Rapid in vitro cloning of a 40-year-old tree of Azadirachta indica A. Juss. (Neem) employing nodal stem segments

Arora, Kavita; Sharma, Meena; Srivastava, J. N.; Ranade, S. A.; Sharma, Ashok Kumar

doi:10.1007/s10457-009-9230-1

Cited by 33 publications

(24 citation statements)

References 24 publications

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“…Similar results of seasonal effects on culture establishment have been reported in mulberry (Chitra and Padmaja 2002), neem (Chaturvedi et al 2004), litchi (Kumar et al 2006) and lotus (Shou et al 2008). The thickness and maturity of the explant with increasing distance from the apex of the shoot not only determine their survival in culture but also contribute to their effective performance in respect of proliferation (Arora et al 2010). This could be the reason for a significant difference (23%) between percent aseptic cultures (87.4%) and percent bud-break (64.1%).…”

Section: Discussionsupporting

confidence: 69%

Improved clonal propagation of Spilanthes acmella Murr. for production of scopoletin

Singh

Chaturvedi

2010

Plant Cell Tiss Organ Cult

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A reproducible protocol for clonal propagation of Spilanthes acmella has been established. Routinely, the cultures were established in spring (January-April) season because of the highest aseptic culture establishment and high frequency shoot proliferation. Incorporation of 5 lM N 6 -benzyladenine (BA) to Murashige and Skoog (MS) basal medium showed 100% bud-break and promoted multiple shoot proliferation in cultures. Interestingly, a higher concentration of BA (7-15 lM) promoted stunted shoots with pale leaves while a lower concentration (1-3 lM) resulted in shoots with long internodes and excessive adventitious root proliferation from all over their surface. For recurrent shoot multiplication, single node segments from in vitro-developed shoots were excised and cultured on MS ? BA (5 lM) medium where 20.3-fold shoot multiplication was achieved every 5 weeks. Finally, these shoots were successfully rooted on half-strength MS medium (major salts reduced to half-strength) with 50 g l -1 sucrose, with a frequency of 100%. Transplantation survival of micropropagated plants was 88.9%. Additionally, accumulation of scopoletin, a phytoalexin, was revealed for the first time in the uninfected leaves of Spilanthes. Further, the quantitative estimation by HPLC with a fluorescence detector showed that the amounts of scopoletin content (0.10 lg g -1 DW) in the leaves of micropropagated plants are comparable to those of field-grown mother plants. The study thus signifies the effectiveness of in vitro methodology for true-to-type plant regeneration of Spilanthes and their later utility for biosynthesis and constant production of scopoletin throughout the year.

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Section: Discussionsupporting

confidence: 69%

Improved clonal propagation of Spilanthes acmella Murr. for production of scopoletin

Singh

Chaturvedi

2010

Plant Cell Tiss Organ Cult

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“…16.0) and significance variation between the concentrations was studied using DMRT at 0.05 % level regenerative which were collected in the months of June to September. The seasonal effect on culture establishment has also been reported in Azadirachta indica (Arora et al 2010),…”

Section: Resultsmentioning

confidence: 66%

In vitro propagation, micromorphological studies and ex vitro rooting of cannon ball tree (Couroupita guianensis aubl.): a multipurpose threatened species

Shekhawat¹,

Manokari²

2016

Physiol Mol Biol Plants

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“…Therefore, it is imperative to use plant tissue culture methods. Micropropagation of neem tree through tissue culture has been reported by employing nodal stem segments (Chaturvedi et al 2004b, Arora et al 2010) and leaf segments (Ramesh and Padhya 1990, Arora et al 2009). Here, we report an in vitro method employing root explants for cloning A. indica leading to production of true-to-type plantlets.…”

mentioning

confidence: 99%

“…were produced employing nodal stem segments of a 40-year-old tree following the protocol reported earlier by Arora et al (2010). Roots measuring about 3 cm in length, excised from in vitro-grown plantlets were cultured in liquid medium which is a modification of Murashige and Skoog (1962;MS) , Na 2 SO 4 (0.70 mM), K 2 SO 4 (0.57 mM), L-arginine-HCl (47.45 µM), L-glutamine (171.06 µM), ascorbic acid (141.95 µM) and removing m-inositol.…”

mentioning

confidence: 99%

In vitro cloning of Azadirachta indica from root explants

Arora¹,

Sharma²,

Srivastava³

et al. 2011

Biol. plant.

