LOCUS       sequence-65577-e        5481 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5481)
  AUTHORS   Machida K, Thompson CM, Dierck K, Jablonowski K, Karkkainen S, Liu 
            B, Zhang H, Nash PD, Newman DK, Nollau P, Pawson T, Renkema GH, 
            Saksela K, Schiller MR, Shin DG, Mayer BJ
  TITLE     High-throughput phosphotyrosine profiling using SH2 domains.
  JOURNAL   Mol Cell. 2007 Jun 22;26(6):899-915.
  PUBMED    17588523
REFERENCE   2  (bases 1 to 5481)
  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: "Mol Cell.";
            date: "2007-06-22"; volume: "26(6)"; pages: "899-915"
COMMENT     Sequence Label: pAC4 Frk-SH2
FEATURES             Location/Qualifiers
     source          1..5481
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(193..198,199..215,216..222)
                     /label=trc promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    230..246
                     /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)."
     promoter        join(470..475,476..492,493..499)
                     /label=trc promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    507..523
                     /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             556..1209
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     1167..1189
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             1216..1233
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1570..1614
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     primer_bind     complement(1664..1681)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1834..1920
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      2012..2039
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        2059..2150
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2151..2219,2220..3011)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPTAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2369..2388)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3182..3770
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3671..3690
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3924..3941
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3956..4096
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4022..4041)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4182..4204
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     promoter        4282..4359
                     /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             4360..5442
                     /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(4379..4398)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 gtttgacagc ttatcatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc
       61 ggaagctgtg gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc
      121 gcactcccgt tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc
      181 tgaaatgagc tgttgacaat taatcatccg gctcgtataa tgtgtggaat tgtgagcgga
      241 taacaatttc acacaggaaa cagaccatgg agctcgagtt tgacagctta tcatcgactg
      301 cacggtgcac caatgcttct ggcgtcaggc agccatcgga agctgtggta tggctgtgca
      361 ggtcgtaaat cactgcataa ttcgtgtcgc tcaaggcgca ctcccgttct ggataatgtt
      421 ttttgcgccg acatcataac ggttctggca aatattctga aatgagctgt tgacaattaa
      481 tcatccggct cgtataatgt gtggaattgt gagcggataa caatttcaca caggaaacag
      541 accatggagc tcgagatgtc ccctatacta ggttattgga aaattaaggg ccttgtgcaa
      601 cccactcgac ttcttttgga atatcttgaa gaaaaatatg aagagcattt gtatgagcgc
      661 gatgaaggtg ataaatggcg aaacaaaaag tttgaattgg gtttggagtt tcccaatctt
      721 ccttattata ttgatggtga tgttaaatta acacagtcta tggccatcat acgttatata
      781 gctgacaagc acaacatgtt gggtggttgt ccaaaagagc gtgcagagat ttcaatgctt
      841 gaaggagcgg ttttggatat tagatacggt gtttcgagaa ttgcatatag taaagacttt
      901 gaaactctca aagttgattt tcttagcaag ctacctgaaa tgctgaaaat gttcgaagat
      961 cgtttatgtc ataaaacata tttaaatggt gatcatgtaa cccatcctga cttcatgttg
     1021 tatgacgctc ttgatgttgt tttatacatg gacccaatgt gcctggatgc gttcccaaaa
     1081 ttagtttgtt ttaaaaaacg tattgaagct atcccacaaa ttgataagta cttgaaatcc
     1141 agcaagtata tagcatggcc tttgcagggc tggcaagcca cgtttggtgg tggcgaccat
     1201 cctccaaaat cggatctggt tccgcgtgga tccgacagaa gcctacaggc agagccgtgg
     1261 ttctttggag caatcggaag atcagatgca gagaaacaac tattatattc agaaaacaag
     1321 accggttcct ttctaatcag agaaagtgaa agccaaaaag gagaattctc tctttcagtt
     1381 ttagatggag cagttgtaaa acactacaga attaaaagac tggatgaagg gggatttttt
     1441 ctcacgcgaa gaagaatctt ttcaacactg aacgaatttg tgagccacta caccaagaca
     1501 agtgacggcc tgtgtgtcaa gctggggaaa ccatgcttaa agatccaggt cgaaagcttg
     1561 cttggtggcg gtctgaacga catcttcgag gctcagaaaa tcgaatggca cgaataatta
