An asymmetric protoplast fusion and screening method for generating celeriac cybrids

Area of Science:

• Plant breeding
• Biotechnology
• Cell biology

Background:

• Celeriac hybrid seed production faces challenges due to unstable cytoplasmic male sterile (CMS) lines.
• Developing alternative alloplasmic CMS lines is crucial for improving celeriac breeding programs.

Purpose of the Study:

• To establish an asymmetric protoplast fusion and hybrid screening methodology for creating novel alloplasmic CMS lines in celeriac.
• To investigate the feasibility of interspecific protoplast fusion between celeriac and other species like carrot, coriander, and white celery.

Main Methods:

• Asymmetric somatic hybridization using celeriac suspension cell protoplasts as acceptors and carrot, coriander, or white celery mesophyll protoplasts as donors.
• Inhibition of acceptor cytoplasmic inheritance with iodoacetamide and donor nuclear genome inheritance with UV exposure.
• Protoplast fusion via electroporation and polyethylene glycol, followed by hybrid shoot regeneration and molecular marker analysis (chloroplast and mitochondrial DNA markers).

Main Results:

• Over 600 plants were analyzed, but no cybrids were initially identified using standard screening methods.
• Pooling microplantlets and using carrot-specific Atp1 marker detected successful fusion events between celeriac and carrot.
• The study demonstrated that viable hybrid creation is possible, albeit at a very low frequency.

Conclusions:

• Asymmetric protoplast fusion is a viable, though low-frequency, method for generating interspecific hybrids in celeriac.
• This research opens avenues for developing new cytoplasmic hybridization techniques and isolating novel CMS lines for celeriac.
• The findings contribute to advancing plant breeding strategies for crops like celeriac.