Shoot Buds Research Articles

Development of an Efficient In vitro Regeneration Protocol for Chrysanthemum (Chrysanthemum morifolium Ramat)

An efficient and rapid in vitro regeneration protocol was developed for chrysanthemum (Chrysanthemum morifolium Ramat) using two local varieties of Bangladesh namely, BARI Chrysanthemum-2 (BARI Chry-2) and local yellow (Y). MS medium supplemented with nine different concentrations and combinations of BAP and IAA was employed to optimize regeneration protocol using young in vitro derived leaf explants. Direct organogenesis was observed from the leaf explants on MS medium supplemented with 0.5 mg/l BAP and 2.0 mg/l IAA (T6) for both the varieties. This treatment (T6) induced shoot buds directly on the adaxial surface of the leaf providing the highest regeneration percentage (90% for BARI Chry-2 and 94.73% for Y), the highest number of shoot/explant (7.6 for BARI Chry-2 and 8.6 for Y) and maximum length of the shoot after six weeks (3 cm for BARI Chry-2 and 2.9 cm for Y) of culture. Explants with initially regenerated shoots were subculture on hormone free MS medium for shoot elongation after 4 weeks of their inoculation. During elongation of shoots, 90-95% of the regenerated shoots produced roots spontaneously in hormone free MS medium within 7-8 weeks of their inoculation. Rooted plantlets were transplanted to the field following hardening where 100% plantlets were survived and produced flower without any variation. Plant Tissue Cult. & Biotech. 31(2): 161-171, 2021 (December)

De Novo Shoot Bud Induction from the Catharanthus roseus Leaf Explants and Agrobacterium tumefaciens-Mediated Technique to Raise Transgenic Plants.

The regeneration of a whole plant from a single cell or organ explant was a valuable task for plant biotechnology. However, important medicinal plants such as Catharanthus roseus have shown recalcitrance to regeneration protocols, thus limiting investigations on MIA metabolism and metabolic engineering in this plant system. In this chapter, successful regeneration protocols were detailed for Catharanthus roseus, either by direct shoot bud induction from leaf explants and Agrobacterium-mediated genetic transformation.

Fenologia do abacate hass nos trópicos andinos de Caldas, Colômbia

Abstract The objective of this study was to document and analyze the effect of climate on the phenological development of ‘Hass’ avocado in two contrasting zones of the Andean tropics of Caldas, located at altitudes of 1,950m and 2,400m a.s.l., respectively. A completely randomized experimental design was carried out using a five to seven year-old ‘Hass’ tree as experimental unit and 15 replicates on each area. Observations were conducted on lateral sylleptic shoots and their corresponding apical buds located on the eastern and western sides of each tree. The destination of the apical bud of lateral shoots was quantified, floral phenology was documented, heat units were calculated from reproductive bud stage 1 to anthesis and to harvest and root growth was followed for each study area. Air temperature, soil temperature, precipitation and photosynthetically active radiation (PAR) values were recorded over time. Differences were found in the number of apical buds of lateral shoots with reproductive and vegetative destination, flowering phenology, heat units, root growth, PAR and precipitation. Differences were found among variables evaluated at the two contrasting altitudes. There is a clear need for further ecophysiological studies on this fruit tree in tropical areas in particular.

Study of anatomical and Acc (1-aminocyclopropane-1-carboxylate) during flower budabscission indicate an important role of alternate bearing

