LOCUS       sequence-193330-        6218 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses C-Terminal segment of C. elegans Piezo1 in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6218)
  AUTHORS   Kamajaya A, Kaiser JT, Lee J, Reid M, Rees DC
  TITLE     The structure of a conserved piezo channel domain reveals a 
            topologically distinct beta sandwich fold.
  JOURNAL   Structure. 2014 Oct 7;22(10):1520-7. doi: 10.1016/j.str.2014.08.009.
            Epub 2014 Sep 18.
  PUBMED    25242456
REFERENCE   2  (bases 1 to 6218)
  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: 
            "10.1016/j.str.2014.08.009"; journalName: "Structure"; date: 
            "2014-10-7- 7"; volume: "22"; issue: "10"; pages: "1520-7"
COMMENT     Sequence Label: CTL2
FEATURES             Location/Qualifiers
     source          1..6218
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      15..470
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(102..121)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     312..333
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        496..600
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(601..669,670..1461)
                     /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(819..838)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1632..2220
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2121..2140
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2374..2391
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2406..2545
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2471..2490)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2631..2653
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2647..2838)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(3647..4729)
                     /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     4691..4710
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4730..4807)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5013..5032
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     5116..5135
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5116..5134
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5135..5159
                     /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             5174..5196
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             6065..6082
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(6135..6153)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      6149..6196
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
ORIGIN
        1 gattggcgaa tgggacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
       61 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
      121 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
      181 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
      241 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
      301 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta
      361 ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat
      421 ttaacaaaaa tttaacgcga attttaacaa aatattaacg cttacaattt aggtggcact
      481 tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg
      541 tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt
      601 atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct
      661 gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca
      721 cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc
      781 gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc
      841 cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg
      901 gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta
      961 tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc
     1021 ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt
     1081 gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg
     1141 cctgcagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct
     1201 tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc
     1261 tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct
     1321 cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac
     1381 acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc
     1441 tcactgatta agcattggta actgtcagac caagtttact catatatact ttagattgat
     1501 ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg
     1561 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     1621 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     1681 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     1741 gtaactggct tcagcagagc gcagatacca aatactgtcc ttctagtgta gccgtagtta
     1801 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     1861 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     1921 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     1981 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     2041 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     2101 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     2161 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     2221 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     2281 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
