LOCUS       sequence-146556-        5436 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Construct for expressing Grb2 lacking the C-terminal SH3 domain in 
            E. Coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5436)
  AUTHORS   Su X, Ditlev JA, Hui E, Xing W, Banjade S, Okrut J, King DS, Taunton
            J, Rosen MK, Vale RD
  TITLE     Phase separation of signaling molecules promotes T cell receptor 
            signal transduction.
  JOURNAL   Science. 2016 Apr 29;352(6285):595-9. doi: 10.1126/science.aad9964. 
            Epub 2016 Apr 7.
  PUBMED    27056844
REFERENCE   2  (bases 1 to 5436)
  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: 
            "10.1126/science.aad9964"; journalName: "Science"; date: 
            "2016-04-29- 29"; volume: "352"; issue: "6285"; pages: "595-9"
COMMENT     Sequence Label: GST-PS-Grb2 1-154 (XSB382)
FEATURES             Location/Qualifiers
     source          1..5436
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(183..188,189..204,205..211)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    219..235
                     /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             258..911
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     869..891
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             918..941
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(1486..1508)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1653..1671)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1739..1843
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1844..1912,1913..2704)
                     /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(2062..2081)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2875..3463
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3364..3383
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3617..3634
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3707..3784
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             3785..4867
                     /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
                     YIPPSTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(3804..3823)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     protein_bind    4880..4901
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4916..4921,4922..4939,4940..4946)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4954..4970
                     /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     4978..4994
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4978..4994
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(4990..5436,1..81)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGITLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5010..5027)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5010..5026)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5019..5041)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 acgttatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc ggaagctgtg
       61 gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc gcactcccgt
      121 tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc tgaaatgagc
      181 tgttgacaat taatcatcgg ctcgtataat gtgtggaatt gtgagcggat aacaatttca
      241 cacaggaaac agtattcatg tcccctatac taggttattg gaaaattaag ggccttgtgc
      301 aacccactcg acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc
      361 gcgatgaagg tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc
      421 ttccttatta tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata
      481 tagctgacaa gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc
      541 ttgaaggagc ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact
      601 ttgaaactct caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag
      661 atcgtttatg tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt
      721 tgtatgacgc tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa
      781 aattagtttg ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat
      841 ccagcaagta tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc
      901 atcctccaaa atcggatctg gaagttctgt tccaggggcc cctgggatcc atggaagcca
      961 tcgccaaata tgacttcaaa gctactgcag acgacgagct gagcttcaaa aggggggaca
     1021 tcctcaaggt tttgaacgaa gaatgtgatc agaactggta caaggcagag cttaatggaa
     1081 aagacggctt cattcccaag aactacatag aaatgaaacc acatccgtgg ttttttggca
     1141 aaatccccag agccaaggca gaagaaatgc ttagcaaaca gcggcacgat ggggcctttc
     1201 ttatccgaga gagtgagagc gctcctgggg acttctccct ctctgtcaag tttggaaacg
     1261 atgtgcagca cttcaaggtg ctccgagatg gagccgggaa gtacttcctc tgggtggtga
     1321 agttcaattc tttgaatgag ctggtggatt atcacagatc tacatctgtc tccagaaacc
     1381 agcagatatt cctgcgggac atagaacagg tgtaagtcga ctcgagcggc cgcatcgtga
     1441 ctgactgacg atctgcctcg cgcgtttcgg tgatgacggt gaaaacctct gacacatgca
     1501 gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac aagcccgtca
     1561 gggcgcgtca gcgggtgttg gcgggtgtcg gggcgcagcc atgacccagt cacgtagcga
     1621 tagcggagtg tataattctt gaagacgaaa gggcctcgtg atacgcctat ttttataggt
     1681 taatgtcatg ataataatgg tttcttagac gtcaggtggc acttttcggg gaaatgtgcg
     1741 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     1801 ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt
     1861 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga
     1921 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga
     1981 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat
     2041 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtgttg acgccgggca
     2101 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt
     2161 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac
     2221 catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct
     2281 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga
     2341 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgcag caatggcaac
     2401 aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat
     2461 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg
     2521 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc
     2581 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc
     2641 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg
     2701 gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta
     2761 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     2821 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     2881 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     2941 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     3001 agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa
     3061 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     3121 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     3181 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     3241 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     3301 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     3361 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     3421 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     3481 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     3541 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     3601 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc tgatgcggta
     3661 ttttctcctt acgcatctgt gcggtatttc acaccgcata aattccgaca ccatcgaatg
     3721 gtgcaaaacc tttcgcggta tggcatgata gcgcccggaa gagagtcaat tcagggtggt
     3781 gaatgtgaaa ccagtaacgt tatacgatgt cgcagagtat gccggtgtct cttatcagac
     3841 cgtttcccgc gtggtgaacc aggccagcca cgtttctgcg aaaacgcggg aaaaagtgga
     3901 agcggcgatg gcggagctga attacattcc caaccgcgtg gcacaacaac tggcgggcaa
     3961 acagtcgttg ctgattggcg ttgccacctc cagtctggcc ctgcacgcgc cgtcgcaaat
     4021 tgtcgcggcg attaaatctc gcgccgatca actgggtgcc agcgtggtgg tgtcgatggt
     4081 agaacgaagc ggcgtcgaag cctgtaaagc ggcggtgcac aatcttctcg cgcaacgcgt
     4141 cagtgggctg atcattaact atccgctgga tgaccaggat gccattgctg tggaagctgc
     4201 ctgcactaat gttccggcgt tatttcttga tgtctctgac cagacaccca tcaacagtat
     4261 tattttctcc catgaagacg gtacgcgact gggcgtggag catctggtcg cattgggtca
     4321 ccagcaaatc gcgctgttag cgggcccatt aagttctgtc tcggcgcgtc tgcgtctggc
     4381 tggctggcat aaatatctca ctcgcaatca aattcagccg atagcggaac gggaaggcga
     4441 ctggagtgcc atgtccggtt ttcaacaaac catgcaaatg ctgaatgagg gcatcgttcc
     4501 cactgcgatg ctggttgcca acgatcagat ggcgctgggc gcaatgcgcg ccattaccga
     4561 gtccgggctg cgcgttggtg cggatatctc ggtagtggga tacgacgata ccgaagacag
     4621 ctcatgttat atcccgccgt caaccaccat caaacaggat tttcgcctgc tggggcaaac
     4681 cagcgtggac cgcttgctgc aactctctca gggccaggcg gtgaagggca atcagctgtt
     4741 gcccgtctca ctggtgaaaa gaaaaaccac cctggcgccc aatacgcaaa ccgcctctcc
     4801 ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac tggaaagcgg
     4861 gcagtgagcg caacgcaatt aatgtgagtt agctcactca ttaggcaccc caggctttac
     4921 actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa tttcacacag
     4981 gaaacagcta tgaccatgat tacggattca ctggccgtcg ttttacaacg tcgtgactgg
     5041 gaaaaccctg gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg
     5101 cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc
     5161 gaatggcgct ttgcctggtt tccggcacca gaagcggtgc cggaaagctg gctggagtgc
     5221 gatcttcctg aggccgatac tgtcgtcgtc ccctcaaact ggcagatgca cggttacgat
     5281 gcgcccatct acaccaacgt aacctatccc attacggtca atccgccgtt tgttcccacg
     5341 gagaatccga cgggttgtta ctcgctcaca tttaatgttg atgaaagctg gctacaggaa
     5401 ggccagacgc gaattatttt tgatggcgtt ggaatt
//
