LOCUS       sequence-445016-        5573 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of GvpK-Linker-Small BiT.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5573)
  AUTHORS   Iburg M, Anderson AP, Wong VT, Anton ED, He A, Lu GJ
  TITLE     Elucidating the assembly of gas vesicles by systematic 
            protein-protein interaction analysis.
  JOURNAL   EMBO J. 2024 Oct;43(19):4156-4172. doi: 10.1038/s44318-024-00178-2. 
            Epub 2024 Sep 3.
  PUBMED    39227754
REFERENCE   2  (bases 1 to 5573)
  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/s44318-024-00178-2"; journalName: "EMBO J"; date: 
            "2024-10"; volume: "43"; issue: "19"; pages: "4156-4172"
COMMENT     Sequence Label: pET26b(modified)-K-Sm
FEATURES             Location/Qualifiers
     source          1..5573
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     41..62
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(291..1106)
                     /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(291..1106)
                     /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     1014..1033
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     1098..1117
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      1228..1773
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     1674..1693
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1939..1956
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    1971..2110
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2036..2055)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2196..2218
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2212..2403)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(3212..4294)
                     /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     4256..4275
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4295..4372)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     4578..4597
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     4681..4700
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4681..4699
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    4700..4724
                     /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             4739..4761
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     primer_bind     complement(5219..5237)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      5233..5280
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      join(5317..5573,1..199)
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5404..5423)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
       61 ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt
      121 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
      181 atattaacgc ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
      241 gtttattttt ctaaatacat tcaaatatgt atccgctcat gaattaattc ttagaaaaac
      301 tcatcgagca tcaaatgaaa ctgcaattta ttcatatcag gattatcaat accatatttt
      361 tgaaaaagcc gtttctgtaa tgaaggagaa aactcaccga ggcagttcca taggatggca
      421 agatcctggt atcggtctgc gattccgact cgtccaacat caatacaacc tattaatttc
      481 ccctcgtcaa aaataaggtt atcaagtgag aaatcaccat gagtgacgac tgaatccggt
      541 gagaatggca aaagtttatg catttctttc cagacttgtt caacaggcca gccattacgc
      601 tcgtcatcaa aatcactcgc atcaaccaaa ccgttattca ttcgtgattg cgcctgagcg
      661 agacgaaata cgcgatcgct gttaaaagga caattacaaa caggaatcga atgcaaccgg
      721 cgcaggaaca ctgccagcgc atcaacaata ttttcacctg aatcaggata ttcttctaat
      781 acctggaatg ctgttttccc ggggatcgca gtggtgagta accatgcatc atcaggagta
      841 cggataaaat gcttgatggt cggaagaggc ataaattccg tcagccagtt tagtctgacc
      901 atctcatctg taacatcatt ggcaacgcta cctttgccat gtttcagaaa caactctggc
      961 gcatcgggct tcccatacaa tcgatagatt gtcgcacctg attgcccgac attatcgcga
     1021 gcccatttat acccatataa atcagcatcc atgttggaat ttaatcgcgg cctagagcaa
     1081 gacgtttccc gttgaatatg gctcataaca ccccttgtat tactgtttat gtaagcagac
     1141 agttttattg ttcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     1201 ccccgtagaa aagatcaaag gatcttcttg agatcgtttt ggtctgcgcg taatctcttg
     1261 ctctgaaaac gaaaaaaccg ccttgcaggt cggtttttcg aaggttctct gaactaccaa
     1321 ctctttgaac cgaggtaact ggcttggagg agcgcagtca ccaaaacttg tcctttcagt
     1381 ttagccttaa ccggcgcatg acttcaagac taactcctct aaatcaatta ccagtggctg
     1441 ctgccagtgg tgcttttgca tgtctttccg ggttggactc aagacgatag ttaccggata
     1501 aggcgcagcg gtcggactga acggggggtt cgtgcataca gtccagcttg gagcgaactg
     1561 cctacccgga actgagtgtc aggcgtggaa tgagacaaac gcggccataa cagcggaatg
     1621 acaccggtaa accgaaaggc aggaacagga gagcgcacga gggagccgcc agggggaaac
     1681 gcctggtatc tttatagtcc tgtcgggttt cgccaccact gatttgagcg tcagatttcg
