LOCUS       sequence-272846-        4808 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of RB3 stathmin-like domain (SLD).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4808)
  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 4808)
  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..4808
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        34..111
                     /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."
     CDS             112..1194
                     /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(131..150)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     primer_bind     1344..1365
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     terminator      complement(1601..1648)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     1644..1662
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     promoter        1720..1738
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1720..1737
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     protein_bind    1769..1793
                     /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             1806..1828
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2277..2294
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2366..2383)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2537..3125)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2617..2636)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3296..4087,4088..4156))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     3919..3938
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4157..4261)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4329..4347
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4385..4407)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4507..4526
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4680..4699)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 agggtgatgg ttcacaattc gagctcggta cccgacacca tcgaatggtg caaaaccttt
       61 cgcggtatgg catgatagcg cccggaagag agtcaattca gggtggtgaa tgtgaaacca
      121 gtaacgttat acgatgtcgc agagtatgcc ggtgtctctt atcagaccgt ttcccgcgtg
      181 gtgaaccagg ccagccacgt ttctgcgaaa acgcgggaaa aagtggaagc ggcgatggcg
      241 gagctgaatt acattcccaa ccgcgtggca caacaactgg cgggcaaaca gtcgttgctg
      301 attggcgttg ccacctccag tctggccctg cacgcgccgt cgcaaattgt cgcggcgatt
      361 aaatctcgcg ccgatcaact gggtgccagc gtggtggtgt cgatggtaga acgaagcggc
      421 gtcgaagcct gtaaagcggc ggtgcacaat cttctcgcgc aacgcgtcag tgggctgatc
      481 attaactatc cgctggatga ccaggatgcc attgctgtgg aagctgcctg cactaatgtt
      541 ccggcgttat ttcttgatgt ctctgaccag acacccatca acagtattat tttctcccat
      601 gaagacggta cgcgactggg cgtggagcat ctggtcgcat tgggtcacca gcaaatcgcg
      661 ctgttagcgg gcccattaag ttctgtctcg gcgcgtctgc gtctggctgg ctggcataaa
      721 tatctcactc gcaatcaaat tcagccgata gcggaacggg aaggcgactg gagtgccatg
      781 tccggttttc aacaaaccat gcaaatgctg aatgagggca tcgttcccac tgcgatgctg
      841 gttgccaacg atcagatggc gctgggcgca atgcgcgcca ttaccgagtc cgggctgcgc
      901 gttggtgcgg atatctcggt agtgggatac gacgataccg aagacagctc atgttatatc
      961 ccgccgttaa ccaccatcaa acaggatttt cgcctgctgg ggcaaaccag cgtggaccgc
     1021 ttgctgcaac tctctcaggg ccaggcggtg aagggcaatc agctgttgcc cgtctcactg
     1081 gtgaaaagaa aaaccaccct ggcgcccaat acgcaaaccg cctctccccg cgcgttggcc
     1141 gattcattaa tgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa
     1201 cgcaattaat gtaagttagc tcactcatta ggcaccccag gctttacact ttatgcttcc
     1261 gacctgcagg catgcaagct gtgggccatc gccctgatag acggtttttc gccctttgac
     1321 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc
     1381 tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa
     1441 aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa cgcttacaat
     1501 ttccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg
     1561 ctattacgcc aggttatccg gatatagttc ctcctttcag caaaaaaccc ctcaagaccc
     1621 gtttagaggc cccaaggggt tatgctagtt attgctcagc ggtggcagca gccaactcag
     1681 cttcctttcg ggctttgtta gcagccggat caagctggga tttaggtgac actatagaat
     1741 actcaagctt wwryaygmsk cmctataggg aattgtgagc ggataacaat tcctctagaa
     1801 ataattttgt ttaactttaa gaaggagata tatccatggc cgacatggaa gtcatcgagc
     1861 tgaacaaatg cacctcgggc caatcctttg aagtcatcct gaagccaccc tcctttgatg
     1921 gggttcccga gttcaacgcc tccctgccaa ggcggcgaga cccatccctg gaagagatcc
     1981 agaagaaact agaagcggct gaggagcgaa ggaagtacca ggaagcggag ctcctgaaac
     2041 acctagcaga gaaacgggaa catgagagag aggtgatcca aaaggccatt gaggaaaaca
     2101 acaacttcat caagatggct aaggaaaaac tggcccagaa gatggaatcc aacaaggaga
     2161 acagggaggc ccacctcgcc gccatgttgg aacggctgca agagaaggac aagcacgccg
     2221 aggaggtgcg gaaaaacaag gagctgaagg aagaggcctc caggggcggc agcggccacc
     2281 atcaccatca ccattgagga tcssrkymmt swakmkgcsr mcgcrygsrk arrskcggtt
     2341 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     2401 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     2461 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     2521 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     2581 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     2641 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     2701 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     2761 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     2821 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     2881 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     2941 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
     3001 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     3061 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     3121 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     3181 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     3241 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     3301 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     3361 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     3421 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     3481 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     3541 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     3601 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     3661 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     3721 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     3781 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     3841 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     3901 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     3961 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     4021 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     4081 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     4141 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     4201 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     4261 gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac cattattatc atgacattaa
     4321 cctataaaaa taggcgtatc acgaggccct ttcgtctcgc gcgtttcggt gatgacggtg
     4381 aaaacctctg acacatgcag ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg
     4441 ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg ggctggctta
     4501 actatgcggc atcagagcag attgtactga gagtgcacca tatgcggtgt gaaataccgc
     4561 acagatgcgt aaggagaaaa taccgcatca ggacgcgccc tgtagcggcg cattaagcgc
     4621 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
     4681 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     4741 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     4801 acttgatt
//
