LOCUS       sequence-112155-        6596 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Dual expression of GBD-LUMP and cdc42.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6596)
  AUTHORS   Hoepker AC, Wang A, Le Marois A, Suhling K, Yan Y, Marriott G
  TITLE     Genetically encoded sensors of protein hydrodynamics and molecular 
            proximity.
  JOURNAL   Proc Natl Acad Sci U S A. 2015 Apr 30. pii: 201424021.
  PUBMED    25931526
REFERENCE   2  (bases 1 to 6596)
  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: "Proc Natl 
            Acad Sci U S A. 2015 Apr 30. pii: 201424021."
COMMENT     Sequence Label: 213-petDuet1-GBD-LUMP--cdc42
FEATURES             Location/Qualifiers
     source          1..6596
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    3..27
                     /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             42..64
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     promoter        859..877
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    878..902
                     /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)."
     CDS             945..1520
                     /codon_start=1
                     /product="human Rho family small GTPase Cdc42"
                     /label=Cdc42
                     /translation="MQTIKCVVVGDGAVGKTCLLISYTTNKFPSEYVPTVFDNYAVTVM
                     IGGEPYTLGLFDTAGQEDYDRLRPLSYPQTDVFLVCFSVVSPSSFENVKEKWVPEITHH
                     CPKTPFLLVGTQIDLRDDPSTIEKLAKNKQKPITPETAEKLARDLKAVKYVECSALTQK
                     GLKNVFDEAILAALEPPEPKKSRRCVLL"
     CDS             1542..1586
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(1624..1642)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1638..1685
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1722..2177
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1809..1828)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2019..2040
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(join(2292..3083,3084..3152))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     2915..2934
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3153..3245)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3326..3914
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3815..3834
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4068..4085
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4100..4242
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4168..4187)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4328..4350
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4344..4535)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5107..6189)
                     /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     6151..6170
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6190..6267)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6473..6492
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        join(6580..6596,1..2)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ggggaattgt gagcggataa caattcccct ctagaaataa ttttgtttaa ctttaagaag
       61 gagatatacc atgggcctga gcgcacagga catcagccag ccgctgcaga acagcttcat
      121 ccataccggc cacggcgaca gcgatcctcg ccattgctgg ggctttccgg accgcatcga
      181 cgagctgtac ctgggcaacg gcagcggcag cgctagcttt agaggtattg ttcaaggtcg
      241 tggtgttatt cgatccattt caaaaagtga agatagtcag cgacatggga ttgcattccc
      301 tgagggtatg tttcaacttg ttgatgtaga tactgtaatg ttagtaaatg gttgttcatt
      361 gacagttgtc cgtattttag gcgatatggt gtactttgat atcgaccaag cattgggtac
      421 gacaacattt gatggattaa aagaaggcga tcaggtaaac ctagagatcc accctaaatt
      481 tggtgaagtg gttggccgtg gtggattaac aggaaatatt aaaggcactg cgctggtcgc
      541 agcaatagaa gagaatgacg caggtttttc tgtactgatt gatatcccta aagggttagc
      601 tgaaaactta acagtgaaag atgatattgg aattgatggg atctctttgc ctatcactga
      661 tatgtcagac agcattatta cattgaatta ctcgcgtgat ttgttagcga gtactaacat
      721 tgcatcgtta gcaaaagatg tgaaagtgaa cgttgaaatt ttaaatgaat ggtaggcggc
      781 cgcactcaag cttgcggccg cataatgctt aagtcgaaca gaaagtaatc gtattgtaca
      841 cggccgcata atcgaaatta atacgactca ctatagggga attgtgagcg gataacaatt
      901 ccccatctta gtatattagt taagtataag aaggagatat acatatgcag acaattaagt
      961 gtgttgttgt gggcgatggt gctgttggta aaacatgtct cctgatatcc tacacaacaa
     1021 acaaatttcc atcggaatat gtaccgactg tttttgacaa ctatgcagtc acagttatga
     1081 ttggtggaga accatatact cttggacttt ttgatactgc agggcaagag gattatgata
     1141 gattacgacc gctgagttat ccacaaacag atgtatttct agtctgtttt tcagtggtct
     1201 ctccatcttc atttgaaaac gtgaaagaaa agtgggtgcc tgagataact caccactgtc
     1261 caaagactcc tttcttgctt gttgggactc aaattgatct cagagatgac ccctctacta
     1321 ttgagaaact tgccaagaac aaacagaagc ctatcactcc agagactgct gaaaagctgg
     1381 cccgtgacct gaaggctgtc aagtatgtgg agtgttctgc acttacacag aaaggcctaa
     1441 agaatgtatt tgacgaagca atattggctg ccctggagcc tccagaaccg aagaagagcc
     1501 gcaggtgtgt gctgctatga gcggccgcac tcgagtctgg taaagaaacc gctgctgcga
     1561 aatttgaacg ccagcacatg gactcgtcta ctagcgcagc ttaattaacc taggctgctg
     1621 ccaccgctga gcaataacta gcataacccc ttggggcctc taaacgggtc ttgaggggtt
     1681 ttttgctgaa aggaggaact atatccggat tggcgaatgg gacgcgccct gtagcggcgc
     1741 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     1801 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     1861 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga
     1921 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     1981 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
