Author

Kangqi Li

Advisor

Reichert, Nancy A.

Committee Member

Klink, Vincent

Committee Member

Peng, Zhaohua

Committee Member

Young, Margaret M.

Date of Degree

5-1-2020

Original embargo terms

Visible to MSU only for 6 months||forever||12/15/2020

Document Type

Graduate Thesis - Open Access

Department

Department of Biological Sciences

Abstract

The potential for Hibiscus cannabinus L. (kenaf) improvement via genome editing using the CRISPR/Cas9 system to generate gene knock-outs was explored. Studies included target gene identification, target guide RNA (gRNA) selection, plant tissue (explant) choice and media composition for plant regeneration. A putative kenaf phytoene desaturase gene (pds, GEED01047592.1) was identified in the kenaf transcriptome, and molecularly confirmed. Kenaf seedling tissues were transformed via Agrobacterium tumefaciens containing the cas9 gene (endonuclease required for gene knock-out) and each gRNA separately; putative transgenic calli and adventitious shoots arose on a medium containing 1-naphthaleneacetic acid, thidiazuron and silver nitrate. Tissues appeared chlorotic/albino and shoots remained diminutive/dwarf-like. These unique morphologies had also been noted by researchers who successfully knocked out the pds gene in other plant species. Cas9 DNA was detected in these putative transgenic kenaf tissues, but initial DNA sequencing analysis did not confirm knock-out/mutations in targeted areas of the pds gene.

URI

https://hdl.handle.net/11668/16695

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