     2341 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
     2401 gagcgcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcaatggt
     2461 gcactctcag tacaatctgc tctgatgccg catagttaag ccagtataca ctccgctatc
     2521 gctacgtgac tgggtcatgg ctgcgccccg acacccgcca acacccgctg acgcgccctg
     2581 acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg
     2641 catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg aggcagctgc ggtaaagctc
     2701 atcagcgtgg tcgtgaagcg attcacagat gtctgcctgt tcatccgcgt ccagctcgtt
     2761 gagtttctcc agaagcgtta atgtctggct tctgataaag cgggccatgt taagggcggt
     2821 tttttcctgt ttggtcactg atgcctccgt gtaaggggga tttctgttca tgggggtaat
     2881 gataccgatg aaacgagaga ggatgctcac gatacgggtt actgatgatg aacatgcccg
     2941 gttactggaa cgttgtgagg gtaaacaact ggcggtatgg atgcggcggg accagagaaa
     3001 aatcactcag ggtcaatgcc agcgcttcgt taatacagat gtaggtgttc cacagggtag
     3061 ccagcagcat cctgcgatgc agatccggaa cataatggtg cagggcgctg acttccgcgt
     3121 ttccagactt tacgaaacac ggaaaccgaa gaccattcat gttgttgctc aggtcgcaga
     3181 cgttttgcag cagcagtcgc ttcacgttcg ctcgcgtatc ggtgattcat tctgctaacc
     3241 agtaaggcaa ccccgccagc ctagccgggt cctcaacgac aggagcacga tcatgcgcac
     3301 ccgtggggcc gccatgccgg cgataatggc ctgcttctcg ccgaaacgtt tggtggcggg
     3361 accagtgacg aaggcttgag cgagggcgtg caagattccg aataccgcaa gcgacaggcc
     3421 gatcatcgtc gcgctccagc gaaagcggtc ctcgccgaaa atgacccaga gcgctgccgg
     3481 cacctgtcct acgagttgca tgataaagaa gacagtcata agtgcggcga cgatagtcat
     3541 gccccgcgcc caccggaagg agctgactgg gttgaaggct ctcaagggca tcggtcgaga
     3601 tcccggtgcc taatgagtga gctaacttac attaattgcg ttgcgctcac tgcccgcttt
     3661 ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg
     3721 cggtttgcgt attgggcgcc agggtggttt ttcttttcac cagtgagacg ggcaacagct
     3781 gattgccctt caccgcctgg ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc
     3841 ccagcaggcg aaaatcctgt ttgatggtgg ttaacggcgg gatataacat gagctgtctt
     3901 cggtatcgtc gtatcccact accgagatat ccgcaccaac gcgcagcccg gactcggtaa
     3961 tggcgcgcat tgcgcccagc gccatctgat cgttggcaac cagcatcgca gtgggaacga
     4021 tgccctcatt cagcatttgc atggtttgtt gaaaaccgga catggcactc cagtcgcctt
     4081 cccgttccgc tatcggctga atttgattgc gagtgagata tttatgccag ccagccagac
     4141 gcagacgcgc cgagacagaa cttaatgggc ccgctaacag cgcgatttgc tggtgaccca
     4201 atgcgaccag atgctccacg cccagtcgcg taccgtcttc atgggagaaa ataatactgt
     4261 tgatgggtgt ctggtcagag acatcaagaa ataacgccgg aacattagtg caggcagctt
     4321 ccacagcaat ggcatcctgg tcatccagcg gatagttaat gatcagccca ctgacgcgtt
     4381 gcgcgagaag attgtgcacc gccgctttac aggcttcgac gccgcttcgt tctaccatcg
     4441 acaccaccac gctggcaccc agttgatcgg cgcgagattt aatcgccgcg acaatttgcg
     4501 acggcgcgtg cagggccaga ctggaggtgg caacgccaat cagcaacgac tgtttgcccg
     4561 ccagttgttg tgccacgcgg ttgggaatgt aattcagctc cgccatcgcc gcttccactt
     4621 tttcccgcgt tttcgcagaa acgtggctgg cctggttcac cacgcgggaa acggtctgat
     4681 aagagacacc ggcatactct gcgacatcgt ataacgttac tggtttcaca ttcaccaccc
     4741 tgaattgact ctcttccggg cgctatcatg ccataccgcg aaaggttttg cgccattcga
     4801 tggtgtccgg gatctcgacg ctctccctta tgcgactcct gcattaggaa gcagcccagt
     4861 agtaggttga ggccgttgag caccgccgcc gcaaggaatg gtgcatgcaa ggagatggcg
     4921 cccaacagtc ccccggccac ggggcctgcc accataccca cgccgaaaca agcgctcatg
     4981 agcccgaagt ggcgagcccg atcttcccca tcggtgatgt cggcgatata ggcgccagca
     5041 accgcacctg tggcgccggt gatgccggcc acgatgcgtc cggcgtagag gatcgagatc
     5101 tcgatcccgc gaaattaata cgactcacta taggggaatt gtgagcggat aacaattccc
     5161 ctctagaaat aattttgttt aactttaaga aggagatata caaggaggta tgtcgctact
     5221 caatcaaatc ggaactatca gtatgccaga aaaagttacg ttgaggattt cgattgaagg
     5281 atatccacca ctttacgaga tggaggcaca aggaagtaat cacgataacg ctgaacttgg
     5341 aatgataaaa cctgatcaat tggcatcttt gaatcaagca ttgacggata gttacacgac
     5401 acgagacact aattctattt taagatcaag aatgtctgta tcatatctga aaggatacac
     5461 atatgaggat attctgattg tgcgattccg tccagaatct gaaatttatt ggccgatttc
     5521 acaggattcg agaaatgcga tgatcgataa attgagccga aacacgtcag tcaattttga
     5581 agtatctctg gaattcactc ggccatatga tccaaatgaa aacgctgctt taaaacattc
     5641 gaaatcgtgg cttgttccaa tctccctgga tatgacgatt cgagcaaaaa ttcaaagtgc
     5701 tctccgagga gatccaggtc atccgattct cattccacag tcaattcctg cgtttattca
     5761 agttccaaat caaggagaat tgacccttcc aacatcaatc ggaaatacta ttatcaatga
     5821 cggaaatcca cggattaata cgaccgggat ggaaaaatca gacgaagctc gggcttggtt
     5881 cgactcgtta accttgaatc tcgaacaagg aaaatcgcaa aacgagaaaa tgtggatcgc
     5941 cacatccgaa catccaggag accaaaatgc gaaactctgg ataaagactg caaatacgac
     6001 gtattcggga aggccgtacc ttcaagttgt cggatttatt gatagagcat tcccatcact
     6061 cgagcaccac caccaccacc actgagatcc ggctgctaac aaagcccgaa aggaagctga
     6121 gttggctgct gccaccgctg agcaataact agcataaccc cttggggcct ctaaacgggt
     6181 cttgaggggt tttttgctga aaggaggaac tatatccg
//