This study investigates whether, and how, 1-aminocyclopropane-1-carboxylic acid (ACC) is concerned in flower bud abscission of pistachio (Pistacia vera L.). Abscission was studied about the activation of the abscission zone, located between shoot and flower bud, from Uzun pistachio cultivar where the abscission profiles and the scanning electron micrographs pictorial differences in the abscission program, under natural conditions of flower bud. ACC content was investigated in the flower bud abscission zone during growth and abscission using liquid chromatography coupled with electrospray tandem mass spectrometry (UPLC/ESI-MS/MS). Results showed that abscission layers were started at first 45 days after full blooming (DAFB) and accomplished on 55 and 65 DAFB from 'On' year sampling. The activities of ACC in the flower bud abscission zone were significantly increased and decreased by flower bud abscission. A strong rise of ACC synthase in leaf and panicle was observed at the beginning of abscission, corresponding to stage 45 DAFB. High levels of ACC activity were found in whole abscission period indicating a possible role of ACC in pistachio flower bud abscission. At the end of the period (65 DAFB), the ACC levels gradually decreased and reached to minimum. Consequently, abscission-promoting effect of ACC depends on the phenological periods. These results provide strong evidence of flower abscission zone and the role of ACC in flower bud abscission possibly for the first time.

In-vitro regeneration in long-day garlic (Allium sativum)

The cloves of garlic (Allium sativum L.) cultivar Mukteshwar Local, after disinfection, were sprouted under in-vitro condition (85.00% sprouting) on MS medium containing 2 mg/l GA3. The cloves were soaked in 100 ppm GA3 solution for 24 hr. Maximum number of shoot buds (7.0) was recorded in treatment containing MS + 4.0 ppm BAP + 1.0 ppm NAA. Among different treatments for shoot multiplication, treatment MS+5.0 ppm BAP+ 1.0 ppm KIN + 0.5 ppm GA3 was the best. The number of shoots also varied with the level of sugar, light intensity and, light and dark cycle on shoot multiplication. They multiplied best in 30 g/l sucrose, 5000 lux light intensity and 16:8 hr of light and dark cycle. The developed protocol may be used for mass multiplication of garlic cultivars as well as regeneration of genetically transformed cell/tissue.

Regeneration potential of different explants during micropropagation of neem tree (Azadirachta indica A. Juss.)

Azadirachta indica A. Juss., (Neem), a prodigious multipurpose tree, has immense potential to benefit mankind and to protect the environment. In order to investigate the effects of three different explants for its regeneration potential, de embryonated cotyledon, immature zygotic embryo and nodal segments from a 30 year old neem plus tree were used. Half strength MS medium with benzyl amino purine (3 mg/L) and naphthalene acetic acid (0.5 mg/L) and casein hydrolysate (1 g/L) was effective in shoot bud sprouting from both nodes and cotyledons. Half strength MS medium fortified with TDZ (0.2 mg/L) was effective for induction of somatic embryogenesis from zygotic embryos. Shoot buds initiated from the cotyledons produced a maximum number of shoots per explants (4.33) which on further sub culturing induced maximum multiple shoots (15) on half strength MS medium fortified with BAP (1.5 mg/L), NAA (0.5 mg/L) and CH (400 g /L) and the nodal explants induced only 4-5 axillary shoots on further sub culturing. Even though immature zygotic embryos produced more number of somatic embryos per explant (24.97) within a short time (30-45 days), the plantlet conversion was poor (25.52 %). In vitro rooting was observed in half strength MS medium supplemented with IBA (2 mg/L). The regeneration potential of de embryonated cotyledons through a simple regeneration system may be beneficial for efficient mass propagation of selected plus trees of neem.

In-vitro propagation and phytochemical profiling of a highly medicinal and endemic plant species of the Himalayan region (Saussurea costus)

