LOCUS       sequence-248404-        7445 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 7445)
  AUTHORS   Oda H, Kumar S, Howley PM
  TITLE     Regulation of the Src family tyrosine kinase Blk through 
            E6AP-mediated ubiquitination.
  JOURNAL   Proc Natl Acad Sci U S A 1999 Aug 17;96(17):9557-62.
  PUBMED    10449731
REFERENCE   2  (bases 1 to 7445)
  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"; date: "1999-08-17"; volume: "96(17)"; pages: 
            "9557-62"
COMMENT     Sequence Label: p4028 GST-E6AP
FEATURES             Location/Qualifiers
     source          1..7445
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     411..430
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     664..681
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        754..831
                     /gene="lacI"
                     /label=lacI promoter
     CDS             832..1914
                     /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(851..870)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1963..1968,1969..1986,1987..1993)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2001..2017
                     /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     2025..2041
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2025..2041
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             2037..2564
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(2057..2074)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2057..2073)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2066..2088)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     promoter        join(2666..2671,2672..2687,2688..2694)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    2702..2718
                     /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             2741..3394
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     3352..3374
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             3401..3418
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(5978..6000)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6145..6163)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6231..6335
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6336..6404,6405..7196)
                     /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(6554..6573)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(7367..7445,1..510)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
       61 aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
      121 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
      181 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
      241 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
      301 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
      361 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
      421 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
      481 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
      541 ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
      601 gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag
      661 cgcagcgagt cagtgagcga ggaagcggaa gagcgcctga tgcggtattt tctccttacg
      721 catctgtgcg gtatttcaca ccgcataaat tccgacacca tcgaatggcg caaaaccttt
      781 cgcggtatgg catgatagcg cccggaagag agtcaattca gggtggtgaa tgtgaaacca
      841 gtaacgttat acgatgtcgc agagtatgcc ggtgtctctt atcagaccgt ttcccgcgtg
      901 gtgaaccagg ccagccacgt ttctgcgaaa acgcgggaaa aagtggaagc ggcgatggcg
      961 gagctgaatt acattcccaa ccgcgtggca caacaactgg cgggcaaaca gtcgttgctg
     1021 attggcgttg ccacctccag tctggccctg cacgcgccgt cgcaaattgt cgcggcgatt
     1081 aaatctcgcg ccgatcaact gggtgccagc gtggtggtgt cgatggtaga acgaagcggc
     1141 gtcgaagcct gtaaagcggc ggtgcacaat cttctcgcgc aacgcgtcag tgggctgatc
