LOCUS       sequence-174922-        6221 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  E. coli expression of MKK3 aa 42-347.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6221)
  TITLE     Mace Lab plasmids
REFERENCE   2  (bases 1 to 6221)
  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"
COMMENT     Sequence Label: His 3C MKK3 42-347
FEATURES             Location/Qualifiers
     source          1..6221
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             7..24
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             34..57
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             999..1016
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1069..1087)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1083..1130
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1167..1622
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1254..1273)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1464..1485
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1714..2529)
                     /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             complement(1714..2529)
                     /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     2437..2456
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2521..2540
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2651..3239
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3140..3159
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3393..3410
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3425..3564
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3490..3509)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3650..3672
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3666..3857)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4666..5748)
                     /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     5710..5729
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5749..5826)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6032..6051
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     6135..6154
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6135..6153
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    6154..6178
                     /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)."
     RBS             6193..6215
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
ORIGIN
        1 atggcacatc accaccacca tcactccgcg gctcttgagg tgctctttca gggacccggt
       61 cggaacctgg actcccggac cttcatcacc attggagaca gaaactttga ggtggaggct
      121 gatgacttgg tgaccatctc agaactgggc cgtggagcct atggggtggt agagaaggtg
      181 cggcacgccc agagcggcac catcatggcc gtgaagcgga tccgggccac cgtgaactca
      241 caggagcaga agcggctgct catggacctg gacatcaaca tgcgcacggt cgactgcttc
      301 tacactgtca ccttctacgg ggcactattc agagagggag acgtgtggat ctgcatggag
      361 ctcatggaca catccttgga caagttctac cggaaggtgc tggataaaaa catgacaatt
      421 ccagaggaca tccttgggga gattgctgtg tctatcgtgc gggccctgga gcatctgcac
      481 agcaagctgt cggtgatcca cagagatgtg aagccctcca atgtccttat caacaaggag
      541 ggccatgtga agatgtgtga ctttggcatc agtggctact tggtggactc tgtggccaag
      601 acgatggatg ccggctgcaa gccctacatg gcccctgaga ggatcaaccc agagctgaac
      661 cagaagggct acaatgtcaa gtccgacgtc tggagcctgg gcatcaccat gattgagatg
      721 gccatcctgc ggttccctta cgagtcctgg gggaccccgt tccagcagct gaagcaggtg
      781 gtggaggagc cgtcccccca gctcccagcc gaccgtttct cccccgagtt tgtggacttc
      841 actgctcagt gcctgaggaa gaaccccgca gagcgtatga gctacctgga gctgatggag
      901 caccccttct tcaccttgca caaaaccaag aagacggaca ttgctgcctt cgtgaaggag
      961 atcctgggag aagactcata accgggcttc tcctcgagca ccaccaccac caccactgag
     1021 atccggctgc taacaaagcc cgaaaggaag ctgagttggc tgctgccacc gctgagcaat
     1081 aactagcata accccttggg gcctctaaac gggtcttgag gggttttttg ctgaaaggag
     1141 gaactatatc cggattggcg aatgggacgc gccctgtagc ggcgcattaa gcgcggcggg
     1201 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt
     1261 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     1321 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     1381 ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
     1441 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc
     1501 tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa
     1561 aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa cgcttacaat
     1621 ttaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     1681 cattcaaata tgtatccgct catgaattaa ttcttagaaa aactcatcga gcatcaaatg
     1741 aaactgcaat ttattcatat caggattatc aataccatat ttttgaaaaa gccgtttctg
     1801 taatgaagga gaaaactcac cgaggcagtt ccataggatg gcaagatcct ggtatcggtc
     1861 tgcgattccg actcgtccaa catcaataca acctattaat ttcccctcgt caaaaataag
     1921 gttatcaagt gagaaatcac catgagtgac gactgaatcc ggtgagaatg gcaaaagttt
     1981 atgcatttct ttccagactt gttcaacagg ccagccatta cgctcgtcat caaaatcact
     2041 cgcatcaacc aaaccgttat tcattcgtga ttgcgcctga gcgagacgaa atacgcgatc
     2101 gctgttaaaa ggacaattac aaacaggaat cgaatgcaac cggcgcagga acactgccag