Efficient protocols for callus induction and micro propagation of Saussurea costus (Falc.) Lipsch were developed and phytochemical diversity of wild and in-vitro propagated material was investigated. Brown and red compact callus was formed with frequency of 80–95%, 78–90%, 70–95% and 65–80% from seeds, leaf, petiole and root explants, respectively. MS media supplemented with BAP (2.0 mgL−1), NAA (1.0 mgL−1) and GA3 (0.25 mgL−1) best suited for multiple shoot buds initiation (82%), while maximum shoot length was formed on media with BAP (1.5 mgL−1), NAA (0.25 mgL−1) and Kinetin (0.5 mgL−1). Full strength media with IAA (0.5 mgL−1) along with IBA (0.5 mgL−1) resulted in early roots initiation. Similarly, maximum rooting (87.57%) and lateral roots formation (up to 6.76) was recorded on full strength media supplemented with BAP (0.5 mgL−1), IAA (0.5 mgL−1) and IBA (0.5 mgL−1). Survival rate of acclimatized plantlets in autoclaved garden soil, farmyard soil, and sand (2:1:1) was 87%. Phytochemical analysis revealed variations in biochemical contents i.e. maximum sugar (808.32 µM/ml), proline (48.14 mg/g), ascorbic acid (373.801 mM/g) and phenolic compounds (642.72 mgL−1) were recorded from callus cultured on different stress media. Nonetheless, highest flavenoids (59.892 mg/g) and anthocyanin contents (32.39 mg/kg) were observed in in-vitro propagated plants. GC–MS analysis of the callus ethyl acetate extracts revealed 24 different phytochemicals. The variability in secondary metabolites of both wild and propagated plants/callus is reported for the first time for this species. This study may provide a baseline for the conservation and sustainable utilization of S. costus with implications for isolation of unique and pharmacologically active compounds from callus or regenerated plantlets.

Multiple shoot induction and regeneration of Vanilla borneensis Rolfe - a critically endangered orchid of Assam, India

In the present investigation, a micropropagation protocol has been developed for Vanilla borneensis Rolfe – a critically endangered orchid through multiple shoot regeneration. Through in vitro multiple shoot regeneration from both nodal and shoot tip explants, maximum (100%) shoot induction was observed. The minimum time required for shoot bud induction was observed from the shoot tip (5–7 days) on medium supplemented with BAP (4.44 mM) + KIN (2.32 mM) as compared to the nodal explants. Maximum multiple shoot regeneration was observed from nodal explants on the medium supplemented with BAP (4.44 mM) + TDZ (6.82 mM). However, maximum shoot length was observed on the medium supplemented with BAP (4.44 mM) + 15% CW and the number of nodes (5.27±0.33) per shoot after 90 days. Maximum (80-100%) of root initiation was observed in almost all the concentrations of NAA. The shortest time of root initiation was found on the medium supplemented with NAA (5.37 mM). Further, acclimatization period was found to be 15 days with 70% acclimatization while 60% of survivability was observed in the field condition. This efficient micropropagation method of V. borneensis could be successfully used for mass propagation as well as conservation of the critically endangered wild orchid.

In vitro organogenesis and plant regeneration of Passiflora xishuangbannaensis, a species with extremely small populations

Passiflora xishuangbannaensis (Passifloraceae) is endemic to a few sites of Mengyang nature reserve in Yunnan, Xishuangbanna and less than 40 individuals have been recorded. Nine Passiflora species are endemic to Yunnan with most species occurring in South America, making P. xishuangbannaensis highly significant and emblematic to the conservation work in the region. This study is designed to provide the first protocol for in vitro organogenesis and plant regeneration for ex situ conservation and reintroduction for an Asian Passiflora species. Using internodes, petioles and tendrils we optimize calli formation and root elongation using several plant growth regulators, individually or in combination. We also assess the genetic stability of regenerated cells. The maximum callus induction and shoot bud differentiation were both achieved on half Murashige and Skoog basal medium supplemented with 4.44 μM 6-Benzylaminopurine and 1.08 μM 1-Naphthaleneacetic acid. The best rooting was achieved from 30 days old, regenerated shoots on half Murashige and Skoog basal medium supplemented with 1.08 μM 1-Naphthaleneacetic acid. Micropropagated plants were subjected to inter simple sequence repeat markers analyses. Collectively, 86 bands were generated from 6 primers of which 12 bands were polymorphic, showing genetic variation between the regenerated plantlets and the original plant. Response to plant growth regulators was more specific than most other studies using South American species, which could be explained by the morphological and physiological differences between South American and Asian Passiflora species. Because very few individuals remain, sexual reproduction is challenging, aerial shoots are annual (rhizomatous) and because genetic variability can be a consequence of repetitive in-vitro regeneration, we advise ex-situ conservation techniques based on limited regeneration events such as cryopreservation.