     1201 attaactatc cgctggatga ccaggatgcc attgctgtgg aagctgcctg cactaatgtt
     1261 ccggcgttat ttcttgatgt ctctgaccag acacccatca acagtattat tttctcccat
     1321 gaagacggta cgcgactggg cgtggagcat ctggtcgcat tgggtcacca gcaaatcgcg
     1381 ctgttagcgg gcccattaag ttctgtctcg gcgcgtctgc gtctggctgg ctggcataaa
     1441 tatctcactc gcaatcaaat tcagccgata gcggaacggg aaggcgactg gagtgccatg
     1501 tccggttttc aacaaaccat gcaaatgctg aatgagggca tcgttcccac tgcgatgctg
     1561 gttgccaacg atcagatggc gctgggcgca atgcgcgcca ttaccgagtc cgggctgcgc
     1621 gttggtgcgg atatctcggt agtgggatac gacgataccg aagacagctc atgttatatc
     1681 ccgccgttaa ccaccatcaa acaggatttt cgcctgctgg ggcaaaccag cgtggaccgc
     1741 ttgctgcaac tctctcaggg ccaggcggtg aagggcaatc agctgttgcc cgtctcactg
     1801 gtgaaaagaa aaaccaccct ggcgcccaat acgcaaaccg cctctccccg cgcgttggcc
     1861 gattcattaa tgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa
     1921 cgcaattaat gtaagttagc tcactcatta ggcaccccag gctttacact ttatgcttcc
     1981 ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa acagctatga
     2041 ccatgattac ggattcactg gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg
     2101 ttacccaact taatcgcctt gcagcacatc cccctttcgc cagctggcgt aatagcgaag
     2161 aggcccgcac cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa tggcgctttg
     2221 cctggtttcc ggcaccagaa gcggtgccgg aaagctggct ggagtgcgat cttcctgagg
     2281 ccgatactgt cgtcgtcccc tcaaactggc agatgcacgg ttacgatgcg cccatctaca
     2341 ccaacgtgac ctatcccatt acggtcaatc cgccgtttgt tcccacggag aatccgacgg
     2401 gttgttactc gctcacattt aatgttgatg aaagctggct acaggaaggc cagacgcgaa
     2461 ttatttttga tggcgttgga attagcttat cgactgcacg gtgcaccaat gcttctggcg
     2521 tcaggcagcc atcggaagct gtggtatggc tgtgcaggtc gtaaatcact gcataattcg
     2581 tgtcgctcaa ggcgcactcc cgttctggat aatgtttttt gcgccgacat cataacggtt
     2641 ctggcaaata ttctgaaatg agctgttgac aattaatcat cggctcgtat aatgtgtgga
     2701 attgtgagcg gataacaatt tcacacagga aacagtattc atgtccccta tactaggtta
     2761 ttggaaaatt aagggccttg tgcaacccac tcgacttctt ttggaatatc ttgaagaaaa
     2821 atatgaagag catttgtatg agcgcgatga aggtgataaa tggcgaaaca aaaagtttga
     2881 attgggtttg gagtttccca atcttcctta ttatattgat ggtgatgtta aattaacaca
     2941 gtctatggcc atcatacgtt atatagctga caagcacaac atgttgggtg gttgtccaaa
     3001 agagcgtgca gagatttcaa tgcttgaagg agcggttttg gatattagat acggtgtttc
     3061 gagaattgca tatagtaaag actttgaaac tctcaaagtt gattttctta gcaagctacc
     3121 tgaaatgctg aaaatgttcg aagatcgttt atgtcataaa acatatttaa atggtgatca
     3181 tgtaacccat cctgacttca tgttgtatga cgctcttgat gttgttttat acatggaccc
     3241 aatgtgcctg gatgcgttcc caaaattagt ttgttttaaa aaacgtattg aagctatccc
     3301 acaaattgat aagtacttga aatccagcaa gtatatagca tggcctttgc agggctggca
     3361 agccacgttt ggtggtggcg accatcctcc aaaatcggat ctggttccgc gtggatccac
     3421 catggaagcc tgcacgaatg agttttgtgc ttcctgtcca acttttcttc gtatggataa
     3481 taaagcagca gctattaaag ccctcgagct ttataagatt aatgcaaaac tctgtgatcc
     3541 tcatccctcc aagaaaggag caagctcagc ttaccttgag aactcgaaag gtgcccccaa
     3601 caactcctgc tctgagataa aaatgaacaa gaaaggcgct agaattgatt ttaaagatgt
     3661 gacttactta acagaagaga aggtatatga aattcttgaa ttatgtagag aaagagagga
     3721 ttattcccct ttaatccgtg ttattggaag agttttttct agtgctgagg cattggtaca
     3781 gagcttccgg aaagttaaac aacacaccaa ggaagaactg aaatctcttc aagcaaaaga
     3841 tgaagacaaa gatgaagatg aaaaggaaaa agctgcatgt tctgctgctg ctatggaaga
     3901 agactcagaa gcatcttcct caaggatagg tgatagctca cagggagaca acaatttgca
     3961 aaaattaggc cctgatgatg tgtctgtgga tattgatgcc attagaaggg tctacaccag
     4021 attgctctct aatgaaaaaa ttgaaactgc ctttctcaat gcacttgtat atttgtcacc
     4081 taacgtggaa tgtgacttga cgtatcacaa tgtatactct cgagatccta attatctgaa
     4141 tttgttcatt atcgtaatgg agaatagaaa tctccacagt cctgaatatc tggaaatggc
     4201 tttgccatta ttttgcaaag cgatgagcaa gctacccctt gcagcccaag gaaaactgat
