LOCUS       sequence-477128-        6237 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of A. thaliana CK2alpha1 
            76-409_K318A_H347A_K350A_K148/149A_K146A_HC.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6237)
  TITLE     Hothorn CK2 plasmids
REFERENCE   2  (bases 1 to 6237)
  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: 
            pMH_AtCK2alpha1_76-409_K318A_H347A_K350A_K148/149A_K146A_HC
FEATURES             Location/Qualifiers
     source          1..6237
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(1..589)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(81..100)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(700..719)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             711..1526
                     /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             711..1526
                     /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(784..803)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(1618..2073)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1755..1776)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1967..1986
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(2110..2157)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     2153..2171
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(2224..2241)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             3262..3284
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    3299..3323
                     /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(3323..3342)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3324..3342)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3426..3445)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        3651..3728
                     /gene="lacI"
                     /label=lacI promoter
     CDS             3729..4811
                     /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(3748..3767)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             5620..5811
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(5805..5827)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    5913..6052
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     5968..5987
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(6067..6084)
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
       61 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
      121 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
      181 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
      241 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
      301 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
      361 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
      421 taactacggc tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac
      481 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
      541 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
      601 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
      661 catgaacaat aaaactgtct gcttacataa acagtaatac aaggggtgtt atgagccata
      721 ttcaacggga aacgtcttgc tctaggccgc gattaaattc caacatggat gctgatttat
      781 atgggtataa atgggctcgc gataatgtcg ggcaatcagg tgcgacaatc tatcgattgt
      841 atgggaagcc cgatgcgcca gagttgtttc tgaaacatgg caaaggtagc gttgccaatg
      901 atgttacaga tgagatggtc agactaaact ggctgacgga atttatgcct cttccgacca
      961 tcaagcattt tatccgtact cctgatgatg catggttact caccactgcg atccccggga
     1021 aaacagcatt ccaggtatta gaagaatatc ctgattcagg tgaaaatatt gttgatgcgc
     1081 tggcagtgtt cctgcgccgg ttgcattcga ttcctgtttg taattgtcct tttaacagcg
     1141 atcgcgtatt tcgtctcgct caggcgcaat cacgaatgaa taacggtttg gttgatgcga
     1201 gtgattttga tgacgagcgt aatggctggc ctgttgaaca agtctggaaa gaaatgcata
     1261 aacttttgcc attctcaccg gattcagtcg tcactcatgg tgatttctca cttgataacc
     1321 ttatttttga cgaggggaaa ttaataggtt gtattgatgt tggacgagtc ggaatcgcag
     1381 accgatacca ggatcttgcc atcctatgga actgcctcgg tgagttttct ccttcattac
     1441 agaaacggct ttttcaaaaa tatggtattg ataatcctga tatgaataaa ttgcagtttc
     1501 atttgatgct cgatgagttt ttctaagaat taattcatga gcggatacat atttgaatgt
     1561 atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctaaa
     1621 ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt
     1681 ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag accgagatag
     1741 ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg
     1801 tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcaccctaat
     1861 caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc
     1921 gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga
     1981 aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac
     2041 ccgccgcgct taatgcgccg ctacagggcg cgtcccattc gccaatccgg atatagttcc
     2101 tcctttcagc aaaaaacccc tcaagacccg tttagaggcc ccaaggggtt atgctagtta
     2161 ttgctcagcg gtggcagcag ccaactcagc ttcctttcgg gctttgttag cagccggatc
     2221 tcagtggtgg tggtggtggt gctcgagtgc ggccgcttga cttctcattc tgctggtttc