Optimisation of micropropagation protocols for temperate eucalypt hybrids in South Africa, with a focus on auxin transport proteins

Globally the demand for forestry resources is booming, and consequently innovative approaches to cultivate valuable hardwoods, such as eucalypts, are crucial. Although micropropagation offers a means to clonally propagate desirable cultivars, this technique is often variety-specific for eucalypts. Thus, recalcitrant genotypes are often excluded from micropropagation programmes as development requires large investments of time, money and expertise. In this study, a minimal maintenance medium was developed to reduce subculturing frequency (∼8 days) and to evaluate each micropropagation stage independently. Furthermore, a generic protocol for use across different varieties was developed by assessing the in vitro performance of three different Eucalyptus grandis x Eucalyptus nitens varieties (varieties 1–3) across various treatments. The purpose of this protocol was to ease the in vitro establishment of novel eucalypt varieties by providing a foundation for future protocol optimisation. During multiplication, medium containing 0.5 mg l−1 meta-topolin with 0.1 mg l−1 of indole acetic acid (IAA) induced the highest bud proliferation and shoot elongation for varieties 1 (59.1 ± 2.9; 1.2 cm ± 0.08), 2 (62.3 ± 2.7; 1.5 cm ± 0.09) and 3 (58.1 ± 3; 1.2 cm ± 0.07). Following multiplication, the rooting treatment that achieved the most consistent rooting percentages among varieties 1 (33.4%), 2 (43.5%) and 3 (34.3%) incorporated 0.029 mg l−1 of racemic-GR24 with 0.5 mg l−1 of IAA. Additionally, root vigour was assessed by measuring the root number and length, which varied considerably based on the variety and treatment in question. Overall, variety 2 was the most amenable to plant growth regulators and upon further investigation was found to possess equal expression levels of the auxin transporters PIN1 and AUX1. Conversely, the other varieties displayed unequal ratios of these transporters. Considering that auxins are principal media components, these expression profiles may serve as markers to identify eucalypt cultivars amenable to micropropagation.

Short-term changes in chemical properties of topsoil (0–10 cm) after low-intensity fires caused by landings of first stages of space rockets Proton-M in Central Kazakhstan

Surface fires affect the species composition and the dynamic development of continental semi-arid ecosystems. We analyzed short-term post-pyrogenic changes in soil properties including pH, concentrations of nitrates, nitrites, ammonium, exchangeable calcium and magnesium and hydrolytic acidity in three key areas affected by low-intensity fires in comparison with background (control) ecosystems in Central Kazakhstan. There, only above-ground vegetation burned, but regeneration shoot buds survived. In all key areas, low-intensity fires were caused by regular landings of fragments of jettisoned first stages of Proton-M rockets launched from the Baikonur Cosmodrome. Low intensities and short periods (1–2 days) of pyrogenic impacts on vegetation predetermined only small changes in soil properties. There were significant (p-value < 0.05) decreases in exchangeable calcium and magnesium concentrations and pH and an increase in hydrolytic acidity in some samples. But changes in concentrations of nitrogen compounds were generally insignificant.

Micropropagation of Rare Scutellaria havanensis Jacq. and Preliminary Studies on Antioxidant Capacity and Anti-Cancer Potential.

