LOCUS       Exported                7235 bp ds-DNA     circular SYN 30-JUN-2022
DEFINITION  For expresseion of recombinant paxillin in bacteria.
ACCESSION   .
VERSION     .
KEYWORDS    SNAP-Paxillin
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7235)
  AUTHORS   Case LB, De Pasquale M, Henry L, Rosen MK
  TITLE     Synergistic phase separation of two pathways promotes integrin 
            clustering and nascent adhesion formation.
  JOURNAL   Elife. 2022 Jan 20;11. pii: 72588. doi: 10.7554/eLife.72588.
  PUBMED    35049497
REFERENCE   2  (bases 1 to 7235)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jun 30, 2022 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7235
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     154..171
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        244..321
                     /gene="lacI"
                     /label=lacI promoter
     CDS             322..1404
                     /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     complement(341..360)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        1453..1483
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1491..1507
                     /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)."
     primer_bind     1515..1531
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1515..1531
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             1527..2054
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(1547..1564)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1547..1563)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1556..1578)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     promoter        2156..2184
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    2192..2208
                     /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             2231..2884
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2842..2864
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2891..2908
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             2909..2929
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             2933..3478
                     /codon_start=1
                     /product="human O6-alkylguanine-DNA-alkyltransferase 
                     variant that forms covalent bonds with benzylguanine 
                     derivatives"
                     /label=SNAP-tag(R)
                     /note="optimized for bacterial expression"
                     /translation="MDKDCEMKRTTLDSPLGKLELSGCEQGLHEIKLLGKGTSAADAVE
                     VPAPAAVLGGPEPLMQATAWLNAYFHQPEAIEEFPVPALHHPVFQQESFTRQVLWKLLK
                     VVKFGEVISYQQLAALAGNPAATAAVKTALSGNPVPILIPCHRVVSSSGAVGGYEGGLA
                     VKEWLLAHEGHRLGKPGLG"
     primer_bind     complement(5258..5280)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5425..5443)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5511..5615
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5616..6476
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5834..5853)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6647..7235
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7136..7155
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
       61 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
      121 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
      181 gagcgcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcataaat
      241 tccgacacca tcgaatggcg caaaaccttt cgcggtatgg catgatagcg cccggaagag
      301 agtcaattca gggtggtgaa tgtgaaacca gtaacgttat acgatgtcgc agagtatgcc
      361 ggtgtctctt atcagaccgt ttcccgcgtg gtgaaccagg ccagccacgt ttctgcgaaa
      421 acgcgggaaa aagtggaagc ggcgatggcg gagctgaatt acattcccaa ccgcgtggca
      481 caacaactgg cgggcaaaca gtcgttgctg attggcgttg ccacctccag tctggccctg
      541 cacgcgccgt cgcaaattgt cgcggcgatt aaatctcgcg ccgatcaact gggtgccagc
      601 gtggtggtgt cgatggtaga acgaagcggc gtcgaagcct gtaaagcggc ggtgcacaat
