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DrAbbasyadollahi     تاريخ انتشار : 2December 2014
Effects of antimicrobial activity of silver nanoparticles on in vitro establishment of G · N15 (hybrid of almond · peach) rootstock
In the present investigation were evaluated the antifungal and antibacterial activities of Nano-silver (NS). Two separate experiments were done to evaluate the potential of silver nanoparticles in controlling the contamination of G · N15 micro-propagation. In the first experiment, a factorial experiment based on a completely randomized design with 15 treatments including five different NS concentrations (0, 50, 100, 150 and 200 ppm) and three soaking time of explants (3, 5 and 7 min) with f

Introduction
Preventing or avoiding microbial contamination of planttissue cultures is critical to successful micropropagationEpiphytic and endophyticorganismscancauseseverelossestomicropropagated plants at each stage of growth [3,6Bacterial contaminants are often difficult to detect because
they remain mostly within the plant tissue [34]. Contaminatedplants may have no visible symptoms, reduce multiplicationand rooting rates, or may die [6G · N15 (Garnem) is a hybrid of Prunus amygdalus(Garfi) · Prunus persica (Nemared). Garnem is the most commonlyused rootstock for almond and peach and has exhibitedtotal graft compatibility with the whole range of peach andalmond cultivars as well as some plum and apricot cultivars[4]. Other appropriate attributes of garnem include good vigorand resistance to root-knot nematodes, adaptation to calcareoussoils and other Mediterranean agroecological conditions[37,47,18]. This rootstock is also compatible with some diploidplums (Prunus salicina Lindl. and related plums) such as ‘SantaRosa’ and ‘Golden Japan’. Compatibility has also beenobserved with some apricot cultivars belonging to the morecompatible apricot group such as ‘Paviot’ but not with theapricot cultivars of the more exigent group such as ‘MoniquıPattern be used foralmond, peach and Japanese plumirrigated and for rainfed almond by showing good overall
compatibility with the varieties of these species [4]. It hasshown a good propagation capacity through the differentsystem in use. ‘Garnem’ is propagated well by hardwood andherbaceous cuttings in aerated and well-drained soils [15The commercialization oftraditionalpeach,nectarine andalmond orchards is considered of great importance in orderto enhance yield; and to achieve this objective the micropropagationof G · N15 as a vigorous rootstock through tissue
culture in a high scale and short period of time is essential forhigh density peach, nectarine and almondorchards.Eliminatingorreducingexogenousandendogenouscontaminatingmicroorganisms is one of the most important steps of a successful micropropagation process
  Materials and methods

lant material and culture initiationPlant material was obtained from 2-year-old G·N15or‘Garnem’ vegetative rootstock grown at the greenhouse ofthe Tarbiat Modares University (TMU), Tehran, Iran. Shootswith length of 15–20 cm of ‘Garnem’ were collected and transferredto the fruit tree micro-propagation laboratory. First ofall, shoots were excised into 1.5–2.5 cm-long nodal segmentsand then for surface disinfection, the explants were agitatedfor 5 min insolution water and dishwashing liquid containing5 drops of Tween-20 in 100 ml of water, finallytheywere
washed underrunningtapwaterfor1htoremovesurfacecontaminationAfterward,theyweresoaked in 0.2 % (w/v) benomyl(fungicide) for half an hour then washed thoroughly withsterile distilled water. After the above steps, in order to internaldisinfection two separate experiments were conducted: in thefirst experiment, the explants were pre-sterilized by immersionin 70%ethanol for 60 s followed by a rinse with sterile distilledwater. These pre-sterilized explants were exposed in fiveconcentrations of NS solution (0, 50, 100, 150 and 200 ppmwiththreedifferentimmersion times (3, 5 and 7 min).Theninordertoreducephenoliccontaminants,theexplantsweresubmerged in 0.7% citric acid 2 times, each time 3 min. Finallythey were cultured in Murashige and Skoog [44] (MS) mediumsupplemented with 0.25 mg L1 BAP, 30 g L1sucroseand8gL1agarAtthesame time and in another experiment, the explantswere treated with 2.5% (v/v)sodiumhypochlorite for 4 minThen, the explants were washed three times with sterilizeddistilled water to remove all the traces of sodium hypochloriteAfter this stage, explants were dipped in 0.7% citric acid 2
times, each time 3 min. Finally they were cultured in MS mediumsupplemented with various concentrations of NS (0, 50100, 150 and 200 ppm), 0.25 mg L1 BAP, 30 g L1 sucroseand 8 g L1 agarThe pH of all media was adjusted to 5.8 before autoclavingand then grown in growth room with light intensity of 25003000 lux, photoperiod of 16/8 h light/dark, relative humidity
of 45% and constant temperature of 25± 1 C. McCarthyglasses were usedfortheestablishment stage of explants. After1 week, fungal contamination percent, bacterial contaminationpercent and viability percent of G · N15 buds were recordedeach day. Viability index was obtained by calculating thepercentage of regenerated explants without contamination(fungal or bacterial

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