     1741 tgatgcttgt caggggggcg gagcctatgg aaaaacgtat ggaaaaacgc cagcaacgcg
     1801 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     1861 tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc
     1921 agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cctgatgcgg
     1981 tattttctcc ttacgcatct gtgcggtatt tcacaccgca atggtgcact ctcagtacaa
     2041 tctgctctga tgccgcatag ttaagccagt atacactccg ctatcgctac gtgactgggt
     2101 catggctgcg ccccgacacc cgccaacacc cgctgacgcg ccctgacggg cttgtctgct
     2161 cccggcatcc gcttacagac aagctgtgac cgtctccggg agctgcatgt gtcagaggtt
     2221 ttcaccgtca tcaccgaaac gcgcgaggca gctgcggtaa agctcatcag cgtggtcgtg
     2281 aagcgattca cagatgtctg cctgttcatc cgcgtccagc tcgttgagtt tctccagaag
     2341 cgttaatgtc tggcttctga taaagcgggc catgttaagg gcggtttttt cctgtttggt
     2401 cactgatgcc tccgtgtaag ggggatttct gttcatgggg gtaatgatac cgatgaaacg
     2461 agagaggatg ctcacgatac gggttactga tgatgaacat gcccggttac tggaacgttg
     2521 tgagggtaaa caactggcgg tatggatgcg gcgggaccag agaaaaatca ctcagggtca
     2581 atgccagcgc ttcgttaata cagatgtagg tgttccacag ggtagccagc agcatcctgc
     2641 gatgcagatc cggaacataa tggtgcaggg cgctgacttc cgcgtttcca gactttacga
     2701 aacacggaaa ccgaagacca ttcatgttgt tgctcaggtc gcagacgttt tgcagcagca
     2761 gtcgcttcac gttcgctcgc gtatcggtga ttcattctgc taaccagtaa ggcaaccccg
     2821 ccagcctagc cgggtcctca acgacaggag cacgatcatg cgcacccgtg gggccgccat
     2881 gccggcgata atggcctgct tctcgccgaa acgtttggtg gcgggaccag tgacgaaggc
     2941 ttgagcgagg gcgtgcaaga ttccgaatac cgcaagcgac aggccgatca tcgtcgcgct
     3001 ccagcgaaag cggtcctcgc cgaaaatgac ccagagcgct gccggcacct gtcctacgag
     3061 ttgcatgata aagaagacag tcataagtgc ggcgacgata gtcatgcccc gcgcccaccg
     3121 gaaggagctg actgggttga aggctctcaa gggcatcggt cgagatcccg gtgcctaatg
     3181 agtgagctaa cttacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     3241 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     3301 gcgccagggt ggtttttctt ttcaccagtg agacgggcaa cagctgattg cccttcaccg
     3361 cctggccctg agagagttgc agcaagcggt ccacgctggt ttgccccagc aggcgaaaat
     3421 cctgtttgat ggtggttaac ggcgggatat aacatgagct gtcttcggta tcgtcgtatc
     3481 ccactaccga gatatccgca ccaacgcgca gcccggactc ggtaatggcg cgcattgcgc
     3541 ccagcgccat ctgatcgttg gcaaccagca tcgcagtggg aacgatgccc tcattcagca
     3601 tttgcatggt ttgttgaaaa ccggacatgg cactccagtc gccttcccgt tccgctatcg
     3661 gctgaatttg attgcgagtg agatatttat gccagccagc cagacgcaga cgcgccgaga
     3721 cagaacttaa tgggcccgct aacagcgcga tttgctggtg acccaatgcg accagatgct
     3781 ccacgcccag tcgcgtaccg tcttcatggg agaaaataat actgttgatg ggtgtctggt
     3841 cagagacatc aagaaataac gccggaacat tagtgcaggc agcttccaca gcaatggcat
     3901 cctggtcatc cagcggatag ttaatgatca gcccactgac gcgttgcgcg agaagattgt
     3961 gcaccgccgc tttacaggct tcgacgccgc ttcgttctac catcgacacc accacgctgg
     4021 cacccagttg atcggcgcga gatttaatcg ccgcgacaat ttgcgacggc gcgtgcaggg
     4081 ccagactgga ggtggcaacg ccaatcagca acgactgttt gcccgccagt tgttgtgcca
     4141 cgcggttggg aatgtaattc agctccgcca tcgccgcttc cactttttcc cgcgttttcg
     4201 cagaaacgtg gctggcctgg ttcaccacgc gggaaacggt ctgataagag acaccggcat
     4261 actctgcgac atcgtataac gttactggtt tcacattcac caccctgaat tgactctctt
     4321 ccgggcgcta tcatgccata ccgcgaaagg ttttgcgcca ttcgatggtg tccgggatct
     4381 cgacgctctc ccttatgcga ctcctgcatt aggaagcagc ccagtagtag gttgaggccg
     4441 ttgagcaccg ccgccgcaag gaatggtgca tgcaaggaga tggcgcccaa cagtcccccg
     4501 gccacggggc ctgccaccat acccacgccg aaacaagcgc tcatgagccc gaagtggcga
     4561 gcccgatctt ccccatcggt gatgtcggcg atataggcgc cagcaaccgc acctgtggcg
     4621 ccggtgatgc cggccacgat gcgtccggcg tagaggatcg agatctcgat cccgcgaaat
     4681 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
     4741 tgtttaactt taagaaggag atatacatat gcaaccggtc agccaagcaa atggacgaat
     4801 ccacttggat cctgatcaag ctgaacaagg cttagcgcag cttgtgatga cagttattga
     4861 gctattgagg caaatcgttg aacgtcatgc catgaggcgg gtggagggtg gaacgttgac
     4921 ggacgaacaa attgaaaact taggaattgc actaatgaac ttagaagaaa aaatggacga
     4981 gttgaaagag gtgttcggtc tggatgcaga agatttaaat attgatcttg gaccgctagg
     5041 cagcctgctt gtttctcaag gcagttcagg tggtggcggg agcggaggtg gaggctcgag
     5101 cggtgtgacc ggctaccggc tgttcgagga gattctgtga ggctgttttg gcggatgaga
     5161 gaagattgag atccggctgc taacaaagcc cgaaaggaag ctgagttggc tgctgccacc
     5221 gctgagcaat aactagcata accccttggg gcctctaaac gggtcttgag gggttttttg
     5281 ctgaaaggag gaactatatc cggattggcg aatgggacgc gccctgtagc ggcgcattaa
     5341 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc
     5401 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
     5461 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     5521 aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag acg
//
