LOCUS       sequence-142446-        5739 bp    DNA     circular SYN 14-APR-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5739)
  AUTHORS   Yaffe MB, Rittinger K, Volinia S, Caron PR, Aitken A, Leffers H, 
            Gamblin SJ, Smerdon SJ, Cantley LC
  TITLE     The structural basis for 14-3-3:phosphopeptide binding specificity.
  JOURNAL   Cell. 1997 Dec 26. 91(7):961-71.
  PUBMED    9428519
REFERENCE   2  (bases 1 to 5739)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Cell. 1997 
            Dec 26. 91(7):961-71."
COMMENT     Sequence Label: pGEX-4T1-14-3-3 epsilon GST
FEATURES             Location/Qualifiers
     source          1..5739
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        63..140
                     /gene="lacI"
                     /label=lacI promoter
     CDS             141..1223
                     /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(160..179)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1272..1277,1278..1295,1296..1302)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1310..1326
                     /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     1334..1350
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1334..1350
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             1346..1873
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(1366..1383)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1366..1382)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1375..1397)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     promoter        join(1975..1980,1981..1996,1997..2003)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    2011..2027
                     /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             2050..2703
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2661..2683
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2710..2727
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(3581..3603)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3748..3766)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3834..3938
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3939..4007,4008..4799)
                     /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(4157..4176)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4970..5558
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5459..5478
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5712..5729
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 agcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcataaatt
       61 ccgacaccat cgaatggcgc aaaacctttc gcggtatggc atgatagcgc ccggaagaga
      121 gtcaattcag ggtggtgaat gtgaaaccag taacgttata cgatgtcgca gagtatgccg
      181 gtgtctctta tcagaccgtt tcccgcgtgg tgaaccaggc cagccacgtt tctgcgaaaa
      241 cgcgggaaaa agtggaagcg gcgatggcgg agctgaatta cattcccaac cgcgtggcac
      301 aacaactggc gggcaaacag tcgttgctga ttggcgttgc cacctccagt ctggccctgc
      361 acgcgccgtc gcaaattgtc gcggcgatta aatctcgcgc cgatcaactg ggtgccagcg
      421 tggtggtgtc gatggtagaa cgaagcggcg tcgaagcctg taaagcggcg gtgcacaatc
      481 ttctcgcgca acgcgtcagt gggctgatca ttaactatcc gctggatgac caggatgcca
      541 ttgctgtgga agctgcctgc actaatgttc cggcgttatt tcttgatgtc tctgaccaga
      601 cacccatcaa cagtattatt ttctcccatg aagacggtac gcgactgggc gtggagcatc
      661 tggtcgcatt gggtcaccag caaatcgcgc tgttagcggg cccattaagt tctgtctcgg
      721 cgcgtctgcg tctggctggc tggcataaat atctcactcg caatcaaatt cagccgatag
      781 cggaacggga aggcgactgg agtgccatgt ccggttttca acaaaccatg caaatgctga
      841 atgagggcat cgttcccact gcgatgctgg ttgccaacga tcagatggcg ctgggcgcaa
      901 tgcgcgccat taccgagtcc gggctgcgcg ttggtgcgga tatctcggta gtgggatacg
      961 acgataccga agacagctca tgttatatcc cgccgttaac caccatcaaa caggattttc
     1021 gcctgctggg gcaaaccagc gtggaccgct tgctgcaact ctctcagggc caggcggtga
     1081 agggcaatca gctgttgccc gtctcactgg tgaaaagaaa aaccaccctg gcgcccaata
     1141 cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt
     1201 cccgactgga aagcgggcag tgagcgcaac gcaattaatg taagttagct cactcattag
     1261 gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga
     1321 taacaatttc acacaggaaa cagctatgac catgattacg gattcactgg ccgtcgtttt
     1381 acaacgtcgt gactgggaaa accctggcgt tacccaactt aatcgccttg cagcacatcc
     1441 ccctttcgcc agctggcgta atagcgaaga ggcccgcacc gatcgccctt cccaacagtt
     1501 gcgcagcctg aatggcgaat ggcgctttgc ctggtttccg gcaccagaag cggtgccgga
     1561 aagctggctg gagtgcgatc ttcctgaggc cgatactgtc gtcgtcccct caaactggca
     1621 gatgcacggt tacgatgcgc ccatctacac caacgtgacc tatcccatta cggtcaatcc
     1681 gccgtttgtt cccacggaga atccgacggg ttgttactcg ctcacattta atgttgatga
     1741 aagctggcta caggaaggcc agacgcgaat tatttttgat ggcgttggaa ttagcttatc