     2161 cgcatcaaca atattttcac ctgaatcagg atattcttct aatacctgga atgctgtttt
     2221 cccggggatc gcagtggtga gtaaccatgc atcatcagga gtacggataa aatgcttgat
     2281 ggtcggaaga ggcataaatt ccgtcagcca gtttagtctg accatctcat ctgtaacatc
     2341 attggcaacg ctacctttgc catgtttcag aaacaactct ggcgcatcgg gcttcccata
     2401 caatcgatag attgtcgcac ctgattgccc gacattatcg cgagcccatt tatacccata
     2461 taaatcagca tccatgttgg aatttaatcg cggcctagag caagacgttt cccgttgaat
     2521 atggctcata acaccccttg tattactgtt tatgtaagca gacagtttta ttgttcatga
     2581 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     2641 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     2701 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     2761 taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag
     2821 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     2881 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     2941 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     3001 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     3061 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     3121 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     3181 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     3241 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
     3301 tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg
     3361 ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag
     3421 agcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcaatggtg
     3481 cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatacac tccgctatcg
     3541 ctacgtgact gggtcatggc tgcgccccga cacccgccaa cacccgctga cgcgccctga
     3601 cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc
     3661 atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga ggcagctgcg gtaaagctca
     3721 tcagcgtggt cgtgaagcga ttcacagatg tctgcctgtt catccgcgtc cagctcgttg
     3781 agtttctcca gaagcgttaa tgtctggctt ctgataaagc gggccatgtt aagggcggtt
     3841 ttttcctgtt tggtcactga tgcctccgtg taagggggat ttctgttcat gggggtaatg
     3901 ataccgatga aacgagagag gatgctcacg atacgggtta ctgatgatga acatgcccgg
     3961 ttactggaac gttgtgaggg taaacaactg gcggtatgga tgcggcggga ccagagaaaa
     4021 atcactcagg gtcaatgcca gcgcttcgtt aatacagatg taggtgttcc acagggtagc
     4081 cagcagcatc ctgcgatgca gatccggaac ataatggtgc agggcgctga cttccgcgtt
     4141 tccagacttt acgaaacacg gaaaccgaag accattcatg ttgttgctca ggtcgcagac
     4201 gttttgcagc agcagtcgct tcacgttcgc tcgcgtatcg gtgattcatt ctgctaacca
     4261 gtaaggcaac cccgccagcc tagccgggtc ctcaacgaca ggagcacgat catgcgcacc
     4321 cgtggggccg ccatgccggc gataatggcc tgcttctcgc cgaaacgttt ggtggcggga
     4381 ccagtgacga aggcttgagc gagggcgtgc aagattccga ataccgcaag cgacaggccg
     4441 atcatcgtcg cgctccagcg aaagcggtcc tcgccgaaaa tgacccagag cgctgccggc
     4501 acctgtccta cgagttgcat gataaagaag acagtcataa gtgcggcgac gatagtcatg
     4561 ccccgcgccc accggaagga gctgactggg ttgaaggctc tcaagggcat cggtcgagat
     4621 cccggtgcct aatgagtgag ctaacttaca ttaattgcgt tgcgctcact gcccgctttc
     4681 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
     4741 ggtttgcgta ttgggcgcca gggtggtttt tcttttcacc agtgagacgg gcaacagctg
     4801 attgcccttc accgcctggc cctgagagag ttgcagcaag cggtccacgc tggtttgccc
     4861 cagcaggcga aaatcctgtt tgatggtggt taacggcggg atataacatg agctgtcttc
     4921 ggtatcgtcg tatcccacta ccgagatatc cgcaccaacg cgcagcccgg actcggtaat
     4981 ggcgcgcatt gcgcccagcg ccatctgatc gttggcaacc agcatcgcag tgggaacgat
     5041 gccctcattc agcatttgca tggtttgttg aaaaccggac atggcactcc agtcgccttc
     5101 ccgttccgct atcggctgaa tttgattgcg agtgagatat ttatgccagc cagccagacg
     5161 cagacgcgcc gagacagaac ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa
     5221 tgcgaccaga tgctccacgc ccagtcgcgt accgtcttca tgggagaaaa taatactgtt
     5281 gatgggtgtc tggtcagaga catcaagaaa taacgccgga acattagtgc aggcagcttc
     5341 cacagcaatg gcatcctggt catccagcgg atagttaatg atcagcccac tgacgcgttg
     5401 cgcgagaaga ttgtgcaccg ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga
     5461 caccaccacg ctggcaccca gttgatcggc gcgagattta atcgccgcga caatttgcga
     5521 cggcgcgtgc agggccagac tggaggtggc aacgccaatc agcaacgact gtttgcccgc
     5581 cagttgttgt gccacgcggt tgggaatgta attcagctcc gccatcgccg cttccacttt
     5641 ttcccgcgtt ttcgcagaaa cgtggctggc ctggttcacc acgcgggaaa cggtctgata
     5701 agagacaccg gcatactctg cgacatcgta taacgttact ggtttcacat tcaccaccct
     5761 gaattgactc tcttccgggc gctatcatgc cataccgcga aaggttttgc gccattcgat
     5821 ggtgtccggg atctcgacgc tctcccttat gcgactcctg cattaggaag cagcccagta
     5881 gtaggttgag gccgttgagc accgccgccg caaggaatgg tgcatgcaag gagatggcgc
     5941 ccaacagtcc cccggccacg gggcctgcca ccatacccac gccgaaacaa gcgctcatga
     6001 gcccgaagtg gcgagcccga tcttccccat cggtgatgtc ggcgatatag gcgccagcaa
     6061 ccgcacctgt ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg atcgagatct
     6121 cgatcccgcg aaattaatac gactcactat aggggaattg tgagcggata acaattcccc
     6181 tctagaaata attttgttta actttaagaa ggagatatac c
//