     2281 tgctgcccta acttgggcga agtaagcgtg cgccatggct tcttttgcag ttaatctgtc
     2341 ttgatgatca taccgaagta gcttgtcgag gaaatcaatc gcctcgggtg aaactaagtg
     2401 ctgattgtct gcgttgatga atttggacca aggtgccctg ctagcccgcc caacaagtgc
     2461 ttccagttgt ggatcaagtt ctaactggta cttgttcaga taagcattca attcgtcagt
     2521 cccaagcacc gcggcgattt tcactagctg atcctggttg tcatggccgt agaagaaagg
     2581 ttccttacgg aatatcatcc ctgcaaacat acagccaaga ctccacatat ccaatgaata
     2641 gtcatagtct tgcaagtcca caagaagttc aggtcccttg aaatatcttg aggcaacacg
     2701 gacattatac tcttttccag ggtggtaaaa ctcagcaaga ccccaatcta taagacgaag
     2761 cttacgcagc tcatggtcaa tcataacatt atgaggcttg acgtctctgt gcattattcc
     2821 ttgtgaatgg cagaagtcta gtgccttcaa cagctcatag atgtagtaac gaatgtcgta
     2881 atcagtcaac gtaggataca aaactttaaa gtctgtgctg ttaacatatt caaatatcaa
     2941 gcttggagtt ttcgaatgtt gatctctaac aacatcaagc agcttgacaa tatttggccc
     3001 accgcatagg ttctgaagta ttttaatctc tcttcttatc gccgctttcg caacaggttt
     3061 gaggattttg ataatgcact tctccttgct gttcacattt ataccctcga aaacttcact
     3121 atattttcca cgccctacct tccttactac ctcataatca tcttgctccc cccattgaac
     3181 aatgagtgac tcgtaatccc aataatcctt gggtcgaatc acattgactt cggtgtacac
     3241 acgagctttc gacatggtat atctccttct taaagttaaa caaaattatt tctagagggg
     3301 aattgttatc cgctcacaat tcccctatag tgagtcgtat taatttcgcg ggatcgagat
     3361 ctcgatcctc tacgccggac gcatcgtggc cggcatcacc ggcgccacag gtgcggttgc
     3421 tggcgcctat atcgccgaca tcaccgatgg ggaagatcgg gctcgccact tcgggctcat
     3481 gagcgcttgt ttcggcgtgg gtatggtggc aggccccgtg gccgggggac tgttgggcgc
     3541 catctccttg catgcaccat tccttgcggc ggcggtgctc aacggcctca acctactact
     3601 gggctgcttc ctaatgcagg agtcgcataa gggagagcgt cgagatcccg gacaccatcg
     3661 aatggcgcaa aacctttcgc ggtatggcat gatagcgccc ggaagagagt caattcaggg
     3721 tggtgaatgt gaaaccagta acgttatacg atgtcgcaga gtatgccggt gtctcttatc
     3781 agaccgtttc ccgcgtggtg aaccaggcca gccacgtttc tgcgaaaacg cgggaaaaag
     3841 tggaagcggc gatggcggag ctgaattaca ttcccaaccg cgtggcacaa caactggcgg
     3901 gcaaacagtc gttgctgatt ggcgttgcca cctccagtct ggccctgcac gcgccgtcgc
     3961 aaattgtcgc ggcgattaaa tctcgcgccg atcaactggg tgccagcgtg gtggtgtcga
     4021 tggtagaacg aagcggcgtc gaagcctgta aagcggcggt gcacaatctt ctcgcgcaac
     4081 gcgtcagtgg gctgatcatt aactatccgc tggatgacca ggatgccatt gctgtggaag
     4141 ctgcctgcac taatgttccg gcgttatttc ttgatgtctc tgaccagaca cccatcaaca
     4201 gtattatttt ctcccatgaa gacggtacgc gactgggcgt ggagcatctg gtcgcattgg
     4261 gtcaccagca aatcgcgctg ttagcgggcc cattaagttc tgtctcggcg cgtctgcgtc
     4321 tggctggctg gcataaatat ctcactcgca atcaaattca gccgatagcg gaacgggaag
     4381 gcgactggag tgccatgtcc ggttttcaac aaaccatgca aatgctgaat gagggcatcg
     4441 ttcccactgc gatgctggtt gccaacgatc agatggcgct gggcgcaatg cgcgccatta
     4501 ccgagtccgg gctgcgcgtt ggtgcggata tctcggtagt gggatacgac gataccgaag
     4561 acagctcatg ttatatcccg ccgttaacca ccatcaaaca ggattttcgc ctgctggggc
     4621 aaaccagcgt ggaccgcttg ctgcaactct ctcagggcca ggcggtgaag ggcaatcagc
     4681 tgttgcccgt ctcactggtg aaaagaaaaa ccaccctggc gcccaatacg caaaccgcct
     4741 ctccccgcgc gttggccgat tcattaatgc agctggcacg acaggtttcc cgactggaaa
     4801 gcgggcagtg agcgcaacgc aattaatgta agttagctca ctcattaggc accgggatct
     4861 cgaccgatgc ccttgagagc cttcaaccca gtcagctcct tccggtgggc gcggggcatg
     4921 actatcgtcg ccgcacttat gactgtcttc tttatcatgc aactcgtagg acaggtgccg
     4981 gcagcgctct gggtcatttt cggcgaggac cgctttcgct ggagcgcgac gatgatcggc
     5041 ctgtcgcttg cggtattcgg aatcttgcac gccctcgctc aagccttcgt cactggtccc
     5101 gccaccaaac gtttcggcga gaagcaggcc attatcgccg gcatggcggc cccacgggtg
     5161 cgcatgatcg tgctcctgtc gttgaggacc cggctaggct ggcggggttg ccttactggt
     5221 tagcagaatg aatcaccgat acgcgagcga acgtgaagcg actgctgctg caaaacgtct
     5281 gcgacctgag caacaacatg aatggtcttc ggtttccgtg tttcgtaaag tctggaaacg
     5341 cggaagtcag cgccctgcac cattatgttc cggatctgca tcgcaggatg ctgctggcta
     5401 ccctgtggaa cacctacatc tgtattaacg aagcgctggc attgaccctg agtgattttt
     5461 ctctggtccc gccgcatcca taccgccagt tgtttaccct cacaacgttc cagtaaccgg
     5521 gcatgttcat catcagtaac ccgtatcgtg agcatcctct ctcgtttcat cggtatcatt
     5581 acccccatga acagaaatcc cccttacacg gaggcatcag tgaccaaaca ggaaaaaacc
     5641 gcccttaaca tggcccgctt tatcagaagc cagacattaa cgcttctgga gaaactcaac
     5701 gagctggacg cggatgaaca ggcagacatc tgtgaatcgc ttcacgacca cgctgatgag
     5761 ctttaccgca gctgcctcgc gcgtttcggt gatgacggtg aaaacctctg acacatgcag
     5821 ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag
     5881 ggcgcgtcag cgggtgttgg cgggtgtcgg ggcgcagcca tgacccagtc acgtagcgat
     5941 agcggagtgt atactggctt aactatgcgg catcagagca gattgtactg agagtgcacc
     6001 attgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggcgctcttc
     6061 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     6121 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     6181 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgtt
//
