LOCUS       sequence-327672-        6507 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses human KANK1 Ank domain in bacterial cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6507)
  TITLE     Bacterial expression plasmids for production of recombinant KANK 
            proteins
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6507)
  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: 
            "Unpublished"
COMMENT     Sequence Label: human KANK1 Ankyrin Repeats pET151
FEATURES             Location/Qualifiers
     source          1..6507
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     106..125
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     209..228
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        209..227
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    228..252
                     /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             267..289
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             300..317
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             318..359
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             360..380
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     primer_bind     complement(1208..1226)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1222..1269
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(1507..1525)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1593..1697
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1698..1766,1767..2558)
                     /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(1916..1935)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2729..3317
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3218..3237
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3471..3488
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3503..3642
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3568..3587)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3728..3750
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3744..3935)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5247..6329)
                     /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     6291..6310
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6330..6407)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 caaggagatg gcgcccaaca gtcccccggc cacggggcct gccaccatac ccacgccgaa
       61 acaagcgctc atgagcccga agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat
      121 ataggcgcca gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta
      181 gaggatcgag atctcgatcc cgcgaaatta atacgactca ctatagggga attgtgagcg
      241 gataacaatt cccctctaga aataattttg tttaacttta agaaggagat atacatatgc
      301 atcatcacca tcaccatggt aagcctatcc ctaaccctct cctcggtctc gattctacgg
      361 aaaacctgta ttttcaggga ggatccgaaa tcagagagag gtatgaatta agtgaaaaga
      421 tgttgtctgc atgcaactta ctgaaaaata ctataaatga ccccaaagct ttgaccagca
      481 aagatatgag gttctgtctg aacaccctcc agcacgagtg gttccgcgtg tccagtcaga
      541 agtcagccat tccagccatg gtgggggact acatagctgc ttttgaggcc atttccccag
      601 atgtcctccg ctatgtcatc aacttggcag acggcaacgg caacacagcc ctccattaca
      661 gcgtgtccca ctccaacttc gagattgtga agctgctgtt agatgccgat gtgtgtaatg
      721 tggatcacca gaacaaggca ggctacaccc ccatcatgtt ggcggccctc gccgctgtgg
      781 aagcagagaa ggacatgcgg attgtggaag aactctttgg ctgtggggat gtgaatgcca
      841 aagctagtca ggcgggacag acggccctca tgctggcggt cagtcacgga cggatagaca
      901 tggtgaaggg ccttctggcc tgtggggctg atgtcaacat ccaggatgac gagggctcca
      961 cggccctcat gtgtgccagc gagcacgggc acgtggagat tgtcaagctg ctgctggccc
     1021 agcccggctg caacggtcac ctagaggaca acgatggcag cactgcgctc tcaatcgccc
     1081 tggaagcagg acacaaggac atcgctgttc ttctgtatgc ccatgtcaac tttgcaaaat
     1141 gactcgagta agagctcaga tccggctgct aacaaagccc gaaaggaagc tgagttggct
     1201 gctgccaccg ctgagcaata actagcataa ccccttgggg cctctaaacg ggtcttgagg
     1261 ggttttttgc tgaaaggagg aactatatcc ggatatcccg caagaggccc ggcagtaccg
     1321 gcataaccaa gcctatgcct acagcatcca gggtgacggt gccgaggatg acgatgagcg
     1381 cattgttaga tttcatacac ggtgcctgac tgcgttagca atttaactgt gataaactac
     1441 cgcattaaag ctagcttatc gatgataagc tgtcaaacat gagaattaat tcttgaagac
     1501 gaaagggcct cgtgatacgc ctatttttat aggttaatgt catgataata atggtttctt
     1561 agacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct
     1621 aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat
     1681 attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg
     1741 cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg
     1801 aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc
     1861 ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat
     1921 gtggcgcggt attatcccgt gttgacgccg ggcaagagca actcggtcgc cgcatacact
     1981 attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca
     2041 tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact
     2101 tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg
     2161 atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg
     2221 agcgtgacac cacgatgcct gcagcaatgg caacaacgtt gcgcaaacta ttaactggcg