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In vitro cultures of Azadirachta indica A. Juss. were raised by first culturing the root segments on modified Murashige and Skoog (MS) medium supplemented with 8.88 µM 6-benzylaminopurine (BAP), 9.84 µM N 6 -(2-isopentenyl) adenine (2iP), 5.71 µM indole-3-acetic acid (IAA), 81.43 µM adenine hemisulphate and 2.27 µM putrescine for 2 d followed by their transfer to the same medium except containing one-tenth of the initially used concentrations of BAP, 2iP and IAA. The regenerated shoots sustained proliferation in the basal medium supplemented with 1.11 µM BAP, 1.43 µM IAA and 135.72 µM adenine hemisulphate. The isolated shoots were rooted to produce plantlets in the presence of 2.46 µM indole-3-butyric acid (IBA). The plantlets showed uniform luxuriant growth under field conditions. True-to-type nature of the field-grown root-regenerated plants was ascertained by random amplified polymorphic DNA (RAPD) analysis.Additional key words: auxins, cytokinins, genetic stability, neem, RAPD, root-regenerated plants.⎯⎯⎯⎯ Azadirachta indica A. Juss. (family Meliaceae), popularly known as neem, is a multipurpose tree which has acquired significance for its commercial uses in pharmaceutical and agro-industries. The main limitations in sexual propagation are the recalcitrant nature of seeds with short span of viability and high heterozygosity (Ezumah 1986, Sacande et al. 2001. Vegetative propagation by cuttings, however, has constraints due to problems of rooting, availability of cuttings of right maturity, particular season of the year and presence of disease causing organisms within the material (Dogra and Thapliyal 1996). Therefore, it is imperative to use plant tissue culture methods. Micropropagation of neem tree through tissue culture has been reported by employing nodal stem segments (Chaturvedi et al. 2004b, Arora et al. 2010) and leaf segments (Ramesh and Padhya 1990, Arora et al. 2009). Here, we report an in vitro method employing root explants for cloning A. indica leading to production of true-to-type plantlets. The fidelity of the method has been proved by carrying out random amplified polymorphic DNA (RAPD) analysis of mother tree as well as of the plants regenerated through root segments.Plantlets of Azadirachta indica A. Juss. were produced employing nodal stem segments of a 40-year-old tree following the protocol reported earlier by Arora et al. (2010). Roots measuring about 3 cm in length, excised from in vitro-grown plantlets were cultured in liquid medium which is a modification of Murashige and Skoog (1962; MS) medium except containing 6.25 mM NH 4 NO 3 , 4.95 mM KNO 3 , 1.36 mM CaCl 2 . 2 H 2 O, 1.10 mM KH 2 PO 4 , 1.83 mM MgSO 4 .7 H 2 O, 0.49 µM pyridoxine-HCl, 39.96 µM glycine, 2 % sucrose and addition of 0.35 mM Na 2 SO 4 , 0.76 mM (NH 4 ) 2 SO 4 , 23.73 µM L-arginine, 68.42 µM L-glutamine, 63.45 µM cysteine-HCl, 0.2 µM D-biotin, 0.13 µM riboflavin, 56.77 µM ascorbic acid and removing m-inositol, supplemented with 0.05 µM indole-3-butyric acid (IBA) and ⎯⎯⎯⎯

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Rapid in vitro cloning of a 40-year-old tree of Azadirachta indica A. Juss. (Neem) employing nodal stem segments

Cited by 33 publications

References 24 publications

Improved clonal propagation of Spilanthes acmella Murr. for production of scopoletin

Improved clonal propagation of Spilanthes acmella Murr. for production of scopoletin

In vitro propagation, micromorphological studies and ex vitro rooting of cannon ball tree (Couroupita guianensis aubl.): a multipurpose threatened species

In vitro cloning of Azadirachta indica from root explants

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