LOCUS       sequence-310693-        5892 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses ClpXP-degradable variant of Anabaena flos-aquae GvpC in 
            E.coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5892)
  AUTHORS   Lakshmanan A, Jin Z, Nety SP, Sawyer DP, Lee-Gosselin A, Malounda D,
            Swift MB, Maresca D, Shapiro MG
  TITLE     Acoustic biosensors for ultrasound imaging of enzyme activity.
  JOURNAL   Nat Chem Biol. 2020 Sep;16(9):988-996. doi: 
            10.1038/s41589-020-0591-0. Epub 2020 Jul 13.
  PUBMED    32661379
REFERENCE   2  (bases 1 to 5892)
  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: 
            "10.1038/s41589-020-0591-0"; journalName: "Nat Chem Biol"; date: 
            "2020-09"; volume: "16"; issue: "9"; pages: "988-996"
COMMENT     Sequence Label: N-His-GvpC-ssrA
FEATURES             Location/Qualifiers
     source          1..5892
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     4..23
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     107..126
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        107..125
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    126..150
                     /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             165..187
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             200..217
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             863..880
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(933..951)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      947..994
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1031..1486
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1118..1137)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1328..1349
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1578..2393)
                     /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             complement(1578..2393)
                     /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     2301..2320
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2385..2404
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2515..3103
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3004..3023
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3257..3274
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3289..3428
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3354..3373)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3514..3536
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3530..3721)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4530..5612)
                     /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     5574..5593
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5613..5690)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 atcggtgatg tcggcgatat aggcgccagc aaccgcacct gtggcgccgg tgatgccggc
       61 cacgatgcgt ccggcgtaga ggatcgagat ctcgatcccg cgaaattaat acgactcact
      121 ataggggaat tgtgagcgga taacaattcc cctctagaaa taattttgtt taactttaag
      181 aaggagatat accatgggac atcaccatca ccatcacagc gggatatctc tgatggcaaa
      241 aattcgtcaa gaacatcaga gcattgcaga aaaagttgca gaactgagcc tggaaacccg
      301 tgaatttctg agcgttacca ccgcaaaacg ccaagaacag gcagaaaaac aggcacaaga
      361 actgcaggca ttctataaag atctgcaaga aaccagccag cagtttctga gtgaaaccgc
      421 acaggcacgt attgcacagg ccgagaaaca agcccaagag ctgctggcct ttcacaaaga
      481 actgcaagag acttctcagc agttcctgtc agcaacagcc caagcccgta tcgcccaagc
      541 ggagaaacag gcgcaagagc tgctggcatt ttatcaagaa gttcgcgaaa cctcacagca
      601 atttctgtca gccactgcgc aagcacgcat tgcgcaagct gaaaaacaag cgcaagagct
      661 gctggcgttc cataaagagc tgcaagaaac gtcacaacag tttctgagcg caacggccga
      721 tgcacgtaca gcacaggcga aagaacagaa agaaagcctg ctgaaatttc gtcaggacct
      781 gtttgttagc attttcggca gcggtgcagc aaacgacgaa aactacgctt tagcagctta
      841 gaagcttgcg gccgcactcg agcaccacca ccaccaccac tgagatccgg ctgctaacaa
      901 agcccgaaag gaagctgagt tggctgctgc caccgctgag caataactag cataacccct
      961 tggggcctct aaacgggtct tgaggggttt tttgctgaaa ggaggaacta tatccggatt
     1021 ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     1081 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
     1141 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     1201 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca
     1261 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     1321 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct
     1381 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
     1441 caaaaattta acgcgaattt taacaaaata ttaacgctta caatttaggt ggcacttttc
     1501 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     1561 cgctcatgaa ttaattctta gaaaaactca tcgagcatca aatgaaactg caatttattc
     1621 atatcaggat tatcaatacc atatttttga aaaagccgtt tctgtaatga aggagaaaac
     1681 tcaccgaggc agttccatag gatggcaaga tcctggtatc ggtctgcgat tccgactcgt
     1741 ccaacatcaa tacaacctat taatttcccc tcgtcaaaaa taaggttatc aagtgagaaa