     1801 gactgcacgg tgcaccaatg cttctggcgt caggcagcca tcggaagctg tggtatggct
     1861 gtgcaggtcg taaatcactg cataattcgt gtcgctcaag gcgcactccc gttctggata
     1921 atgttttttg cgccgacatc ataacggttc tggcaaatat tctgaaatga gctgttgaca
     1981 attaatcatc ggctcgtata atgtgtggaa ttgtgagcgg ataacaattt cacacaggaa
     2041 acagtattca tgtcccctat actaggttat tggaaaatta agggccttgt gcaacccact
     2101 cgacttcttt tggaatatct tgaagaaaaa tatgaagagc atttgtatga gcgcgatgaa
     2161 ggtgataaat ggcgaaacaa aaagtttgaa ttgggtttgg agtttcccaa tcttccttat
     2221 tatattgatg gtgatgttaa attaacacag tctatggcca tcatacgtta tatagctgac
     2281 aagcacaaca tgttgggtgg ttgtccaaaa gagcgtgcag agatttcaat gcttgaagga
     2341 gcggttttgg atattagata cggtgtttcg agaattgcat atagtaaaga ctttgaaact
     2401 ctcaaagttg attttcttag caagctacct gaaatgctga aaatgttcga agatcgttta
     2461 tgtcataaaa catatttaaa tggtgatcat gtaacccatc ctgacttcat gttgtatgac
     2521 gctcttgatg ttgttttata catggaccca atgtgcctgg atgcgttccc aaaattagtt
     2581 tgttttaaaa aacgtattga agctatccca caaattgata agtacttgaa atccagcaag
     2641 tatatagcat ggcctttgca gggctggcaa gccacgtttg gtggtggcga ccatcctcca
     2701 aaatcggatc tggttccgcg tggatccacc atggatgatc gggaggatct ggtgtaccag
     2761 gcgaagctgg cagagcaggc cgagcgatac gacgaaatgg tggaatcaat gaagaaagta
     2821 gcaggaatgg acgtggagct gacagttgaa gaacgaaacc ttttatctgt tgcatataaa
     2881 aatgtgattg gagccagaag agcatcctgg agaataatca gcagcattga acagaaggaa
     2941 gaaaacaagg gaggagagga caaattaaag atgattcggg agtaccggca aatggttgaa
     3001 actgagctca agttaatctg ttgtgacatt ctggatgtac tggacaagca cctcattcca
     3061 gcagctaaca ctggcgagtc caaggttttc tattataaaa tgaaagggga ctaccacagg
     3121 tatctggctg agtttgccac aggaaacgac aggaaggagg cagcagagaa cagccttgtg
     3181 gcttacaaag ctgctagtga cattgcgatg acagaacttc ctccaacgca ccccattcgt
     3241 ttaggtcttg ctctcaactt ttccgtattc tactatgaaa ttcttaattc ccccgaccgt
     3301 gcctgcaggt tggcaaaagc agcttttgat gacgcaattg cagaactgga cacgctgagt
     3361 gaagaaagct ataaggactc cacactcatc atgcagctgc tacgtgataa cctgacgctg
     3421 tggacctcag acatgcaggg cgatggtgaa gagcagaata aagaagcgct gcaggatgtg
     3481 gaagatgaga atcagtgatc tagagttcgg gtcgactcga gcggccgcat cgtgactgac
     3541 tgacgatctg cctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac atgcagctcc
     3601 cggagacggt cacagcttgt ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg
     3661 cgtcagcggg tgttggcggg tgtcggggcg cagccatgac ccagtcacgt agcgatagcg
     3721 gagtgtataa ttcttgaaga cgaaagggcc tcgtgatacg cctattttta taggttaatg
     3781 tcatgataat aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa
     3841 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     3901 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg
     3961 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
     4021 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
     4081 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
     4141 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tgttgacgcc gggcaagagc
     4201 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
     4261 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
     4321 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
     4381 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
     4441 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgcagcaatg gcaacaacgt
     4501 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     4561 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     4621 ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg
     4681 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     4741 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac
     4801 tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta
     4861 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     4921 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
     4981 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
     5041 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
     5101 agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
     5161 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
     5221 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     5281 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
     5341 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
     5401 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
     5461 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat
     5521 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt
     5581 tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg
     5641 attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa
     5701 cgaccgagcg cagcgagtca gtgagcgagg aagcggaag
//
