Effect of AgNO3 and BAP on Root as a Novel Explant in Date Palm (Phoenix dactylifera cv. Medjool) Somatic Embryogenesis.

BACKGROUND AND OBJECTIVE: Somatic embryogenesis techniques are used for cloning a wide range of varieties of date palms around the world. The aim of the present study was to develop an efficient method with the lowest cost and the greatest potential to obtain in vitro plantlets of date palm cv. Medjool. Also, produce embryogenic callus and somatic embryos without using 2,4-dichlorophenoxyacetic acid (2,4-D). METHODOLOGY: In this study, produced plantlets through somatic embryogenesis were used in vitro roots as explant cultured on Murashige and Skoog (MS) media containing three level of Silver Nitrate (AgNO3) (0, 3 and 6 mg L-1) plus two level of 6-benzylaminopurine (BAP) (0 and 2 mg L-1) plus 0.1 mg L-1 1-naphthylacetic acid (NAA) for callus induction. After 12 weeks of culture, callus induction and after 16 weeks, production of embryogenic callus and embryos were occurred from root explants. RESULTS: According to the results, medium containing 2 mg L-1 BAP and 3 mg L-1 silver nitrate+0.1 mg L-1 NAA showed the highest amount of embryogenic callus fresh weight (1.38 g). This treatment also cause the highest number and length of embryos by production of 90.04 embryogenic callus with length of 11.18 mm. On the other hand, shoots were appeared from germinated embryos and white roots began to appear within 8 weeks. Medium contains 3 mg L-1 BAP and 0.1 mg L-1 NAA with average of 12.27 cm shoot length and 15.48 cm root length was the best. Control treatment had the lowest average shoot (3.71 cm) and root (5.03 cm) length. CONCLUSION: This study showed that certain concentration of silver nitrate and BAP has stimulating effect on growth of produced embryonic callus from root segments of Medjool cultivar of date palm.

Microcalli Induction in Protoplasts Isolated from Embryogenic Callus of Date Palm.

Date palm (Phoenix dactylifera L.) production is severely hampered due to several pests and diseases. Biotechnological tools such as protoplast fusion appear as an alternative to ensure rapid genetic improvement and multiplication of this species. However, establishment of an effective system of plant regeneration from protoplasts culture is a prerequisite for date palm somatic hybridization. In this chapter, we describe an effective protocol to induce microcalli in protoplasts isolated from nodular callus of important Algerian date palm cultivars. In this protocol, the main factors influencing the isolation (i.e., enzymatic solution, mannitol concentration, duration, and mode of maceration) of protoplasts from the calli of Algerian date palm cultivars were optimized. Purified protoplasts were cultured on a semisolid medium supplemented with a hormonal balance of auxin and cytokinin to obtain microcalli formation.

Genetic Transformation of Date Palm Via Microprojectile Bombardment.

Efficient protocols for date palm embryogenic callus and somatic embryo transformation with uidA gene are described in this chapter. The embryogenic callus transformation procedure is 1.6 µm gold particle size coated with 2.5 µg DNA (pAct1-D plasmid), 1100 psi helium pressure, 9 cm target distance, 26 inHg vacuum pressure, 3 mm distance between the rupture disk and macrocarrier, and osmotic pretreatment with 0.4 M mannitol followed by 60 min air desiccation. The somatic embryo transformation procedure is 0.6 µm gold particle size coated with 2.5 µg DNA (pAct1-D plasmid), 1350 psi helium pressure, 6 cm target distance, 28 inHg vacuum pressure, 3 mm distance between the rupture disk and macrocarrier, and osmotic pretreatment with 0.4 M mannitol followed by 60 min air desiccation. Protocols for analysis of the transgenic plantlets have also been described.

Microprojectile Bombardment Transformation of Date Palm Using the Insecticidal Cholesterol Oxidase (ChoA) Gene.

The overall objective of this work is to optimize the transformation system for date palm as a first step toward production of date palm clones resistant to noxious pests. A construct harboring the cholesterol oxidase (ChoA) gene, which renders plant resistance against insect attack, is introduced into embryogenic date palm callus using the PDS-1000/He particle bombardment system. The process involves the establishment of embryogenic callus cultures as well as immature embryo-derived microcalli that are used as target tissues for shooting and optimization of transformation conditions. This chapter in addition explains molecular and histochemical assays conducted to confirm gene integration and expression.

Bioreactor Steroid Production and Analysis of Date Palm Embryogenic Callus.

