LOCUS       sequence-258680-        6349 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Bacterial expression of ETS gene with A86D mutation for solubility.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6349)
  AUTHORS   Selvaraj N, Kedage V, Hollenhorst PC
  TITLE     Comparison of MAPK specificity across the ETS transcription factor 
            family identifies a high-affinity ERK interaction required for ERG 
            function in prostate cells.
  JOURNAL   Cell Commun Signal. 2015 Feb 19;13:12. doi: 
            10.1186/s12964-015-0089-7.
  PUBMED    25885538
REFERENCE   2  (bases 1 to 6349)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Cell Commun
            Signal."; date: "2015-02-19"; volume: "13:12. doi"; pages: " 
            10.1186/s12964-015-0089-7"
COMMENT     Sequence Label: ETV7_A86D_pet28a
FEATURES             Location/Qualifiers
     source          1..6349
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(26..73)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     69..87
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(140..157)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(1227..1244)
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             complement(1254..1271)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             1290..1312
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    1327..1351
                     /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     complement(1351..1370)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1352..1370)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1454..1473)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1679..1756
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1757..2839
                     /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(1776..1795)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3648..3839
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3833..3855)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3941..4080
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3996..4015
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4095..4112)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4266..4854)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4346..4365)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4965..4984)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4976..5791
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             4976..5791
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(5049..5068)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(5883..6338)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6020..6041)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6232..6251
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
       61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
      121 tgttagcagc cggatctcag tggtggtggt ggtggtgctc gagtgcggcc gcaagcttgt
      181 cgacggagct cgaattctca cggagagatt tctggcctct tgtccttgaa ctctattctg
      241 tcctgctcct ggctctccag cggctccagg tggctgtgct tgtcctggac catctttccc
      301 ggagtcttta gaaatctgaa caggagtttc tgccccggtt ccttcttaat gatattaagc
      361 ttataatagt ggcgcagggc acgagacatc ttctcgtagg tcatgttcac ccggttcttg
      421 tgatttcccc agagtctggc gagcccattt ggatccacaa ctcggaagat cttggcgtcc
      481 ttgtcttccc acttgatgta gggctcatat cgggtatcaa ggagcagctg atacacgtaa
      541 tcccacagca ggcggcagtc agcgatcctg ccgtcaatgg gggcctgcgg catcgcgggg
      601 aaggaacaga ccccctgggt cctgcagccg agctctgcac agtgacataa gttgagggac
      661 tcctccttgc caggggtcca ccttgccagg ccagggtcat ccaggtggcc gaagttgctg
      721 gtaagccctg ggtctggtgg ctgcagcagg tggccccttc gggtgtccat ctgagagggg
      781 ccagtcacct cttccggggg gactggagag tgctgggtgg gcgtcttcag cctgaagatc
      841 cctccaaaaa agggcccaca caccagggct cgccgctggg tcttgatgta ctggagcagc
      901 tcatacagga cgtcacctga gctgggcgca cggtgccgga agtcgtcctt ggtgaggatg
      961 cagaggtcgc gtccgttcat ctcgaacccg tgctccgcgg tgcatggcag agagtactcc
     1021 tgctctgccc agcgcagcca gtgcagcacg tcctccctgc tccacagtgc gggctggatg
     1081 cggagtcttc ctggcagctt gcagatcccc ccttcaccca gcaggttaat ttgagcttca
     1141 catctggctt gcacgtgggt gcctagggga ggcatggctg ccacagggct tataggagaa
     1201 atagccaatt ctccctcctg catatggctg ccgcgcggca ccaggccgct gctgtgatga
     1261 tgatgatgat ggctgctgcc catggtatat ctccttctta aagttaaaca aaattatttc
     1321 tagaggggaa ttgttatccg ctcacaattc ccctatagtg agtcgtatta atttcgcggg
     1381 atcgagatct cgatcctcta cgccggacgc atcgtggccg gcatcaccgg cgccacaggt
     1441 gcggttgctg gcgcctatat cgccgacatc accgatgggg aagatcgggc tcgccacttc
     1501 gggctcatga gcgcttgttt cggcgtgggt atggtggcag gccccgtggc cgggggactg
     1561 ttgggcgcca tctccttgca tgcaccattc cttgcggcgg cggtgctcaa cggcctcaac
     1621 ctactactgg gctgcttcct aatgcaggag tcgcataagg gagagcgtcg agatcccgga
     1681 caccatcgaa tggcgcaaaa cctttcgcgg tatggcatga tagcgcccgg aagagagtca
     1741 attcagggtg gtgaatgtga aaccagtaac gttatacgat gtcgcagagt atgccggtgt
     1801 ctcttatcag accgtttccc gcgtggtgaa ccaggccagc cacgtttctg cgaaaacgcg
     1861 ggaaaaagtg gaagcggcga tggcggagct gaattacatt cccaaccgcg tggcacaaca
     1921 actggcgggc aaacagtcgt tgctgattgg cgttgccacc tccagtctgg ccctgcacgc