We report the development of in vitro propagation protocols through an adventitious shoot induction pathway for a rare and medicinal Scutellaria havanensis. In vitro propagation studies using nodal explants showed MS medium supplemented with 10 µM 6-Benzylaminopurine induced the highest number of adventitious shoots in a time-dependent manner. A ten-day incubation was optimum for shoot bud induction as longer exposures resulted in hyperhydricity of the explants and shoots induced. We also report preliminary evidence of Agrobacterium tumefaciens EHA105-mediated gene transfer transiently expressing the green fluorescent protein in this species. Transformation studies exhibited amenability of various explant tissues, internode being the most receptive. As the plant has medicinal value, research was carried out to evaluate its potential antioxidant capacity and the efficacy of methanolic leaf extracts in curbing the viability of human colorectal cancer cell line HCT116. Comparative total polyphenol and flavonoid content measurement of fresh and air-dried leaf extract revealed that the fresh leaf extracts contain higher total polyphenol and flavonoid content. The HCT 116 cell viability was assessed by colorimetric assay using a 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide, showed a steady growth inhibition after 24 h of incubation. Scanning electron microscopy of leaf surface revealed a high density of glandular and non-glandular trichomes. This research provides a basis for the conservation of this rare plant and future phytochemical screening and clinical research.

A Reliable Regeneration Method in Genome-Editable Bell Pepper ‘Dempsey’

A reliable regeneration technique is critical for the improvement of pepper traits in the genome editing era. Recently, we reported that peppers were successfully and specifically edited using CRISPR tools, CRISPR/Cas9 and CRISPR/Cas12a (LbCpf1). Although genome-editing tools can be applied to modify peppers at the cellular level, feasible pepper regeneration techniques have not been developed. Therefore, we studied a pepper regeneration protocol for Capsicum annuum L. ‘Dempsey’, a bell pepper species that has been proven to be genome-editable. Three explant types were used in this study, including the first leaves, cotyledons and hypocotyls of pepper seedlings. The shoot buds of the tested explants were produced using 8 mg/L 6-benzylaminopurine (BAP)- and 6 mg/L indole-3-acetic acid (IAA)-containing shoot induction medium (SIM). The first leaves of the ‘Dempsey’ seedlings showed an average shooting rate of 69.8%, whereas the hypocotyls and cotyledons had approximately 25.5% and 19.5% shooting rates, respectively. The regenerated ‘Dempsey’ plants exhibited no alterations in fruit and fertile seed phenotypes. Furthermore, the parent ‘Dempsey’ and progenies of the regenerants were cytogenetically stable with the same chromosome numbers (2n = 24). Therefore, this regeneration protocol enables the precise molecular breeding of ‘Dempsey’ peppers when coupled with CRISPR tools.

In vitro propagation of an economically important medicinal plant Lawsonia inermis L. through nodal segments

The present investigation aimed to standardize efficient plant regeneration protocol through in vitro culture by using nodal segment for mass multiplication of Lawsonia inermis an economically important medicinal plant species. Mass multiplication of shoots induced on Murashige and Skoog (MS) medium supplemented with different growth regulators like auxins and cytokinins separately and in different combinations. The medium fortified with 6-Benzylaminopurine ( BAP) 1.0 mg/l + kinetin (KN) 1.5mg/l explained best compared to all other combinations. In vitro raised plantlets were excised and transferred in half strength MS medium supplemented with different growth regulators like Indole Butyric acid ( IBA) and naphthalene acetic acid (NAA ) (0.5-3.0 mg/l) in an experiment that gave rise to rooting. The half strength of MS medium additive with IBA in separate and in different combinations with NAA concentrations (0.5-3.0 mg/l) supported root development. The best response of rooting was obtained on half MS medium fortified with 1.0 mg/l IBA. The regenerated plantlets were successfully transplanted to pots. Regenerants were transferred to the field conditions and recorded the survival rate.. Among all the carbon sources and gelling agents used, sucrose (3%) in combination with 0.8 per cent agar-agar has proved significantly better. Multiple shoots formation with longer shoots were achieved on medium with 1.0mg/l BAP and 1.5mg/l Kn. Thus, it is possible to develop a large number of plants of L. inermis through shoot bud regeneration which can cater for the need of pharmaceutical as well as other industries.

