Morphological, Cytological, and Molecular-Based Genetic Stability Analysis of In Vitro-Propagated Plants from Newly Induced Aneuploids in Caladium

Abstract

Aneuploids are valuable materials of genetic diversity for genetic analysis and improvement in diverse plant species, which can be propagated mainly via in vitro culture methods. However, somaclonal variation is common in tissue culture-derived plants including euploid caladium. In the present study, the genetic stability of in vitro-propagated plants from the leaf cultures of two types of caladium (Caladium × hortulanum Birdsey) aneuploids obtained previously was analyzed morphologically, cytologically, and molecularly. Out of the randomly selected 23 and 8 plants regenerated from the diploid aneuploid SVT9 (2n = 2x − 2 = 28) and the tetraploid aneuploid SVT14 (2n = 4x − 6 = 54), respectively, 5 plants from the SVT9 and 3 plants from the SVT14 exhibited morphological differences from their corresponding parent. Stomatal analysis indicated that both the SVT9-derived variants and the SVT14-originated plants showed significant differences in stomatal guard cell length and width. In addition, the variants from the SVT14 were observed to have rounder and thicker leaves with larger stomatal guard cells and significantly reduced stomatal density compared with the regenerants of the SVT9. Amongst the established plants from the SVT9, two morphological variants containing 3.14–3.58% less mean fluorescence intensity (MFI) lost one chromosome, and four variants containing 4.55–11.02% more MFI gained one or two chromosomes. As for the plants regenerated from the SVT14, one variant with significantly higher MFI gained two chromosomes and three plants having significantly lower MFI resulted in losing four chromosomes. Three, out of the twelve, simple sequence repeat (SSR) markers identified DNA band profile changes in four variants from the SVT9, whereas no polymorphism was detected among the SVT14 and its regenerants. These results indicated that a relatively high frequency of somaclonal variation occurred in the in vitro-propagated plants from caladium aneuploids, especially for the tetraploid aneuploid caladium. Newly produced aneuploid plants are highly valuable germplasm for future genetic improvement and research in caladium.

Plant Regeneration via Organogenesis in Jerusalem Artichokes and Comparative Analysis of Endogenous Hormones and Antioxidant Enzymes in Typical and Atypical Shoots

“…Therefore, an assessment of the genetic stability of the in vitroregenerated plants is essential for a tissue culture procedure. Generally, the genetic fidelity of the regenerants can be confirmed by morphological, cytological, and DNA-based marker analysis [45,46]. In this study, the genetic homogeneity of the in vitro-produced plants was confirmed by flow cytometry and SSR analysis.…”

“…The reactions were conducted in an FC-96G thermocycler (BigFish, Hangzhou, China) as follows: initial denaturalization of the DNA at 94 • C for 2 min, followed by 40 cycles of amplification consisting of denaturation at 94 • C for 30 s, annealing at a temperature depending on the primer pairs for 30 s, and extension at 72 • C for 30 s, with a final extension at 72 • C for 10 min. PCR products were analyzed by electrophoresis in 8% polyacrylamide gels under non-denaturing conditions according to Yu et al [46].…”

In Vitro Microrhizome Production, Genetic Homogeneity Assessment, and Field Performance Evaluation in Ginger

“…The amplification was performed in an FC-96G thermocycler (BigFish, Hangzhou, China) under the following program: initial denaturation for 2 min at 94 • C, followed by 40 cycles of denaturation at 94 • C for 30 s, annealing at a temperature according to each primer for 30 s and extension at 72 • C for 30 s, and final extension at 72 • C for 10 min. PCR products were separated on an 8% (w/v) non-denaturing polyacrylamide gel and visualized by silver staining according to Yu et al [41].…”

Somatic Embryogenesis Induction and Genetic Stability Assessment of Plants Regenerated from Immature Seeds of Akebia trifoliate (Thunb.) Koidz

“…The PCR amplification was carried out on a T100 Thermo Cycler (BIO-RAD, Singapore) as follows: initial denaturalization of the DNA at 94 • C for 4 min, followed by 32 cycles of amplification consisting of denaturation at 94 • C for 60 s, annealing at a temperature depending on the Tm of the primer pairs used for 30 s, and extension at 72 • C for 30 s, with a final extension at 72 • C for 10 min. The microsatellite alleles were separated on non-denaturing 8% polyacrylamide gels in 1 × TBE buffer using a vertical electrophoresis instrument (DYCZ-24G, Beijing Liuyi Biotechnology Co., Ltd., Beijing, China) and visualized by silver staining, as described previously [29]. A 1000 bp DNA marker was used to evaluate the molecular weight of the detected bands.…”