Somatic Embryogenesis and Plant Regeneration from Sugi (Japanese Cedar, Cryptomeria japonica D. Don, Cupressaceae) Seed Families by Marker Assisted Selection for the Male Sterility Allele ms1.

Tsuyoshi E Maruyama,Takumi Kaneeda,Yoshinari Moriguchi,Saneyoshi Ueno,Satoko Hirayama

doi:10.3390/plants9081029

Abstract

One of the possible countermeasures for pollinosis caused by sugi (Cryptomeria japonica), a serious public health problem in Japan, is the use of male sterile plants (MSPs; pollen-free plants). However, the production efficiencies of MSPs raised by conventional methods are extremely poor, time consuming, and resulting in a high seedling cost. Here, we report the development of a novel technique for efficient production of MSPs, which combines marker-assisted selection (MAS) and somatic embryogenesis (SE). SE from four full sib seed families of sugi, carrying the male sterility gene MS1, was initiated using megagametophyte explants that originated from four seed collections taken at one-week intervals during the month of July 2017. Embryogenic cell lines (ECLs) were achieved in all families, with initiation rates varying from 0.6% to 59%. Somatic embryos were produced from genetic marker-selected male sterile ECLs on medium containing maltose, abscisic acid (ABA), polyethylene glycol (PEG), and activated charcoal (AC). Subsequently, high frequencies of germination and plant conversion (≥76%) were obtained on plant growth regulator-free medium. Regenerated plantlets were acclimatized successfully, and the initial growth of male sterile somatic plants was monitored in the field.

Highlights

Sugi, which accounts for 44% of Japan’s planted forest area, is the most important tree species in forestry
We examined whether use of a DNA marker for marker-assisted selection (MAS) to achieve early selection of male sterile embryogenic cell lines (ECLs) at the undifferentiated cell stage can be combined with large-scale somatic embryo propagation to produce a possible 100% male sterile plants (MSPs) production rate
The whole megagametophyte containing the zygotic embryo was used as the initial explant for induction and culture of Embryogenic cell lines (ECLs) to be used for somatic embryogenesis (SE)

Summary

Introduction

Sugi, which accounts for 44% of Japan’s planted forest area, is the most important tree species in forestry. Over 30% of the total population in Japan Tokyo) suffer from sugi pollinosis, an allergic reaction, resulting in an estimated economic loss of more than 600 billion yens per year, which represents a serious social and public health problem. One possible countermeasure against sugi pollinosis is to use male sterile plants (MSPs), which produce no pollen. The first natural male sterile sugi was discovered in Toyama Prefecture in 1992 [1], and its frequency in the planted forest area is estimated to be one in several thousand [2]. As a result of vigorous selection across the country, 23 male sterile sugi individuals have been discovered [3].

Methods

Results

Conclusion