     4261 cagactgtgg tctaaataca atgcagacca gattcggaga atgatggaga catttcagca
     4321 acttattacc tataaagtca taagcaatga atttaacagt cgaaatctag tgaatgatga
     4381 tgatgccatt gttgctgctt cgaagtgctt gaaaatggtt tactatgcaa atgtagtggg
     4441 aggggaagtg gacacaaatc acaatgaaga agatgatgaa gagcccatcc ctgagtccag
     4501 cgagctgaca cttcaggaac ttttgggaga agaaagaaga aacaagaaag gtcctcgagt
     4561 ggaccccctg gaaaccgaac ttggtgttaa aaccctggat tgtcgaaaac cacttatccc
     4621 ttttgaagag tttattaatg aaccactgaa tgaggttcta gaaatggata aagattatac
     4681 ttttttcaaa gtagaaacag agaacaaatt ctcttttatg acatgtccct ttatattgaa
     4741 tgctgtcaca aagaatttgg gattatatta tgacaataga attcgcatgt acagtgaacg
     4801 aagaatcact gttctctaca gcttagttca aggacagcag ttgaatccat atttgagact
     4861 caaagttaga cgtgaccata tcatagatga tgcacttgtc cggctagaga tgatcgctat
     4921 ggaaaatcct gcagacttga agaagcagtt gtatgtggaa tttgaaggag aacaaggagt
     4981 tgatgaggga ggtgtttcca aagaattttt tcagctggtt gtggaggaaa tcttcaatcc
     5041 agatattggt atgttcacat acgatgaatc tacaaaattg ttttggttta atccatcttc
     5101 ttttgaaact gagggtcagt ttactctgat tggcatagta ctgggtctgg ctatttacaa
     5161 taactgtata ctggatgtac attttcccat ggttgtctac aggaagctaa tggggaaaaa
     5221 aggaactttt cgtgacttgg gagactctca cccagttcta tatcagagtt taaaagattt
     5281 attggagtat gaagggaatg tggaagatga catgatgatc actttccaga tatcacagac
     5341 agatcttttt ggtaacccaa tgatgtatga tctaaaggaa aatggtgata aaattccaat
     5401 tacaaatgaa aacaggaagg aatttgtcaa tctttattct gactacattc tcaataaatc
     5461 agtagaaaaa cagttcaagg cttttcggag aggttttcat atggtgacca atgaatctcc
     5521 cttaaagtac ttattcagac cagaagaaat tgaattgctt atatgtggaa gccggaatct
     5581 agatttccaa gcactagaag aaactacaga atatgacggt ggctatacca gggactctgt
     5641 tctgattagg gagttctggg aaatcgttca ttcatttaca gatgaacaga aaagactctt
     5701 cttgcagttt acaacgggca cagacagagc acctgtggga ggactaggaa aattaaagat
     5761 gattatagcc aaaaatggcc cagacacaga aaggttacct acatctcata cttgctttaa
     5821 tgtgctttta cttccggaat actcaagcaa agaaaaactt aaagagagat tgttgaaggc
     5881 catcacgtat gccaaaggat ttggcatgct gtaaaacgcg gccgcatcgt gactgactga
     5941 cgatctgcct cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg
     6001 agacggtcac agcttgtctg taagcggatg ccgggagcag acaagcccgt cagggcgcgt
     6061 cagcgggtgt tggcgggtgt cggggcgcag ccatgaccca gtcacgtagc gatagcggag
     6121 tgtataattc ttgaagacga aagggcctcg tgatacgcct atttttatag gttaatgtca
     6181 tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc
     6241 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     6301 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
     6361 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     6421 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
     6481 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
     6541 cttttaaagt tctgctatgt ggcgcggtat tatcccgtgt tgacgccggg caagagcaac
     6601 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
     6661 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     6721 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
     6781 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     6841 aagccatacc aaacgacgag cgtgacacca cgatgcctgc agcaatggca acaacgttgc
     6901 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
     6961 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     7021 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     7081 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     7141 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
     7201 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     7261 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
     7321 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     7381 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     7441 tgccg
//
