Open AccessPreparation of optimized concanavalin A-conjugated Dynabeads® magnetic beads for CUT&Tag
Author(s) -
Yasuhiro Fujiwara,
Yuji Tanno,
Hiroki Sugishita,
Yusuke Kishi,
Yoshinori Makino,
Yuki Okada
Publication year - 2021
Publication title -
plos one
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.99
H-Index - 332
ISSN - 1932-6203
DOI - 10.1371/journal.pone.0259846
Subject(s) - chromatin , concanavalin a , magnetic bead , chromatography , chemistry , conjugated system , biology , biophysics , biochemistry , dna , in vitro , polymer , organic chemistry
Epigenome research has employed various methods to identify the genomic location of proteins of interest, such as transcription factors and histone modifications. A recently established method called CUT&Tag uses a Protein-A Tn5 transposase fusion protein, which cuts the genome and inserts adapter sequences nearby the target protein. Throughout most of the CUT&Tag procedure, cells are held on concanavalin A (con A)-conjugated magnetic beads. Proper holding of cells would be decisive for the accessibility of Tn5 to the chromatin, and efficacy of the procedure of washing cells. However, BioMag®Plus ConA magnetic beads, used in the original CUT&Tag protocol, often exhibit poor suspendability and severe aggregation. Here, we compared the BioMag beads and Dynabeads® magnetic particles of which conjugation of con A was done by our hands, and examined the performance of these magnetic beads in CUT&Tag. Among tested, one of the Dynabeads, MyOne-T1, kept excessive suspendability in a buffer even after overnight incubation. Furthermore, the MyOne-T1 beads notably improved the sensitivity in CUT&Tag assay for H3K4me3. In conclusion, the arrangement and the selection of MyOne-T1 refine the suspendability of beads, which improves the association of chromatin with Tn5, which enhances the sensitivity in CUT&Tag assay.