Somatic embryogenesis in mature zygotic embryos of Picea pungens

This study developed somatic embryogenesis protocols for Picea pungens (Engelm), an important ornamental species, including initiation, proliferation, maturation, germination, and acclimation. Somatic embryogenic tissues were induced from mature zygotic embryos of five families, with a frequency of $$\ge $$ ≥ 22% for each. Embryogenic tissues (ET) from 13 clones of three families were proliferated for one week, achieving an average rate of 179.1%. The ET of 38 clones of three families were cultured in maturation medium for six weeks; 188 mature embryos on average were counted per gram ET cultured, of which $$\ge $$ ≥ 81.1% appeared normal, and each clone developed at least 28 normally matured embryos. A total of 69.9% or more of cotyledonary somatic embryos germinated normally and developed into normal emblings. The experiment of transplanting the emblings into a greenhouse had an average survival rate of 68.5%. Considerable variation among and within families during initiation and proliferation was observed, but this variation decreased in the maturation and germination. Changing the concentration of plant growth regulator of the initiation medium did not significantly change the initiation frequency. We recommend incorporating these protocols into the current Picea pungens practical programs, although further research is essential to increase efficiencies and reduce cost.

Rice embryogenic trigger BABY BOOM1 promotes somatic embryogenesis by upregulation of auxin biosynthesis genes

ABSTRACTSomatic embryogenesis, a powerful tool for clonal propagation and for plant transformation, involves cellular reprogramming of differentiated somatic cells to acquire pluripotency. Somatic embryogenesis can be induced by treating explants with plant growth regulators. However, several plant species including agronomically important cereal crops remain recalcitrant to dedifferentiation and transformation except from embryonic tissues. Somatic embryogenesis can also be induced by ectopic expression of select embryonic factors, including in cereals by BABY BOOM (BBM) transcription factors. How BBM genes bypass the need for exogenous hormones is not well understood. Here, we investigated downstream targets during induction of somatic embryogenesis in rice by OsBBM1 ((Oryza sativa BABY BOOM1). Transient induction of OsBBM1 led to the upregulation of auxin biosynthesis OsYUCCA genes. Continued induction of OsBBM1 resulted in somatic embryogenesis without the need for exogenous auxins. Genetic mutant analysis of OsBBM1 downstream targets, OsYUCCA6, OsYUCCA7 and OsYUCCA9, show that they are required for normal rice development including root and shoot development. Somatic embryogenic potential of OsYUCCA triple mutants was highly compromised despite the presence of exogenous auxin. Additionally, we show that somatic embryogenesis induction by exogenous auxin in rice requires functional BBM genes. Thus, OsBBM1 mediated cellular reprogramming and somatic embryogenesis likely involves increased localized auxin through direct upregulation of OsYUCCA genes. This study reveals mechanistic details of how somatic embryogenesis is established in differentiated tissues in rice, a monocot model and agronomically important cereal crop, with the potential utility to improve regeneration from tissue culture for recalcitrant plants in future.One-sentence summaryRice BABY BOOM1 induces somatic embryogenesis from differentiated tissues by promoting auxin biosynthesis through direct upregulation of YUCCA genes.

YUCCA-Mediated Biosynthesis of the Auxin IAA Is Required during the Somatic Embryogenic Induction Process in Coffea canephora

Despite the existence of considerable research on somatic embryogenesis (SE), the molecular mechanism that regulates the biosynthesis of auxins during the SE induction process remains unknown. Indole-3-acetic acid (IAA) is an auxin that is synthesized in plants through five pathways. The biosynthetic pathway most frequently used in this synthesis is the conversion of tryptophan to indol-3-pyruvic acid (IPA) by tryptophan aminotransferase of Arabidopsis (TAA) followed by the conversion of IPA to IAA by enzymes encoded by YUCCA (YUC) genes of the flavin monooxygenase family; however, it is unclear whether YUC-mediated IAA biosynthesis is involved in SE induction. In this study, we report that the increase of IAA observed during SE pre-treatment (plants in MS medium supplemented with 1-naphthaleneacetic acid (NAA) 0.54 µM and kinetin (Kin) 2.32 µM for 14 days) was due to its de novo biosynthesis. By qRT-PCR, we demonstrated that YUC gene expression was consistent with the free IAA signal found in the explants during the induction of SE. In addition, the use of yucasin to inhibit the activity of YUC enzymes reduced the signal of free IAA in the leaf explants and dramatically decreased the induction of SE. The exogenous addition of IAA restored the SE process in explants treated with yucasin. Our findings suggest that the biosynthesis and localization of IAA play an essential role during the induction process of SE in Coffea canephora.