     1801 tcaccatgag tgacgactga atccggtgag aatggcaaaa gtttatgcat ttctttccag
     1861 acttgttcaa caggccagcc attacgctcg tcatcaaaat cactcgcatc aaccaaaccg
     1921 ttattcattc gtgattgcgc ctgagcgaga cgaaatacgc gatcgctgtt aaaaggacaa
     1981 ttacaaacag gaatcgaatg caaccggcgc aggaacactg ccagcgcatc aacaatattt
     2041 tcacctgaat caggatattc ttctaatacc tggaatgctg ttttcccggg gatcgcagtg
     2101 gtgagtaacc atgcatcatc aggagtacgg ataaaatgct tgatggtcgg aagaggcata
     2161 aattccgtca gccagtttag tctgaccatc tcatctgtaa catcattggc aacgctacct
     2221 ttgccatgtt tcagaaacaa ctctggcgca tcgggcttcc catacaatcg atagattgtc
     2281 gcacctgatt gcccgacatt atcgcgagcc catttatacc catataaatc agcatccatg
     2341 ttggaattta atcgcggcct agagcaagac gtttcccgtt gaatatggct cataacaccc
     2401 cttgtattac tgtttatgta agcagacagt tttattgttc atgaccaaaa tcccttaacg
     2461 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     2521 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     2581 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     2641 agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa
     2701 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     2761 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     2821 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     2881 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     2941 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     3001 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     3061 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     3121 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     3181 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     3241 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc tgatgcggta
     3301 ttttctcctt acgcatctgt gcggtatttc acaccgcaat ggtgcactct cagtacaatc
     3361 tgctctgatg ccgcatagtt aagccagtat acactccgct atcgctacgt gactgggtca
     3421 tggctgcgcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc
     3481 cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt
     3541 caccgtcatc accgaaacgc gcgaggcagc tgcggtaaag ctcatcagcg tggtcgtgaa
     3601 gcgattcaca gatgtctgcc tgttcatccg cgtccagctc gttgagtttc tccagaagcg
     3661 ttaatgtctg gcttctgata aagcgggcca tgttaagggc ggttttttcc tgtttggtca
     3721 ctgatgcctc cgtgtaaggg ggatttctgt tcatgggggt aatgataccg atgaaacgag
     3781 agaggatgct cacgatacgg gttactgatg atgaacatgc ccggttactg gaacgttgtg
     3841 agggtaaaca actggcggta tggatgcggc gggaccagag aaaaatcact cagggtcaat
     3901 gccagcgctt cgttaataca gatgtaggtg ttccacaggg tagccagcag catcctgcga
     3961 tgcagatccg gaacataatg gtgcagggcg ctgacttccg cgtttccaga ctttacgaaa
     4021 cacggaaacc gaagaccatt catgttgttg ctcaggtcgc agacgttttg cagcagcagt
     4081 cgcttcacgt tcgctcgcgt atcggtgatt cattctgcta accagtaagg caaccccgcc
     4141 agcctagccg ggtcctcaac gacaggagca cgatcatgcg cacccgtggg gccgccatgc
     4201 cggcgataat ggcctgcttc tcgccgaaac gtttggtggc gggaccagtg acgaaggctt
     4261 gagcgagggc gtgcaagatt ccgaataccg caagcgacag gccgatcatc gtcgcgctcc
     4321 agcgaaagcg gtcctcgccg aaaatgaccc agagcgctgc cggcacctgt cctacgagtt
     4381 gcatgataaa gaagacagtc ataagtgcgg cgacgatagt catgccccgc gcccaccgga
     4441 aggagctgac tgggttgaag gctctcaagg gcatcggtcg agatcccggt gcctaatgag
     4501 tgagctaact tacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     4561 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     4621 gccagggtgg tttttctttt caccagtgag acgggcaaca gctgattgcc cttcaccgcc
     4681 tggccctgag agagttgcag caagcggtcc acgctggttt gccccagcag gcgaaaatcc
     4741 tgtttgatgg tggttaacgg cgggatataa catgagctgt cttcggtatc gtcgtatccc
     4801 actaccgaga tatccgcacc aacgcgcagc ccggactcgg taatggcgcg cattgcgccc
     4861 agcgccatct gatcgttggc aaccagcatc gcagtgggaa cgatgccctc attcagcatt
     4921 tgcatggttt gttgaaaacc ggacatggca ctccagtcgc cttcccgttc cgctatcggc
     4981 tgaatttgat tgcgagtgag atatttatgc cagccagcca gacgcagacg cgccgagaca
     5041 gaacttaatg ggcccgctaa cagcgcgatt tgctggtgac ccaatgcgac cagatgctcc
     5101 acgcccagtc gcgtaccgtc ttcatgggag aaaataatac tgttgatggg tgtctggtca
     5161 gagacatcaa gaaataacgc cggaacatta gtgcaggcag cttccacagc aatggcatcc
     5221 tggtcatcca gcggatagtt aatgatcagc ccactgacgc gttgcgcgag aagattgtgc
     5281 accgccgctt tacaggcttc gacgccgctt cgttctacca tcgacaccac cacgctggca
     5341 cccagttgat cggcgcgaga tttaatcgcc gcgacaattt gcgacggcgc gtgcagggcc
     5401 agactggagg tggcaacgcc aatcagcaac gactgtttgc ccgccagttg ttgtgccacg
     5461 cggttgggaa tgtaattcag ctccgccatc gccgcttcca ctttttcccg cgttttcgca
     5521 gaaacgtggc tggcctggtt caccacgcgg gaaacggtct gataagagac accggcatac
     5581 tctgcgacat cgtataacgt tactggtttc acattcacca ccctgaattg actctcttcc
     5641 gggcgctatc atgccatacc gcgaaaggtt ttgcgccatt cgatggtgtc cgggatctcg
     5701 acgctctccc ttatgcgact cctgcattag gaagcagccc agtagtaggt tgaggccgtt
     5761 gagcaccgcc gccgcaagga atggtgcatg caaggagatg gcgcccaaca gtcccccggc
     5821 cacggggcct gccaccatac ccacgccgaa acaagcgctc atgagcccga agtggcgagc
     5881 ccgatcttcc cc
//