Reversion of fruit‐dependent inhibition of flowering in Citrus requires sprouting of buds with epigenetically silenced CcMADS19

In Citrus, the response to environmental floral inductive signals is inhibited by the presence of developing fruits. The mechanism involves epigenetic activation of the CcMADS19 locus (FLC orthologue), encoding a floral repressor. To understand how this epigenetic regulation is reverted to allow flowering in the following season, we have forced precocious sprouting of axillary buds in fruit-bearing shoots, and examined the competence to floral inductive signals of old and new leaves derived from them. We have found that CcMADS19 is enriched in repressive H3K27me3 marks in young, but not old leaves, revealing that axillary buds retain a silenced version of the floral repressor that is mitotically transmitted to the newly emerging leaves, which are able to induce flowering. Therefore, we propose that flowering in Citrus is necessarily preceded by vegetative sprouting, so that the competence to respond to floral inductive signals is reset in the new leaves.

Effect of medium composition, genotype and age of explant on the regeneration of hexaploid plants from endosperm culture of tetraploid kiwiberry (Actinidia arguta)

Endosperm, an ephemeral and storage tissue, serves as a source of nutrition and protection during embryo development and germination. It can be used for the cultivation of polyploid plants in vitro. Here, results of plant regeneration and acclimatization from the endosperm-derived calli of four cultivars of Actinidia arguta has been presented. Seeds excised from fresh fruit and dry seeds stored for one year served as the sources of endosperm explants of selected tetraploid cultivars of A. arguta. Callus Induction Medium (CIM; containing 0.25, 0.5, or 1 mg/l of TDZ) and Actinidia Endosperm Medium (AEM; containing 2 mg/l of 2,4-D and 5 mg/l of kinetin) were used to study the organogenic responses of the calli. On AEM, the source of explant did not significantly affect the rate of callus induction for any of the tested cultivars; no organogenic events were observed. In contrast, on CIM both the source of explants and the cultivar origin caused significant differences in callus formation and subsequent organogenic events. Histological and ultrastructural analyses revealed the adventitious nature of shoot bud formation on these media. The most efficient elongation of shoot buds was achieved after transferring organogenic calli with adventitious shoot buds to a medium supplemented with zeatin or meta-topolin. Robust root induction with minimal basal callus formation occurred on the medium with indole-3-acetic acid. Flow cytometric analysis revealed that the nuclear DNA content in the leaves of some regenerants was approximately 50 % higher (4.5 pg/2C) than that in leaves from the tetraploid seedlings (3.1 pg/2C),which confirmed that those regenerants originated from the endosperm. The regeneration of such hexaploid plants was more efficient when endosperm from fresh seeds served as an explant; therefore, fresh rather than dry seeds are recommended for endosperm-derived plant production. The hexaploid plants of A. arguta can serve as an important source of breeding material.

Identification of apple TFL1-interacting proteins uncovers an expanded flowering network

MdTFL1, a floral repressor, forms protein complexes with several proteins and could compete with MdFT1 to regulate reproductive development in apple. Floral transition is a key developmental stage in the annual growth cycle of perennial fruit trees that directly determines the fruit development in the subsequent stage. FLOWERING LOCUS T (FT)/TERMINAL FLOWER1 (TFL1) family is known to play a vital regulatory role in plant growth and flowering. In apple, the two TFL1-like genes (MdTFL1-1 and MdTFL1-2) function as floral inhibitors; however, their mechanism of action is still largely unclear. This study aimed to functionally validate MdTFL1 and probe into its mechanism of action in apple. MdTFL1-1 and MdTFL1-2 were expressed mainly in stem and apical buds of vegetative shoots, with little expression in flower buds and young fruit. Expression of MdTFL1-1 and MdTFL1-2 rapidly decreased during floral induction. On the other hand, transgenic Arabidopsis, which ectopically expressed MdTFL1-1 or MdTFL1-2, flowered later than wild-type plants; demonstrating their in planta capability to function redundantly as flower repressors. Furthermore, we identified hundreds of novel interaction proteins of the two apple MdTFL1 proteins using yeast two-hybrid screens. Independent experiments for several proteins confirmed the yeast two-hybrid interactions. Among them, the transcription factor Nuclear Factor-Y subunit C (MdNF-YC2) functions as a promoter of flowering in Arabidopsis by activating LEAFY (LFY) and APETALA1 (AP1) expression. MdFT1 showed a similar interaction pattern as MdTFL1, implying a possible antagonistic action in the regulation of flowering. These newly identified TFL1-interacting proteins (TIPs) not only expand the floral regulatory network, but may also introduce new roles for TFL1 in plant development.

