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15 views15 pages

Shiv,,,,, 1

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shortswallahedx
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ANAND AGRICULTURE UNIVERSITY

VASO-387380

Seed Production Technology Department of Genetics & /Plant


Tissue Culture Technology
(ELP 8.2)

Submitted By : Patel Shivanikumari Prakashbhai


Roll No. : 38
Reg. No. : 30-1022-0043

Submitted To:
Ms. Nisha B Patel
MICROPROPAGATION OF
SANDALWOOD (SANTALUM ALBUM L.)
WHAT IS MICRO-PROPAGATION

Micropropagation is the artificial process of producing


plants vegetatively through tissue culture or cell culture
techniques. In this artificial process of propagation, plants
are produced invitro by asexual means of reproduction or by
vegetative propagation.
INTRODUCTION
 Sandalwood (Santalum album L.) is a commercially and
culturally important plant species belonging the family
santalaceae.
 The world's second-most expensive wood is this one.

 One of the best natural carving materials is the tree's


heartwood, which is prized for its scent.
 Pharmaceuticals, cosmetics, aromatherapy, and
fragrances all use sandalwood oil.
 Conventional methods of asexual propagation like
grafting, budding, layering etc. for many of the plants and
trees are often too slow or fail completely.
 In vitro regeneration techniques can be used to encounter
difficulties of traditional vegetative propagation methods
of superior clones.
 In vitro propagation of sandalwood was attempted as early
as 1963. Induction of callus from mature endosperm on
modified White's medium was reported, but the callus did
not proliferate further (Rangaswamy and Rao, 1963).
 Various explants such as embryo, hypocotyls, shoot tip,
nodal segment and cell suspension cultures with varying
degree of success have been used for rapid multiplication
of sandal trees.
 There is only one published report on shoot bud formation
directly from in vitro cultured leaves for sandal (Mujib,
2005).
 In woody species relatively little information is available
on shoots formed directly on leaves without a callus
phase (Preece et al., 1993).
 Leaves obtainedfrom in vitro grown plants provide a
useful source of explants which eliminate the risk of
contamination.
 Micropropagation using tissue culture allows much
greater control and manipulation of the development of
tissues within the culture tube than conventional
methods.
 The aim of the present study is to develop a potential
system of in vitro regeneration of the highly valued tree
Santalum album.
INITIATION
 PLANT MATERIALS (EXPLANT)
 Nodal segments and leaves were collected from selected
sandalwood.
 The explants (nodes) of 2.0-2.5 cm in length and first and
second leaves of tender shoots were dissected out from the
mother plant and collected in sterile distilled water and brought
to the lab.
 A slant cut was given at the base of the nodal explants to
expose more surface area for better transport of nutrients.
 The explants washed with distilled water and then disinfected
with Tween 20 solution (0.1%) for 5-10 minutes and washed
with distilled water.
 After an hour, the explants were treated with 1% Bavistin for
10 min in horizontal shaker and surface sterilized in 0.1%
HgCl2 for 1 min.
 MEDIA PREPARATION

 1. Assemble all chemicals in work area


before beginning.
 2. Accurately weight the media base
calculated "powder" by electronic
balance.
 3. Add the powder into flask.
 4. Add distilled water to the flask, for
making the correct volume.
 5. Heat & stir (agar will be burn if is
not stirred) until all of the ingredients
go into solution. When the media
boils, it is ready for sterilization.
 6. For sterilization, autoclave should
be used, test tube top should be
covered by caps to prevent
contamination.
 INCUBATION

• The explants were inoculated in MS basal medium supplemented with


various concentrations and combinations of BAP and 2ip. The basal MS
medium served as control.
• Inocubation done in proper sterile area(Laminar air flow).
•The cultures were exposed to a light (approximately 2000 lux) and
darkness cycle of 16 hours and 8 hours respectively.
• The culture room temperature was maintained at 25 ± 2°C.
 MULTIPLICATION
 The highest shoot multiplication was achieved on MS
medium containing 1.0 mg L-1 (21P) showed better
growth response and produced 41.0±2.0 shootlets with an
average length of 2.90±0.10 cm after 45 days of culture.
The cluster of shoots were cultured on same media with
10% coconut Milk (CoM) showed shoot elongation up to
4.6 cm.
ROOTING
 After completion of shoot multiplication and elongation
cycle, the elongated shoots transfer to half strength MS
medium supplemented with 0.5 mg L-1 IBA and 0.25 mg
L-¹ NAA produced 4.2 ± 0.10 roots with an average
height of 4.8±0.2 cm after six weeks in culture.
HARDENING
Hardening is lab to land transfer
technology. It provides healthy growth of
the plant under milder conditions before
the plant exposed to environmental
Two type of hardening
1. Primary hardening:
The In vitro Plantlets raised after pre-
hardening were removed from culture jars,
washed thoroughly under disttiled water
remove traces of medium from the roots.
Plantlets were transferred to hardening
trays. One set of hardening traywas consist
of 3:1 ratio of coco peat and
vermicompost.
2. Secondary hardening:
Afterwards the plants are
transferred to polybags filled with
potting mixture and grown under
shaded house for 6-8 weeks. This is
called Secondary hardening. After
secondary hardening the plants are
suitable for growing in farmer's
fields.

 The rooted plantlets were


transferred for hardening 70% of
plantlets survived and resumed
growth in the mixture of soil,
vermiculite and farm yard manure
(1:1:1).
WHOLE PROCESS
THANK YOU

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