LOCUS       sequence-146528-        1066 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Expresses Hook domain and first coiled coil of Hook3 in bacteria for
            purification.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 1066)
  AUTHORS   Schroeder CM, Vale RD
  TITLE     Assembly and activation of dynein-dynactin by the cargo adaptor 
            protein Hook3.
  JOURNAL   J Cell Biol. 2016 Aug 1;214(3):309-18. doi: 10.1083/jcb.201604002.
  PUBMED    27482052
REFERENCE   2  (bases 1 to 1066)
  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: "10.1083/jcb"; 
            journalName: "J Cell Biol"; date: "2016-08-1- 1"; volume: "214"; 
            issue: "3"; pages: "309-18"
COMMENT     Sequence Label: pET28a-Hook3 aa 1-239
FEATURES             Location/Qualifiers
     source          1..1066
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     primer_bind     70..89
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        70..88
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    89..113
                     /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             128..150
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             168..185
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             195..212
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             216..248
                     /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(264..287,288..323,324..347)
                     /codon_start=1
                     /product="two Strep-Tag(R)II moieties connected by a linker
                     for enhanced binding to Strep-Tactin(R), an engineered form
                     of streptavidin (Schmidt et al., 2013)"
                     /label=Twin-Strep-tag
                     /translation="WSHPQFEKGGGSGGGSGGSAWSHPQFEK"
     CDS             join(354..356,357..1064)
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /note="mammalian codon-optimized"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELY"
     CDS             354..1064
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Nager et al., 2011)"
                     /label=superfolder GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELY"
     primer_bind     complement(384..412)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     1007..1027
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
ORIGIN
        1 cctgtggcgc cggtgatgcc ggccacgatg cgtccggcgt agaggatcga gatctcgatc
       61 ccgcgaaatt aatacgactc actatagggg aattgtgagc ggataacaat tcccctctag
      121 aaataatttt gtttaacttt aagaaggaga tatacatggg cagcagccat catcatcatc
      181 atcacagcag cggcctggtg ccgcgcggca gccatatggc tagcatgact ggtggacagc
      241 aaatgggtcg cggatccgaa ttctggagtc atccacaatt tgagaagggc ggcggcagtg
      301 gcggcggcag tggcggcagt gcctggagcc acccgcagtt cgaaaaaggc agtatgagca
      361 aaggagaaga acttttcact ggagttgtcc caattcttgt tgaattagat ggtgatgtta
      421 atgggcacaa attttctgtc cgtggagagg gtgaaggtga tgctacaaac ggaaaactca
      481 cccttaaatt tatttgcact actggaaaac tacctgttcc gtggccaaca cttgtcacta
      541 ctctgaccta tggtgttcaa tgcttttccc gttatccgga tcacatgaaa cggcatgact
      601 ttttcaagag tgccatgccc gaaggttatg tacaggaacg cactatatct ttcaaagatg
      661 acgggaccta caagacgcgt gctgaagtca agtttgaagg tgataccctt gttaatcgta
      721 tcgagttaaa gggtattgat tttaaagaag atggaaacat tcttggacac aaactcgaat
      781 acaactttaa ctcacacaat gtatacatca cggcagacaa acaaaagaat ggaatcaaag
      841 ctaacttcaa aattcgccac aacgttgaag atggttccgt tcaactagca gaccattatc
      901 aacaaaatac tccaattggc gatggccctg tccttttacc agacaaccat tacctgtcga
      961 cacaatctgt cctttcgaaa gatcccaacg aaaagcgtga ccacatggtc cttcttgagt
     1021 ttgtaactgc tgctgggatt acacatggca tggatgagct ctacaa
//
