LOCUS       sequence-257853-        4416 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Bacterial expression of RB3 stathmin-like domain (SLD).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 4416)
  AUTHORS   Monda JK, Whitney IP, Tarasovetc EV, Wilson-Kubalek E, Milligan RA, 
            Grishchuk EL, Cheeseman IM
  TITLE     Microtubule Tip Tracking by the Spindle and Kinetochore Protein Ska1
            Requires Diverse Tubulin-Interacting Surfaces.
  JOURNAL   Curr Biol. 2017 Dec 4;27(23):3666-3675.e6. doi: 
            10.1016/j.cub.2017.10.018. Epub 2017 Nov 16.
  PUBMED    29153323
REFERENCE   2  (bases 1 to 4416)
  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.cub.2017.10.018"; journalName: "Curr Biol"; date: 
            "2017-12-4- 4"; volume: "27"; issue: "23"; pages: "3666-3675.e6"
COMMENT     Sequence Label: pJM160
FEATURES             Location/Qualifiers
     source          1..4416
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    19..43
                     /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             56..78
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     promoter        complement(158..176)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(159..176)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(234..252)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      248..295
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(531..552)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     protein_bind    671..692
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     CDS             complement(705..1787)
                     /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
                     YIPPSTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     1749..1768
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1788..1865)
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     primer_bind     2011..2030
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     complement(2186..2205)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2305..2327
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2365..2383)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2451..2555
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2556..2624,2625..3416)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCHTLLSRIDAGQEQLGRRARYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      3587..4175
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4076..4095
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4329..4346
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 taatacgact cactataggg aattgtgagc ggataacaat tcctctagaa ataattttgt
       61 ttaactttaa gaaggagata tatccatgga tatcgaattc gtcgacctcg agggatccgg
      121 gccctctaga tgcggccgca tgcataagct tgagtattct atagtgtcac ctaaatccca
      181 gcttgatccg gctgctaaca aagcccgaaa ggaagctgag ttggctgctg ccaccgctga
      241 gcaataacta gcataacccc ttggggcctc taaacgggtc ttgaggggtt ttttgctgaa
      301 aggaggaact atatccggat aacctggcgt aatagcgaag aggcccgcac cgatcgccct
      361 tcccaacagt tgcgcagcct gaatggcgaa tggaaattgt aaacgttaat attttgttaa
      421 aattcgcgtt aaatatttgt taaatcagct cattttttaa ccaataggcc gaaatcggca
      481 aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga
      541 acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc
      601 agggcgatgg cccactacag cttgcatgcc tgcaggtcgg aagcataaag tgtaaagcct
      661 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
      721 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
      781 gtttgcgtat tgggcgccag ggtggttttt cttttcacca gtgagacggg caacagctga
      841 ttgcccttca ccgcctggcc ctgagagagt tgcagcaagc ggtccacgct ggtttgcccc
      901 agcaggcgaa aatcctgttt gatggtggtt gacggcggga tataacatga gctgtcttcg
      961 gtatcgtcgt atcccactac cgagatatcc gcaccaacgc gcagcccgga ctcggtaatg
     1021 gcgcgcattg cgcccagcgc catctgatcg ttggcaacca gcatcgcagt gggaacgatg
     1081 ccctcattca gcatttgcat ggtttgttga aaaccggaca tggcactcca gtcgccttcc
     1141 cgttccgcta tcggctgaat ttgattgcga gtgagatatt tatgccagcc agccagacgc
     1201 agacgcgccg agacagaact taatgggccc gctaacagcg cgatttgctg gtgacccaat
     1261 gcgaccagat gctccacgcc cagtcgcgta ccgtcttcat gggagaaaat aatactgttg
     1321 atgggtgtct ggtcagagac atcaagaaat aacgccggaa cattagtgca ggcagcttcc
     1381 acagcaatgg catcctggtc atccagcgga tagttaatga tcagcccact gacgcgttgc
     1441 gcgagaagat tgtgcaccgc cgctttacag gcttcgacgc cgcttcgttc taccatcgac
     1501 accaccacgc tggcacccag ttgatcggcg cgagatttaa tcgccgcgac aatttgcgac
     1561 ggcgcgtgca gggccagact ggaggtggca acgccaatca gcaacgactg tttgcccgcc
     1621 agttgttgtg ccacgcggtt gggaatgtaa ttcagctccg ccatcgccgc ttccactttt
     1681 tcccgcgttt tcgcagaaac gtggctggcc tggttcacca cgcgggaaac ggtctgataa
     1741 gagacaccgg catactctgc gacatcgtat aacgttactg gtttcacatt caccaccctg
     1801 aattgactct cttccgggcg ctatcatgcc ataccgcgaa aggttttgca ccattcgatg
     1861 gtgtcgggta ccgagctcga atttacgtga accatcaccc aaatcaagtt ttttgcggtc
     1921 gaggtgccgt aaagctctaa atcggaaccc taaagggagc ccccgattta gagcttgacg
     1981 gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag cgggcgctag
     2041 ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc
     2101 gccgctacag ggcgcgtcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt
     2161 cacaccgcat atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc
     2221 cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg
     2281 cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat
     2341 caccgaaacg cgcgagacga aagggcctcg tgatacgcct atttttatag gttaatgtca
     2401 tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc
     2461 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     2521 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
     2581 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     2641 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
     2701 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
     2761 cttttaaagt tctgctatgt catacactat tatcccgtat tgacgccggg caagagcaac
     2821 tcggtcgccg ggcgcggtat tctcagaatg acttggttga gtactcacca gtcacagaaa
     2881 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     2941 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
     3001 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     3061 aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc
     3121 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
     3181 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     3241 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     3301 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     3361 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
     3421 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     3481 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
     3541 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     3601 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     3661 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     3721 taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     3781 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     3841 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     3901 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     3961 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     4021 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     4081 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     4141 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     4201 ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt
     4261 ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga
     4321 ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cccaatacgc aaaccgcctc
     4381 tccccgcgcg ttggccgatt cattaatgca gctgat
//