Several compounds and families of compounds of date palm secondary metabolites have been investigated. The analysis of date palm tissue has shown the abundance of secondary metabolites including phytosterols, e.g., steroids, an important group of pharmaceutical compounds. Biotechnology offers the opportunity to utilize cells, tissues, and organs grown in vitro and manipulated to obtain desired compounds. This chapter presents a protocol for the production, determination, and identification of steroids in date palm callus tissue. The addition of 0.01 mg/L pyruvic acid as a precursor to MS liquid culture medium enhances steroid production. In addition, the chapter describes the sterol analytical techniques based on gas-liquid chromatography and gas chromatography-mass spectrometry.

Somatic Embryogenesis of Date Palm (Phoenix dactylifera L.) Through Cell Suspension Culture.

Date palm (Phoenix dactylifera L.) is the oldest and most economically important plant species distributed in the hot arid regions of the world. Propagation of date palm by seeds produces heterogeneous offspring with inferior field performance and poor fruit quality. Traditionally, date palm is propagated by offshoots, but this method is inefficient for mass propagation because of limited availability of offshoots. Plant regeneration through tissue culture is able to provide technologies for the large-scale propagation of healthy true-to-type plants. The most commonly used technology approach is somatic embryogenesis which presents a great potential for the rapid propagation and genetic resource preservation of this species. Significant progress has been made in the development and optimization of this regeneration pathway through the establishment of embryogenic suspension cultures. This chapter focuses on the methods employed for the induction of callus from shoot tip explants, establishment of cell suspension culture, and subsequent somatic embryogenesis and plant regeneration.

The Domestication Syndrome in Phoenix dactylifera Seeds: Toward the Identification of Wild Date Palm Populations.

Investigating crop origins is a priority to understand the evolution of plants under domestication, develop strategies for conservation and valorization of agrobiodiversity and acquire fundamental knowledge for cultivar improvement. The date palm (Phoenix dactylifera L.) belongs to the genus Phoenix, which comprises 14 species morphologically very close, sometimes hardly distinguishable. It has been cultivated for millennia in the Middle East and in North Africa and constitutes the keystone of oasis agriculture. Yet, its origins remain poorly understood as no wild populations are identified. Uncultivated populations have been described but they might represent feral, i.e. formerly cultivated, abandoned forms rather than truly wild populations. In this context, this study based on morphometrics applied to 1625 Phoenix seeds aims to (1) differentiate Phoenix species and (2) depict the domestication syndrome observed in cultivated date palm seeds using other Phoenix species as a "wild" reference. This will help discriminate truly wild from feral forms, thus providing new insights into the evolutionary history of this species. Seed size was evaluated using four parameters: length, width, thickness and dorsal view surface. Seed shape was quantified using outline analyses based on the Elliptic Fourier Transform method. The size and shape of seeds allowed an accurate differentiation of Phoenix species. The cultivated date palm shows distinctive size and shape features, compared to other Phoenix species: seeds are longer and elongated. This morphological shift may be interpreted as a domestication syndrome, resulting from the long-term history of cultivation, selection and human-mediated dispersion. Based on seed attributes, some uncultivated date palms from Oman may be identified as wild. This opens new prospects regarding the possible existence and characterization of relict wild populations and consequently for the understanding of the date palm origins. Finally, we here describe a pipeline for the identification of the domestication syndrome in seeds that could be used in other crops.

Effect of potassium nitrate on antioxidants production of date palm (Phoenix dactylifera L.) in vitro.

Antioxidants present in dates are necessary for all physiological processes of humans and animals. In Saudi Arabia date palm is a national fruit tree, produces millions of tons of dates for consumption and is considered a major source of antioxidants. The main aim of this study was to determine the role of potassium nitrate (KNO3) in the formation of antioxidants from explants collected from date palm cultivars in Al-Ahsa Oasis, Kingdom of Saudi Arabia (KSA) and to monitor the extent of its effect on growth and development of cells during callus formation stage via somatic embryogenesis. The results showed that full concentration of KNO3 was the best for callus formation in general. While, the half concentration of KNO3 played an important role for stimulating the explants to form phenolic compounds and the browning emergence for all the cultivars under investigation. On the other hand, the chemical analysis for measuring the phenolic compounds in the explants showed that all the explants formed antioxidants but with varying degrees. The highest mean of phenolic contents was found in those explants cultured with the half concentration of KNO3 for Shishi cv 2.053 ± 0.010a mg g(-1) and antioxidant activity by ABTS Inhibition and UM Trolox was 80.694 ± 0.439 and 801.575 ± 2.391, respectively.