MICROPROPAGATION OF RHYNCHOSTYLIS GIGANTEA ORCHID BY SOMATIC EMBRYOGENIC CULTURES

Rhynchostylis gigantea is a wild orchid species commonly grown. Flowering at the traditional Tet holiday, the flower season always blooms in December of the lunar calendar. The flower has a faint aroma, so it is called Nghinh spring and is a kind of orchid with a national soul and national essence. Rhynchostylis gigantea is found in many areas in Vietnam as it is distributed in the South Central Highlands, especially the areas bordering Laos and Cambodia at low altitudes <600m. Rhynchostylis gigantea is a type of heat-resistant orchid, with suitable temperatures for orchid being from 26-30oC. Rhynchostylis gigantea species have many different colors such as red, white, shrimp brick color, blue and pink. The single-member tree does not assume pseudobulbs, grows vertically, has many pneumatophores and grows straight from the stem. Rhynchostylis gigantea grows slowly and regenerates with seeds. Due to high genetic diversity, it is difficult to maintain the parent’s characteristics. Applying techniques of somatic embryo culture in conservation and development of the Rhynchostylis gigantea tree to meet the increasing demand for seedlings is urgent.

Regeneration Rate of Eggplant Somatic Embryogenic In Various Maturation Media

Ralstonia solanacearum is one of the most important pathogen that causes bacterial wilt disease in eggplant and inhibits eggplant production. Improvement of eggplant varieties resistant to bacterial wilt can be accomplished through genetic manipulation. Regeneration of in vitro plants isone of the important tools to supports plant improvementthrough biotechnology. This study was aimed to determine the rate of eggplant regeneration in various maturation media, and to find the best medium for eggplant regeneration based on maturation rate and the number of cotyledon produced. We used resistant eggplant (accession 032) as the material to produce somatic embryogenic.There were 7 types of regeneration media used in this research. MS medium was supplemented with a certain concentration of plant growt regulators , such as: 1 mg / L + BAP 1 mg / L, NAA 4mg / L, TDZ 0.005 mg / L, TDZ 0.001 mg / L, CuSO4 2mM + BAP 1 mg / L, CuSO4 2mM + BAP 2 mg / L and Kinetin 1 mg / L + CuSO4 2mM. Three clumps of callus per plate with three replications were transferred to MS suplemented medium. The parameters observed were the color of callus before and after they were transfered to regeneration medium, the day of formation of globular, heart-shaped, tubular and cotyledonary phase, and the number of cotyledons formed. The results obtained showed the somatic embryogenic color of the 032 genotype was white with friable structure before being transferred to regeneration medium and was turned to yellowish white after being transferred to the regeneration medium. On the day sixth, friable embryogenic somatic of eggplant was developed into nodule on medium MS + NAA 4 mg / L, MS + CuSO4 2mM + BAP medium 1 mg / L, and MS + CuSO4 2mM + BAP 2 mg / L. Somatic embryogenic callus of accession 032 were able to pass complete globular, heart-shaped, tubular and cotyledonary phase. The most responsive medium for somatic embryogenic callus regeneration, based on the days of the callus phases formation and the number of early-phase cotyledons obtained, were MS medium suplemented with CuSO4 2mM + BAP,and CuSO4 2 mM + BAP 2 mg / L.Keywords: eggplant, Ralstonia solanacearum, regeneration, cotyledonary, clump, BAP