      661 cttctcgcgc aacgcgtcag tgggctgatc attaactatc cgctggatga ccaggatgcc
      721 attgctgtgg aagctgcctg cactaatgtt ccggcgttat ttcttgatgt ctctgaccag
      781 acacccatca acagtattat tttctcccat gaagacggta cgcgactggg cgtggagcat
      841 ctggtcgcat tgggtcacca gcaaatcgcg ctgttagcgg gcccattaag ttctgtctcg
      901 gcgcgtctgc gtctggctgg ctggcataaa tatctcactc gcaatcaaat tcagccgata
      961 gcggaacggg aaggcgactg gagtgccatg tccggttttc aacaaaccat gcaaatgctg
     1021 aatgagggca tcgttcccac tgcgatgctg gttgccaacg atcagatggc gctgggcgca
     1081 atgcgcgcca ttaccgagtc cgggctgcgc gttggtgcgg atatctcggt agtgggatac
     1141 gacgataccg aagacagctc atgttatatc ccgccgttaa ccaccatcaa acaggatttt
     1201 cgcctgctgg ggcaaaccag cgtggaccgc ttgctgcaac tctctcaggg ccaggcggtg
     1261 aagggcaatc agctgttgcc cgtctcactg gtgaaaagaa aaaccaccct ggcgcccaat
     1321 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
     1381 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtaagttagc tcactcatta
     1441 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
     1501 ataacaattt cacacaggaa acagctatga ccatgattac ggattcactg gccgtcgttt
     1561 tacaacgtcg tgactgggaa aaccctggcg ttacccaact taatcgcctt gcagcacatc
     1621 cccctttcgc cagctggcgt aatagcgaag aggcccgcac cgatcgccct tcccaacagt
     1681 tgcgcagcct gaatggcgaa tggcgctttg cctggtttcc ggcaccagaa gcggtgccgg
     1741 aaagctggct ggagtgcgat cttcctgagg ccgatactgt cgtcgtcccc tcaaactggc
     1801 agatgcacgg ttacgatgcg cccatctaca ccaacgtgac ctatcccatt acggtcaatc
     1861 cgccgtttgt tcccacggag aatccgacgg gttgttactc gctcacattt aatgttgatg
     1921 aaagctggct acaggaaggc cagacgcgaa ttatttttga tggcgttgga attagcttat
     1981 cgactgcacg gtgcaccaat gcttctggcg tcaggcagcc atcggaagct gtggtatggc
     2041 tgtgcaggtc gtaaatcact gcataattcg tgtcgctcaa ggcgcactcc cgttctggat
     2101 aatgtttttt gcgccgacat cataacggtt ctggcaaata ttctgaaatg agctgttgac
     2161 aattaatcat cggctcgtat aatgtgtgga attgtgagcg gataacaatt tcacacagga
     2221 aacagtattc atgtccccta tactaggtta ttggaaaatt aagggccttg tgcaacccac
     2281 tcgacttctt ttggaatatc ttgaagaaaa atatgaagag catttgtatg agcgcgatga
     2341 aggtgataaa tggcgaaaca aaaagtttga attgggtttg gagtttccca atcttcctta
     2401 ttatattgat ggtgatgtta aattaacaca gtctatggcc atcatacgtt atatagctga
     2461 caagcacaac atgttgggtg gttgtccaaa agagcgtgca gagatttcaa tgcttgaagg
     2521 agcggttttg gatattagat acggtgtttc gagaattgca tatagtaaag actttgaaac
     2581 tctcaaagtt gattttctta gcaagctacc tgaaatgctg aaaatgttcg aagatcgttt
     2641 atgtcataaa acatatttaa atggtgatca tgtaacccat cctgacttca tgttgtatga
     2701 cgctcttgat gttgttttat acatggaccc aatgtgcctg gatgcgttcc caaaattagt
     2761 ttgttttaaa aaacgtattg aagctatccc acaaattgat aagtacttga aatccagcaa
     2821 gtatatagca tggcctttgc agggctggca agccacgttt ggtggtggcg accatcctcc
     2881 aaaatcggat ctggttccgc gtggatctga aaacctgtat tttcagggcc atatggacaa
     2941 agattgcgaa atgaaacgta ccaccctgga tagcccgctg ggcaaactgg aactgagcgg
     3001 ctgcgaacag ggcctgcatg aaattaaact gctgggtaaa ggcaccagcg cggccgatgc
     3061 ggttgaagtt ccggccccgg ccgccgtgct gggtggtccg gaaccgctga tgcaggcgac
     3121 cgcgtggctg aacgcgtatt ttcaccagcc ggaagcgatt gaagaatttc cggttccggc
     3181 gctgcatcat ccggtgtttc agcaggagag ctttacccgt caggtgctgt ggaaactgct
     3241 gaaagtggtt aaatttggcg aagtgattag ctatcagcag ctggcggccc tggcgggtaa
     3301 tccggcggcc accgccgccg ttaaaaccgc gctgagcggt aacccggtgc cgattctgat
     3361 tccgtgccat cgtgtggtta gctctagcgg tgcggttggc ggttatgaag gtggtctggc
     3421 ggtgaaagag tggctgctgg cccatgaagg tcatcgtctg ggtaaaccgg gtctgggagg
     3481 tagcggtagc ggtagcggtg gaggtggaag cagccatatg gacgacctcg acgccctgct
     3541 ggcggacttg gagtctacca cctcccacat ctccaaacgg cctgtgttct tgtcggagga
     3601 gaccccctac tcatacccaa ctggaaacca cacataccag gagattgccg tgccaccccc
     3661 cgtcccccca cccccgtcca gcgaggccct caatggcaca atccttgacc ccttagacca
     3721 gtggcagccc agcggctccc gattcatcca ccagcagcct cagtcctcat cacctgtgta
     3781 cggctccagt gccaaaactt ccagtgtctc caaccctcag gacagtgttg gctctccgtg