In vitro shoot multiplication using meta-Topolin and leaf-based regeneration of a withaferin A rich accession of Withania somnifera (L.) Dunal

Withania somnifera (L.) Dunal is a popular medicinal herb used in traditional Ayurvedic medicines. The pharmacological properties of this plant are largely due to the presence of withanolides such as withanone, withanolide A, withanolide D and withaferin A. Among these, withaferin A has gained a lot of attention because of its anti-cancer activity against a number of cancer cell lines. The current study deals with the selection of an elite accession of W. somnifera with high withaferin A content and the establishment of aseptic cultures under in vitro conditions for clonal mass propagation. Here, a novel cytokinin meta-Topolin (mT) was used for the first time in W. somnifera for shoot proliferation. mT (2.5 μM) was recorded as an efficient cytokinin for successful shoot multiplication and their growth. Further, an effective and reproducible regeneration protocol using leaf explant of W. somnifera has been developed for in vitro mass propagation and genetic transformation studies. The best shoot regeneration frequency and the maximum number of shoot buds were recorded in MS medium amended with BAP (10.0 μM). Among various auxins such as IAA, IBA, and NAA, IAA (10.0 μM) was proved to be optimal for both shoot and root growth. The regenerated plants were transferred to the greenhouse for their hardening and a high rate of survival (80 %) was recorded. The genetic uniformity of in vitro regenerated shoots was verified with RAPD, ISSR, and SCoT markers.

Development of high frequency cost-effective micropropagation protocol for Juncus rigidus using liquid culture medium and extraction of cellulose from their in vitro shoots - An important rush

Juncus rigidus is important halophyte with high cellulose content, but no studies has been made on micropropagation of the plant and cellulose extraction from in vitro shoots. To optimize an effective system for shoot multiplication, explants were grown in agar medium, liquid static culture, and bioreactor. Best shoot bud induction was obtained in liquid MS medium supplemented with2.27 μM thidiazuron (TDZ) over 0.46–4.65 μM Kinetin (KN) and 2.22–6.66 μM 6-benzyl aminopurine (BAP). Maximum number of shoot bud (36 ± 1.3) were achieved in static liquid MS medium supplemented with 2.22 μM BAP and 0.57 μM indole acetic acid (IAA). Upon subsequent subcultures >80 shoot buds were achieved in static liquid MS medium supplemented with 1.11 μM BAP, 0.57 μM IAA, and 0.46 μMKN within 30 days of culture. Being halophytic plant NaCl did not have any significant effect on shoot proliferation. Elongated shoots cultured in MS medium supplemented with 0.44 μM BAP, and 0.57 μM IAA developed 97% rooting in filter paper boat culture vessel. Sapling production cost in liquid culture medium was 90% less as compared to solid medium. Cellulose was extracted from the in vitro shoots and 17.64% cellulose achieved from liquid medium cultured shoots. For the first time micro-shoots are tested for in vitro cellulose production. • Maximum shoot bud (36 ± 1.2) induction was achieved in static liquid medium. • More than 80 shoot buds generated in static liquid MS medium supplemented with BAP, IAA, and KN. • 33.4 multiplication rate was recorded in static liquid medium. • 17.64 % cellulose extracted form in vitro shoots of J. rigidus. • In vitro plant based cellulose synthesis is important with future industrial application and bioengineering.

Shoot growth and bud topology in ‘Chandler’ walnut

Shoot growth and bud topology in ‘Chandler’ walnut