Nguyen Van Doai has been a researcher at the Vietnam Academy
of Science and Technology since 2012. He earned his Bachelor of Biotechnology
at VNU University of Science, Vietnam National University in 2012, and his Ms
in Experimental Biology at the Institute of Ecology and Biological Resource at
Vietnam Academy of Science and Technology. He has completed his PhD program in
Integrated biological science at Pusan University (Busan – Korea). He also
worked as an internship student at Sejong University (2017-2018) and Pusan
University (2019-2020) where he joined several plant genetic and gene function
study projects. Currently, he is focusing on applied bioscience, especially the
application of genome editing systems in plants, animals, and bacterial.
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An efficient hairy root system for validation
of plant transformation vector and CRISPR/Cas construct activities in cucumber
(Cucumis sativus L.)
D.V. NGUYEN,
T.T. HOANG, N.T. LE, H.T. TRAN, C.X.
NGUYEN, Y.H. MOON, H.H. CHU, & P.T. DO
Laboratory of Plant Cell Biotechnology, Institute of
Biotechnology, Vietnam Academy of Science and Technology – A10, 18 – Hoang Quoc
Viet, Cau Giay, Hanoi 11355, Vietnam
Hairy root induction system has been applied in various
plant species as an effective method to study gene expression and function.
Recently, these systems have shown to be an effective tool to evaluate
activities of CRISPR/Cas9 systems for genome editing. In this study, Rhizobium rhizogenes mediated hairy root
induction was optimized to provide an effective tool for validation of plant
transformation vector, CRISPR/Cas9 construct activities as well as selection of
targeted gRNAs for gene editing in cucumber. Under the optimized conditions
including OD650 at 0.4 for infection and 5 days of co-cultivation,
the highest hairy root induction frequency reached 100% for the cucumber
variety Choka F1. This procedure was successfully utilized to overexpress a
reporter gene (gus) and induce
mutations in two Lotus japonicus
ROOTHAIRLESS1 homolog genes CsbHLH66
and CsbHLH82 using CRISPR/Cas9
system. For induced mutation, about 78% of transgenic hairy roots exhibited
mutant phenotypes including sparse root hair and root hair-less. The targeted
mutations were obtained in individual CsbHLH66,
CsbHLH82, or both CsbHLH66 and CsbHLH82 genes by heteroduplex analysis and sequencing. The hairy
root transformation system established in this study is sufficient and
potential for further research in genome editing of cucumber as well as other
cucumis plants.