     3841 ctcccgagtg ggtgaggagg agcacgtcta cagcttcccc aacaagcaga aatcagctga
     3901 gccttcaccc accgtaatga gcacgtccct gggcagcaac ctttctgaac tcgaccgcct
     3961 gctgctggaa ctgaacgctg tacagcataa cccgccaggc ttccctgcag atgaggccaa
     4021 ctcaagcccc ccgcttcctg gggccctgag ccccctctat ggtgtcccag agactaacag
     4081 ccccttggga ggcaaagctg ggcccctgac gaaagagaag cctaagcgga atgggggccg
     4141 gggcctggag gacgtgcggc ccagtgtgga gagtctcttg gatgaactgg agagctccgt
     4201 gcccagcccc gtccctgcca tcactgtgaa ccagggcgag atgagcagcc cgcagcgcgt
     4261 cacctccacc caacagcaga cacgcatctc ggcctcctct gccaccaggg agctggacga
     4321 gctgatggct tcgctgtcgg atttcaagtt catggcccag gggaagacag ggagcagctc
     4381 accccctggg gggcccccga agcccgggag ccagctggac agcatgctgg ggagcctgca
     4441 gtctgacctg aacaagctgg gggtcgccac agtcgccaaa ggagtctgcg gggcctgcaa
     4501 gaagcccatc gccgggcagg ttgtgaccgc catggggaag acgtggcacc ccgagcactt
     4561 cgtctgcacc cactgccagg aggagatcgg ttcacggaac ttcttcgagc gggatggaca
     4621 gccctactgt gaaaaggact accacaacct cttctccccg cgctgctact actgcaacgg
     4681 ccccatcctg gataaagtgg tgacagccct tgaccggacg tggcaccctg aacacttctt
     4741 ctgtgcacag tgtggagcct tctttggtcc cgaagggttc cacgagaagg acggcaaggc
     4801 ctactgtcgc aaggactact tcgacatgtt cgcacccaag tgtggcggct gcgcccgggc
     4861 catcctggag aactatatct cagccctcaa cacgctgtgg catcctgagt gctttgtgtg
     4921 ccgggaatgc ttcacgccat tcgtgaacgg cagcttcttc gagcacgacg ggcagcccta
     4981 ctgtgaggtg cactaccacg agcggcgcgg ctcgctgtgt tctggctgcc agaagcccat
     5041 caccggccgc tgcatcaccg ccatggccaa gaagttccac cccgagcact tcgtctgtgc
     5101 cttctgcctc aagcagctca acaagggcac cttcaaggag cagaacgaca agccttactg
     5161 tcagaactgc ttcctcaagc tcttctgcta gggatccgcg gccgcatcgt gactgactga
     5221 cgatctgcct cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg
     5281 agacggtcac agcttgtctg taagcggatg ccgggagcag acaagcccgt cagggcgcgt
     5341 cagcgggtgt tggcgggtgt cggggcgcag ccatgaccca gtcacgtagc gatagcggag
     5401 tgtataattc ttgaagacga aagggcctcg tgatacgcct atttttatag gttaatgtca
     5461 tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc
     5521 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     5581 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
     5641 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     5701 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
     5761 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
     5821 cttttaaagt tctgctatgt ggcgcggtat tatcccgtgt tgacgccggg caagagcaac
     5881 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
     5941 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     6001 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
     6061 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     6121 aagccatacc aaacgacgag cgtgacacca cgatgcctgc agcaatggca acaacgttgc
     6181 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
     6241 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     6301 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     6361 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     6421 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
     6481 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     6541 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
     6601 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     6661 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     6721 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     6781 taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     6841 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     6901 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     6961 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     7021 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     7081 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     7141 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     7201 tgtgatgctc gtcagggggg cggagcctat ggaaa
//