     2041 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc
     2101 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat
     2161 attaacgttt acaatttctg gcggcacgat ggcatgagat tatcaaaaag gatcttcacc
     2221 tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact
     2281 tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt
     2341 cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta
     2401 ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta
     2461 tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc
     2521 gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat
     2581 agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt
     2641 atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg
     2701 tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca
     2761 gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta
     2821 agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg
     2881 cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact
     2941 ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg
     3001 ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt
     3061 actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga
     3121 ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaatc atgattgaag
     3181 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     3241 acaaataggt catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
     3301 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
     3361 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
     3421 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag
     3481 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
     3541 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
     3601 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
     3661 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
     3721 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
     3781 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
     3841 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
     3901 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
     3961 cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg
     4021 cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga
     4081 gcgaggaagc ggaagagcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt
     4141 cacaccgcat atatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca
     4201 gtatacactc cgctatcgct acgtgactgg gtcatggctg cgccccgaca cccgccaaca
     4261 cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg
     4321 accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgagg
     4381 cagctgcggt aaagctcatc agcgtggtcg tgaagcgatt cacagatgtc tgcctgttca
     4441 tccgcgtcca gctcgttgag tttctccaga agcgttaatg tctggcttct gataaagcgg
     4501 gccatgttaa gggcggtttt ttcctgtttg gtcactgatg cctccgtgta agggggattt
     4561 ctgttcatgg gggtaatgat accgatgaaa cgagagagga tgctcacgat acgggttact
     4621 gatgatgaac atgcccggtt actggaacgt tgtgagggta aacaactggc ggtatggatg
     4681 cggcgggacc agagaaaaat cactcagggt caatgccagc gcttcgttaa tacagatgta
     4741 ggtgttccac agggtagcca gcagcatcct gcgatgcaga tccggaacat aatggtgcag
     4801 ggcgctgact tccgcgtttc cagactttac gaaacacgga aaccgaagac cattcatgtt
     4861 gttgctcagg tcgcagacgt tttgcagcag cagtcgcttc acgttcgctc gcgtatcggt
     4921 gattcattct gctaaccagt aaggcaaccc cgccagccta gccgggtcct caacgacagg
     4981 agcacgatca tgctagtcat gccccgcgcc caccggaagg agctgactgg gttgaaggct
     5041 ctcaagggca tcggtcgaga tcccggtgcc taatgagtga gctaacttac attaattgcg
     5101 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     5161 ggccaacgcg cggggagagg cggtttgcgt attgggcgcc agggtggttt ttcttttcac
     5221 cagtgagacg ggcaacagct gattgccctt caccgcctgg ccctgagaga gttgcagcaa
     5281 gcggtccacg ctggtttgcc ccagcaggcg aaaatcctgt ttgatggtgg ttaacggcgg
     5341 gatataacat gagctgtctt cggtatcgtc gtatcccact accgagatgt ccgcaccaac
     5401 gcgcagcccg gactcggtaa tggcgcgcat tgcgcccagc gccatctgat cgttggcaac
     5461 cagcatcgca gtgggaacga tgccctcatt cagcatttgc atggtttgtt gaaaaccgga
     5521 catggcactc cagtcgcctt cccgttccgc tatcggctga atttgattgc gagtgagata
     5581 tttatgccag ccagccagac gcagacgcgc cgagacagaa cttaatgggc ccgctaacag
     5641 cgcgatttgc tggtgaccca atgcgaccag atgctccacg cccagtcgcg taccgtcttc
     5701 atgggagaaa ataatactgt tgatgggtgt ctggtcagag acatcaagaa ataacgccgg
     5761 aacattagtg caggcagctt ccacagcaat ggcatcctgg tcatccagcg gatagttaat
     5821 gatcagccca ctgacgcgtt gcgcgagaag attgtgcacc gccgctttac aggcttcgac
     5881 gccgcttcgt tctaccatcg acaccaccac gctggcaccc agttgatcgg cgcgagattt
     5941 aatcgccgcg acaatttgcg acggcgcgtg cagggccaga ctggaggtgg caacgccaat
     6001 cagcaacgac tgtttgcccg ccagttgttg tgccacgcgg ttgggaatgt aattcagctc
     6061 cgccatcgcc gcttccactt tttcccgcgt tttcgcagaa acgtggctgg cctggttcac
     6121 cacgcgggaa acggtctgat aagagacacc ggcatactct gcgacatcgt ataacgttac
     6181 tggtttcaca ttcaccaccc tgaattgact ctcttccggg cgctatcatg ccataccgcg
     6241 aaaggttttg cgccattcga tggtgtccgg gatctcgacg ctctccctta tgcgactcct
     6301 gcattaggaa gcagcccagt agtaggttga ggccgttgag caccgccgcc gcaaggaatg
     6361 gtgcatgcaa ggagatggcg cccaacagtc ccccggccac ggggcctgcc accataccca
     6421 cgccgaaaca agcgctcatg agcccgaagt ggcgagcccg atcttcccca tcggtgatgt
     6481 cggcgatata ggcgccagca accgcacctg tggcgccggt gatgccggcc acgatgcgtc
     6541 cggcgtagag gatcgagatc gatctcgatc ccgcgaaatt aatacgactc actata
//
