Analyses of several seed viability markers in individual recalcitrant seeds of Eugenia stipitata McVaugh with totipotent germination.

The use of biochemical seed viability markers is often compromised by the unknown partitioning of analytes in bulk seed lots consisting of inseparable populations of viable and nonviable seeds. We took advantage of an unusual morphological syndrome found in the recalcitrant, undifferentiated seeds of Eugenia stipitata: one seed can be cut into several parts, each of which can germinate and develop into seedlings. We used four seed parts from one individual seed to analyse seed moisture content (MC), seed viability and the antioxidant glutathione (γ-glutamyl-cysteinyl-glycine; GSH), glutathione disulphide (GSSG) and intermediates of glutathione synthesis and breakdown. Seeds were exposed to different environmental MC to induce various levels of desiccation stress. Upon storage at high seed MC, seed viability was maintained, while GSH concentration increased and the glutathione half-cell reduction potential (EGSSG/2GSH ) was less negative than -215 mV, indicating GSH production and highly reducing conditions. Storage at low seed MC led to loss of GSH, resulting in a shift in EGSSG/2GSH , and seed death. In contrast, the cyst(e)ine half-cell reduction potential (ECySS/2CYS ) could not distinguish between the viability categories. Previous studies on seed populations revealed that the probability for a seed being alive is 50% at EGSSG/2GSH values between -180 and -160 mV. The single seed approach revealed that the window in which seed viability was lost could be slightly shifted towards more negative values. We discuss the contribution of cellular pH to EGSSG/2GSH and recommend E. stipitata as a recalcitrant seed model to study stress response on a single seed basis.

Inducing autotetraploids in cassava using oryzalin and colchicine and their in vitro morphophysiological effects.

Polyploid induction has been used for plant breeding to produce bigger and more robust plants than diploid types. The present study aimed to develop a methodology for in vitro induction of polyploidy in cassava. Apical and lateral microcuttings from the BRS Formosa variety were treated with six oryzalin concentrations for 24 and 48 h. The same methodology was used for colchicine with different concentrations. After 45 days of cultivation and an additional 45 days of subculture, the viability of the explants was assessed and plant acclimatization was performed. Ploidy was determined using flow cytometry. Oryzalin dose and exposure negatively affected cassava explant growth and development compared to untreated explants. Furthermore, apical and lateral explants responded differently to the treatments, showing a diversity in antimitotic sensitivity and effect that is tissue-type specific. In contrast, the doses of 1.25 to 6.25 mM colchicine resulted in high mortality of cassava explants. Therefore, the type of antimitotic affects the morphophysiological behavior of cassava plants in vitro, although apical explants have higher viability and regenerative capacity compared to lateral explants. In addition, the lateral explants have lower mixoploid rates compared to apical explants. Of the 310 plants generated by oryzalin treatments, 277 were diploid, 31 were mixoploid, and 2 were tetraploid. Exposure to oryzalin led to low rate of tetraploids and colchicine caused phytotoxic reactions and death of the explants. The tetraploids were multiplied in vitro to evaluate their yield in the field as well as their behavior against abiotic and biotic stress.