     2281 aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg
     2341 caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag
     2401 ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc
     2461 gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga
     2521 tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat
     2581 atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc
     2641 tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag
     2701 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
     2761 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
     2821 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgtccttc
     2881 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg
     2941 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
     3001 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
     3061 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
     3121 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
     3181 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
     3241 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
     3301 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
     3361 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta
     3421 ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag
     3481 tgagcgagga agcggaagag cgcctgatgc ggtattttct ccttacgcat ctgtgcggta
     3541 tttcacaccg caatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca
     3601 gtatacactc cgctatcgct acgtgactgg gtcatggctg cgccccgaca cccgccaaca
     3661 cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg
     3721 accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgagg
     3781 cagctgcggt aaagctcatc agcgtggtcg tgaagcgatt cacagatgtc tgcctgttca
     3841 tccgcgtcca gctcgttgag tttctccaga agcgttaatg tctggcttct gataaagcgg
     3901 gccatgttaa gggcggtttt ttcctgtttg gtcactgatg cctccgtgta agggggattt
     3961 ctgttcatgg gggtaatgat accgatgaaa cgagagagga tgctcacgat acgggttact
     4021 gatgatgaac atgcccggtt actggaacgt tgtgagggta aacaactggc ggtatggatg
     4081 cggcgggacc agagaaaaat cactcagggt caatgccagc gcttcgttaa tacagatgta
     4141 ggtgttccac agggtagcca gcagcatcct gcgatgcaga tccggaacat aatggtgcag
     4201 ggcgctgact tccgcgtttc cagactttac gaaacacgga aaccgaagac cattcatgtt
     4261 gttgctcagg tcgcagacgt tttgcagcag cagtcgcttc acgttcgctc gcgtatcggt
     4321 gattcattct gctaaccagt aaggcaaccc cgccagccta gccgggtcct caacgacagg
     4381 agcacgatca tgcgcacccg tggccaggac ccaacgctgc ccgagatgcg ccgcgtgcgg
     4441 ctgctggaga tggcggacgc gatggatatg ttctgccaag ggttggtttg cgcattcaca
     4501 gttctccgca agaattgatt ggctccaatt cttggagtgg tgaatccgtt agcgaggtgc
     4561 cgccggcttc cattcaggtc gaggtggccc ggctccatgc accgcgacgc aacgcgggga
     4621 ggcagacaag gtatagggcg gcgcctacaa tccatgccaa cccgttccat gtgctcgccg
     4681 aggcggcata aatcgccgtg acgatcagcg gtccaatgat cgaagttagg ctggtaagag
     4741 ccgcgagcga tccttgaagc tgtccctgat ggtcgtcatc tacctgcctg gacagcatgg
     4801 cctgcaacgc gggcatcccg atgccgccgg aagcgagaag aatcataatg gggaaggcca
     4861 tccagcctcg cgtcgcgaac gccagcaaga cgtagcccag cgcgtcggcc gccatgccgg
     4921 cgataatggc ctgcttctcg ccgaaacgtt tggtggcggg accagtgacg aaggcttgag
     4981 cgagggcgtg caagattccg aataccgcaa gcgacaggcc gatcatcgtc gcgctccagc
     5041 gaaagcggtc ctcgccgaaa atgacccaga gcgctgccgg cacctgtcct acgagttgca
     5101 tgataaagaa gacagtcata agtgcggcga cgatagtcat gccccgcgcc caccggaagg
     5161 agctgactgg gttgaaggct ctcaagggca tcggtcgaga tcccggtgcc taatgagtga
     5221 gctaacttac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     5281 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgcc
     5341 agggtggttt ttcttttcac cagtgagacg ggcaacagct gattgccctt caccgcctgg
     5401 ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc ccagcaggcg aaaatcctgt
     5461 ttgatggtgg ttaacggcgg gatataacat gagctgtctt cggtatcgtc gtatcccact
     5521 accgagatat ccgcaccaac gcgcagcccg gactcggtaa tggcgcgcat tgcgcccagc
     5581 gccatctgat cgttggcaac cagcatcgca gtgggaacga tgccctcatt cagcatttgc
     5641 atggtttgtt gaaaaccgga catggcactc cagtcgcctt cccgttccgc tatcggctga
     5701 atttgattgc gagtgagata tttatgccag ccagccagac gcagacgcgc cgagacagaa
     5761 cttaatgggc ccgctaacag cgcgatttgc tggtgaccca atgcgaccag atgctccacg
     5821 cccagtcgcg taccgtcttc atgggagaaa ataatactgt tgatgggtgt ctggtcagag
     5881 acatcaagaa ataacgccgg aacattagtg caggcagctt ccacagcaat ggcatcctgg
     5941 tcatccagcg gatagttaat gatcagccca ctgacgcgtt gcgcgagaag attgtgcacc
     6001 gccgctttac aggcttcgac gccgcttcgt tctaccatcg acaccaccac gctggcaccc
     6061 agttgatcgg cgcgagattt aatcgccgcg acaatttgcg acggcgcgtg cagggccaga
     6121 ctggaggtgg caacgccaat cagcaacgac tgtttgcccg ccagttgttg tgccacgcgg
     6181 ttgggaatgt aattcagctc cgccatcgcc gcttccactt tttcccgcgt tttcgcagaa
     6241 acgtggctgg cctggttcac cacgcgggaa acggtctgat aagagacacc ggcatactct
     6301 gcgacatcgt ataacgttac tggtttcaca ttcaccaccc tgaattgact ctcttccggg
     6361 cgctatcatg ccataccgcg aaaggttttg cgccattcga tggtgtccgg gatctcgacg
     6421 ctctccctta tgcgactcct gcattaggaa gcagcccagt agtaggttga ggccgttgag
     6481 caccgccgcc gcaaggaatg gtgcatg
//