     1981 gccgtcgcaa attgtcgcgg cgattaaatc tcgcgccgat caactgggtg ccagcgtggt
     2041 ggtgtcgatg gtagaacgaa gcggcgtcga agcctgtaaa gcggcggtgc acaatcttct
     2101 cgcgcaacgc gtcagtgggc tgatcattaa ctatccgctg gatgaccagg atgccattgc
     2161 tgtggaagct gcctgcacta atgttccggc gttatttctt gatgtctctg accagacacc
     2221 catcaacagt attattttct cccatgaaga cggtacgcga ctgggcgtgg agcatctggt
     2281 cgcattgggt caccagcaaa tcgcgctgtt agcgggccca ttaagttctg tctcggcgcg
     2341 tctgcgtctg gctggctggc ataaatatct cactcgcaat caaattcagc cgatagcgga
     2401 acgggaaggc gactggagtg ccatgtccgg ttttcaacaa accatgcaaa tgctgaatga
     2461 gggcatcgtt cccactgcga tgctggttgc caacgatcag atggcgctgg gcgcaatgcg
     2521 cgccattacc gagtccgggc tgcgcgttgg tgcggatatc tcggtagtgg gatacgacga
     2581 taccgaagac agctcatgtt atatcccgcc gttaaccacc atcaaacagg attttcgcct
     2641 gctggggcaa accagcgtgg accgcttgct gcaactctct cagggccagg cggtgaaggg
     2701 caatcagctg ttgcccgtct cactggtgaa aagaaaaacc accctggcgc ccaatacgca
     2761 aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg
     2821 actggaaagc gggcagtgag cgcaacgcaa ttaatgtaag ttagctcact cattaggcac
     2881 cgggatctcg accgatgccc ttgagagcct tcaacccagt cagctccttc cggtgggcgc
     2941 ggggcatgac tatcgtcgcc gcacttatga ctgtcttctt tatcatgcaa ctcgtaggac
     3001 aggtgccggc agcgctctgg gtcattttcg gcgaggaccg ctttcgctgg agcgcgacga
     3061 tgatcggcct gtcgcttgcg gtattcggaa tcttgcacgc cctcgctcaa gccttcgtca
     3121 ctggtcccgc caccaaacgt ttcggcgaga agcaggccat tatcgccggc atggcggccc
     3181 cacgggtgcg catgatcgtg ctcctgtcgt tgaggacccg gctaggctgg cggggttgcc
     3241 ttactggtta gcagaatgaa tcaccgatac gcgagcgaac gtgaagcgac tgctgctgca
     3301 aaacgtctgc gacctgagca acaacatgaa tggtcttcgg tttccgtgtt tcgtaaagtc
     3361 tggaaacgcg gaagtcagcg ccctgcacca ttatgttccg gatctgcatc gcaggatgct
     3421 gctggctacc ctgtggaaca cctacatctg tattaacgaa gcgctggcat tgaccctgag
     3481 tgatttttct ctggtcccgc cgcatccata ccgccagttg tttaccctca caacgttcca
     3541 gtaaccgggc atgttcatca tcagtaaccc gtatcgtgag catcctctct cgtttcatcg
     3601 gtatcattac ccccatgaac agaaatcccc cttacacgga ggcatcagtg accaaacagg
     3661 aaaaaaccgc ccttaacatg gcccgcttta tcagaagcca gacattaacg cttctggaga
     3721 aactcaacga gctggacgcg gatgaacagg cagacatctg tgaatcgctt cacgaccacg
     3781 ctgatgagct ttaccgcagc tgcctcgcgc gtttcggtga tgacggtgaa aacctctgac
     3841 acatgcagct cccggagacg gtcacagctt gtctgtaagc ggatgccggg agcagacaag
     3901 cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg cgcagccatg acccagtcac
     3961 gtagcgatag cggagtgtat actggcttaa ctatgcggca tcagagcaga ttgtactgag
     4021 agtgcaccat tgcggtgtga aataccgcac agatgcgtaa ggagaaaata ccgcatcagg
     4081 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
     4141 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     4201 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     4261 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     4321 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     4381 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     4441 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     4501 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     4561 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     4621 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     4681 tggcctaact acggctacac tagaaggaca gtatttggta tctgcgctct gctgaagcca
     4741 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     4801 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     4861 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     4921 ttggtcatga acaataaaac tgtctgctta cataaacagt aatacaaggg gtgttatgag
     4981 ccatattcaa cgggaaacgt cttgctctag gccgcgatta aattccaaca tggatgctga
     5041 tttatatggg tataaatggg ctcgcgataa tgtcgggcaa tcaggtgcga caatctatcg
     5101 attgtatggg aagcccgatg cgccagagtt gtttctgaaa catggcaaag gtagcgttgc
     5161 caatgatgtt acagatgaga tggtcagact aaactggctg acggaattta tgcctcttcc
     5221 gaccatcaag cattttatcc gtactcctga tgatgcatgg ttactcacca ctgcgatccc
     5281 cgggaaaaca gcattccagg tattagaaga atatcctgat tcaggtgaaa atattgttga
     5341 tgcgctggca gtgttcctgc gccggttgca ttcgattcct gtttgtaatt gtccttttaa
     5401 cagcgatcgc gtatttcgtc tcgctcaggc gcaatcacga atgaataacg gtttggttga
     5461 tgcgagtgat tttgatgacg agcgtaatgg ctggcctgtt gaacaagtct ggaaagaaat
     5521 gcataaactt ttgccattct caccggattc agtcgtcact catggtgatt tctcacttga
     5581 taaccttatt tttgacgagg ggaaattaat aggttgtatt gatgttggac gagtcggaat
     5641 cgcagaccga taccaggatc ttgccatcct atggaactgc ctcggtgagt tttctccttc
     5701 attacagaaa cggctttttc aaaaatatgg tattgataat cctgatatga ataaattgca
     5761 gtttcatttg atgctcgatg agtttttcta agaattaatt catgagcgga tacatatttg
     5821 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
     5881 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
     5941 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
     6001 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
     6061 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
     6121 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
     6181 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
     6241 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
     6301 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgcca
//