     1621 attaagagct tggctgtttt ggcggatgag agaagatttt cagcctgata cagattaaat
     1681 cagaacgcag aagcggtctg ataaaacaga atttgcctgg cggcagtagc gcggtggtcc
     1741 cacctgaccc catgccgaac tcagaagtga aacgccgtag cgccgatggt agtgtggggt
     1801 ctccccatgc gagagtaggg aactgccagg catcaaataa aacgaaaggc tcagtcgaaa
     1861 gactgggcct ttcgttttat ctgttgtttg tcggtgaacg ctctcctgag taggacaaat
     1921 ccgccgggag cggatttgaa cgttgcgaag caacggcccg gagggtggcg ggcaggacgc
     1981 ccgccataaa ctgccaggca tcaaattaag cagaaggcca tcctgacgga tggccttttt
     2041 gcgtttctac aaactctttt tgtttatttt tctaaataca ttcaaatatg tatccgctca
     2101 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc
     2161 aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc
     2221 acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt
     2281 acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt
     2341 ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtgttgacg
     2401 ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact
     2461 caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg
     2521 ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga
     2581 aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg
     2641 aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctacagcaa
     2701 tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac
     2761 aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc
     2821 cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca
     2881 ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga
     2941 gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta
     3001 agcattggta actgtcagac caagtttact catatatact ttagattgat ttaaaacttc
     3061 atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc
     3121 cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt
     3181 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac
     3241 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct
     3301 tcagcagagc gcagatacca aatactgtcc ttctagtgta gccgtagtta ggccaccact
     3361 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg
     3421 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata
     3481 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga
     3541 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag
     3601 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg
     3661 agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac
     3721 ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca
     3781 acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg
     3841 cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc
     3901 gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcctga
     3961 tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcatatgg tgcactctca
     4021 gtacaatctg ctctgatgcc gcatagttaa gccagtatac actccgctat cgctacgtga
     4081 ctgggtcatg gctgcgcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg
     4141 tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca
     4201 gaggttttca ccgtcatcac cgaaacgcgc gaggcagcag atcaattcgc gcgcgaaggc
     4261 gaagcggcat gcatttacgt tgacaccatc gaatggtgca aaacctttcg cggtatggca
     4321 tgatagcgcc cggaagagag tcaattcagg gtggtgaatg tgaaaccagt aacgttatac
     4381 gatgtcgcag agtatgccgg tgtctcttat cagaccgttt cccgcgtggt gaaccaggcc
     4441 agccacgttt ctgcgaaaac gcgggaaaaa gtggaagcgg cgatggcgga gctgaattac
     4501 attcccaacc gcgtggcaca acaactggcg ggcaaacagt cgttgctgat tggcgttgcc
     4561 acctccagtc tggccctgca cgcgccgtcg caaattgtcg cggcgattaa atctcgcgcc
     4621 gatcaactgg gtgccagcgt ggtggtgtcg atggtagaac gaagcggcgt cgaagcctgt
     4681 aaagcggcgg tgcacaatct tctcgcgcaa cgcgtcagtg ggctgatcat taactatccg
     4741 ctggatgacc aggatgccat tgctgtggaa gctgcctgca ctaatgttcc ggcgttattt
     4801 cttgatgtct ctgaccagac acccatcaac agtattattt tctcccatga agacggtacg
     4861 cgactgggcg tggagcatct ggtcgcattg ggtcaccagc aaatcgcgct gttagcgggc
     4921 ccattaagtt ctgtctcggc gcgtctgcgt ctggctggct ggcataaata tctcactcgc
     4981 aatcaaattc agccgatagc ggaacgggaa ggcgactgga gtgccatgtc cggttttcaa
     5041 caaaccatgc aaatgctgaa tgagggcatc gttcccactg cgatgctggt tgccaacgat
     5101 cagatggcgc tgggcgcaat gcgcgccatt accgagtccg ggctgcgcgt tggtgcggat
     5161 atctcggtag tgggatacga cgataccgaa gacagctcat gttatatccc gccgttaacc
     5221 accatcaaac aggattttcg cctgctgggg caaaccagcg tggaccgctt gctgcaactc
     5281 tctcagggcc aggcggtgaa gggcaatcag ctgttgcccg tctcactggt gaaaagaaaa
     5341 accaccctgg cgcccaatac gcaaaccgcc tctccccgcg cgttggccga ttcattaatg
     5401 cagctggcac gacaggtttc ccgactggaa agcgggcagt gagcgcaacg caattaatgt
     5461 gagttagcgc gaattgatct g
//
