LOCUS       sequence-271807-        5537 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of his-CENP-C 700-943.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5537)
  AUTHORS   McKinley KL, Cheeseman IM
  TITLE     Polo-like kinase 1 licenses CENP-A deposition at centromeres.
  JOURNAL   Cell. 2014 Jul 17;158(2):397-411. doi: 10.1016/j.cell.2014.06.016.
  PUBMED    25036634
REFERENCE   2  (bases 1 to 5537)
  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"; doi: 
            "10.1016/j.cell.2014.06"; journalName: "Cell"; date: "2014-07-17- 
            17"; volume: "158"; issue: "2"; pages: "397-411"
COMMENT     Sequence Label: pKM136
FEATURES             Location/Qualifiers
     source          1..5537
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     65..84
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        65..83
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             157..179
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             199..216
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             220..252
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             join(256..264,265..279)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             280..303
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(1131..1149)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1145..1192
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(1558..1563,1564..1580,1581..1586))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(1613..1631)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1699..1803
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1804..1872,1873..2664)
                     /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(2022..2041)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2835..3423
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3324..3343
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3577..3594
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3609..3748
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3674..3693)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3834..3856
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3850..4041)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     5499..5518
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg atcgagatct cgatcccgcg
       61 aaattaatac gactcactat agggagacca caacggtttc cctctagtgc cggctccgga
      121 gagctccaat tggaattcgc tagctctaga aataattttg tttaacttta agaaggagat
      181 atacgtatgc ggggttctca tcatcatcat catcatggta tggctagcat gactggtgga
      241 cagcaaatgg gtcgggatct gtacgacgat gacgataagc tggaagttct gttccagggg
      301 ccccatatgg atgtggacga tgaggaagtt cacggtagta gcgacgattc aaaacagagt
      361 aaagtcatcc ctaaaaatcg tattcaccat aaactggtgc tgccgagtaa tactccgaac
      421 gttcgtcgta caaaacgtac acgcctgaaa cctctggaat attggcgtgg tgaacgtatc
      481 gactaccagg gtcgtccatc tggtggtttt gtaatcagcg gtgtgctgag tcctgatacg
      541 attagcagca aacgtaaagc caaagagaac atcggcaaag tcaacaaaaa atccaacaaa
      601 aaacgcatct gcctggataa cgatgagcgt aaaacgaatc tgatggtgaa tctgggtatt
      661 cctctgggag atcctctgca gcctacccgt gtaaaagatc ctgagacacg tgagattatc
      721 ctgatggacc tggttcgtcc tcaagacacc tatcagtttt tcgtcaaaca cggcgagctg
      781 aaagtctata aaaccctgga caccccgttc tttagtaccg gcaaactgat tctgggtcct
      841 caggaggaaa aaggcaaaca gcacgtgggt caagacatcc tggtctttta tgtgaatttt
      901 ggggacctgc tgtgtactct gcatgaaacc ccgtatattc tgtctaccgg tgacagcttc
      961 tatgtgccgt ctggaaacta ttataacatc aaaaacctgc gcaacgaaga aagtgttctg
     1021 ctgttcacac agatcaaacg ctgaggatcc agatctaagc ttggtaccac gcgtgcgcgc
     1081 tgatccggct gctaacaaag cccgaaagga agctgagttg gctgctgcca ccgctgagca
     1141 ataactagca taaccccttg gggcctctaa acgggtcttg aggggttttt tgctgaaagg
     1201 aggaactata tccggatatc cacaggacgg gtgtggtcgc catgatcgcg tagtcgatag
     1261 tggctccaag tagcgaagcg agcaggactg ggcggcggcc aaagcggtcg gacagtgctc
     1321 cgagaacggg tgcgcataga aattgcatca acgcatatag cgctagcagc acgccatagt
     1381 gactggcgat gctgtcggaa tggacgatat cccgcaagag gcccggcagt accggcataa
     1441 ccaagcctat gcctacagca tccagggtga cggtgccgag gatgacgatg agcgcattgt
     1501 tagatttcat acacggtgcc tgactgcgtt agcaatttaa ctgtgataaa ctaccgcatt
     1561 aaagcttatc gatgataagc tgtcaaacat gagaattctt gaagacgaaa gggcctcgtg
     1621 atacgcctat ttttataggt taatgtcatg ataataatgg tttcttagac gtcaggtggc
     1681 acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat
     1741 atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag
     1801 agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt
     1861 cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt
     1921 gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga gagttttcgc
     1981 cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta
     2041 tcccgtgttg acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac
     2101 ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa
     2161 ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg
     2221 atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc
     2281 cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg
     2341 atgcctgcag caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta
     2401 gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg accacttctg
     2461 cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg
     2521 tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc
     2581 tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt
     2641 gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt
     2701 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
     2761 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     2821 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     2881 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     2941 aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag
     3001 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     3061 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     3121 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     3181 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     3241 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     3301 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     3361 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     3421 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     3481 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
     3541 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg
     3601 gaagagcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcaat
     3661 ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagtat acactccgct
     3721 atcgctacgt gactgggtca tggctgcgcc ccgacacccg ccaacacccg ctgacgcgcc
     3781 ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag
     3841 ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgaggcagc tgcggtaaag
     3901 ctcatcagcg tggtcgtgaa gcgattcaca gatgtctgcc tgttcatccg cgtccagctc
     3961 gttgagtttc tccagaagcg ttaatgtctg gcttctgata aagcgggcca tgttaagggc
     4021 ggttttttcc tgtttggtca ctgatgcctc cgtgtaaggg ggatttctgt tcatgggggt
     4081 aatgataccg atgaaacgag agaggatgct cacgatacgg gttactgatg atgaacatgc
     4141 ccggttactg gaacgttgtg agggtaaaca actggcggta tggatgcggc gggaccagag
     4201 aaaaatcact cagggtcaat gccagcgctt cgttaataca gatgtaggtg ttccacaggg
     4261 tagccagcag catcctgcga tgcagatccg gaacataatg gtgcagggcg ctgacttccg
     4321 cgtttccaga ctttacgaaa cacggaaacc gaagaccatt catgttgttg ctcaggtcgc
     4381 agacgttttg cagcagcagt cgcttcacgt tcgctcgcgt atcggtgatt cattctgcta
     4441 accagtaagg caaccccgcc agcctagccg ggtcctcaac gacaggagca cgatcatgcg
     4501 cacccgtggc caggacccaa cgctgcccga gatgcgccgc gtgcggctgc tggagatggc
     4561 ggacgcgatg gatatgttct gccaagggtt ggtttgcgca ttcacagttc tccgcaagaa
     4621 ttgattggct ccaattcttg gagtggtgaa tccgttagcg aggtgccgcc ggcttccatt
     4681 caggtcgagg tggcccggct ccatgcaccg cgacgcaacg cggggaggca gacaaggtat
     4741 agggcggcgc ctacaatcca tgccaacccg ttccatgtgc tcgccgaggc ggcataaatc
     4801 gccgtgacga tcagcggtcc aatgatcgaa gttaggctgg taagagccgc gagcgatcct
     4861 tgaagctgtc cctgatggtc gtcatctacc tgcctggaca gcatggcctg caacgcgggc
     4921 atcccgatgc cgccggaagc gagaagaatc ataatgggga aggccatcca gcctcgcgtc
     4981 gcgaacgcca gcaagacgta gcccagcgcg tcggccgcca tgccggcgat aatggcctgc
     5041 ttctcgccga aacgtttggt ggcgggacca gtgacgaagg cttgagcgag ggcgtgcaag
     5101 attccgaata ccgcaagcga caggccgatc atcgtcgcgc tccagcgaaa gcggtcctcg
     5161 ccgaaaatga cccagagcgc tgccggcacc tgtcctacga gttgcatgat aaagaagaca
     5221 gtcataagtg cggcgacgat agtcatgccc cgcgcccacc ggaaggagct gactgggttg
     5281 aaggctctca agggcatcgg tcgacgctct cccttatgcg actcctgcat taggaagcag
     5341 cccagtagta ggttgaggcc gttgagcacc gccgccgcaa ggaatggtgc atgcaaggag
     5401 atggcgccca acagtccccc ggccacgggg cctgccacca tacccacgcc gaaacaagcg
     5461 ctcatgagcc cgaagtggcg agcccgatct tccccatcgg tgatgtcggc gatataggcg
     5521 ccagcaaccg cacctgt
//
