Next-generation sequencing coupled with a cell-free display technology for high-throughput production of reliable interactome data.
Next-generation sequencing (NGS) has been applied to various kinds of omics studies, resulting in many biological and medical discoveries. However, high-throughput protein-protein interactome datasets derived from detection by sequencing are scarce, because protein-protein interaction analysis requires many cell manipulations to examine the interactions. The low reliability of the high-throughput data is also a problem. Here, we describe a cell-free display technology combined with NGS that can improve both the coverage and reliability of interactome datasets. The completely cell-free method gives a high-throughput and a large detection space, testing the interactions without using clones. The quantitative information provided by NGS reduces the number of false positives. The method is suitable for the in vitro detection of proteins that interact not only with the bait protein, but also with DNA, RNA and chemical compounds. Thus, it could become a universal approach for exploring the large space of protein sequences and interactome networks.
Division of Interactome Medical Sciences, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan.
Shigeo
S
Fujimori
Naoya
N
Hirai
Hiroyuki
H
Ohashi
Kazuyo
K
Masuoka
Akihiko
A
Nishikimi
Yoshinori
Y
Fukui
Takanori
T
Washio
Tomohiro
T
Oshikubo
Tatsuhiro
T
Yamashita
Etsuko
E
Miyamoto-Sato
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002474:Cell-Free System; D019295:Computational Biology; D018076:DNA, Complementary; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D051379:Mice; D025941:Protein Interaction Mapping / Q000379:methods*; D011506:Proteins / Q000737:chemistry / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods; D015203:Reproducibility of Results; D017422:Sequence Analysis, DNA
D018076:DNA, Complementary; D011506:Proteins; D012333:RNA, Messenger