accgagatat ccgcaccaac
     4861 gcgcagcccg gactcggtaa tggcgcgcat tgcgcccagc gccatctgat cgttggcaac
     4921 cagcatcgca gtgggaacga tgccctcatt cagcatttgc atggtttgtt gaaaaccgga
     4981 catggcactc cagtcgcctt cccgttccgc tatcggctga atttgattgc gagtgagata
     5041 tttatgccag ccagccagac gcagacgcgc cgagacagaa cttaatgggc ccgctaacag
     5101 cgcgatttgc tggtgaccca atgcgaccag atgctccacg cccagtcgcg taccgtcttc
     5161 atgggagaaa ataatactgt tgatgggtgt ctggtcagag acatcaagaa ataacgccgg
     5221 aacattagtg caggcagctt ccacagcaat ggcatcctgg tcatccagcg gatagttaat
     5281 gatcagccca ctgacgcgtt gcgcgagaag attgtgcacc gccgctttac aggcttcgac
     5341 gccgcttcgt tctaccatcg acaccaccac gctggcaccc agttgatcgg cgcgagattt
     5401 aatcgccgcg acaatttgcg acggcgcgtg cagggccaga ctggaggtgg caacgccaat
     5461 cagcaacgac tgtttgcccg ccagttgttg tgccacgcgg ttgggaatgt aattcagctc
     5521 cgccatcgcc gcttccactt tttcccgcgt tttcgcagaa acgtggctgg cctggttcac
     5581 cacgcgggaa acggtctgat aagagacacc ggcatactct gcgacatcgt ataacgttac
     5641 tggtttcaca ttcaccaccc tgaattgact ctcttccggg cgctatcatg ccataccgcg
     5701 aaaggttttg cgccattcga tggtgtccgg gatctcgacg ctctccctta tgcgactcct
     5761 gcattaggaa gcagcccagt agtaggttga ggccgttgag caccgccgcc gcaaggaatg
     5821 gtgcatgcaa ggagatggcg cccaacagtc ccccggccac ggggcctgcc accataccca
     5881 cgccgaaaca agcgctcatg agcccgaagt ggcgagcccg atcttcccca tcggtgatgt
     5941 cggcgatata ggcgccagca accgcacctg tggcgccggt gatgccggcc acgatgcgtc
     6001 cggcgtagag gatcgagatc tcgatcccgc gaaat
//
