International Journal of Horticulture, 2017, Vol.7, No.18, 146-153
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agriculture. In tissue culture techniques, especially the micropropagation technique is very important and
effective technique which can produce thousands of vegetatively propagated plants in less time and by using less
space. Micropropagation technique facilitates the production, multiplication and maintenance of disease free
potato clones (Venkatasalam et al.
,
2013).
Potato crop is prone to stresses by different biotic and abiotic factors and among these; frost is a very important
factor. Frost tolerant germplasm of potato crop was imported but now in Pakistan, different research institutes are
working on this aspect. A number of frost tolerant genotypes have been approved. Hence, this proposed research
study was aimed at the establishment and optimization of an efficient, expedite and genotype-independent
micropropagation system in selected genotypes of potato to produce healthy, disease free material in short time
and less space for the farmers. For this research study, axially buds of sprouts were used as explants. The explants
were induced into plantlets on tissue culture regime containing different growth regulators at different levels and
even in different combinations. However in this study effect of best micropropagation medium on secondary
metabolites of in vitro micropropagated shoots was also assessed and estimated.
2 Material and Methods
Germplasm to be used:
Two local genotypes of potato (
Solanum tuberosum
L.) viz., PRI-Red and FD 69-1 were
used for this research study.
2.1 Explant preparation
a)
Sprouting of Potato Tubers:
For expedite sprouting, potatoes of both genotypes were incubated under
dark and cold regimes. Potatoes were washed with ultra pure water and kept in cold condition at 4˚C for
5 days and subsequent storage in dark condition at 26±1˚C for one month.
b)
Sterilization of Explants:
Surface sterilization of explants (Axillary bud of sprout) was done by using
different detergents and antiseptic solutions.
Shoots induction/in vitro micropropagation:
Eight micropropagation media (Table 1) were used in this
experiment. These media were investigated for efficient and genotype independent in vitro micropropagation
system in
Solanum tuberosum
L. The explants were induced into plantlets on tissue culture regime containing
NAA and BAP growth regulators at different levels and even in different combinations. For culturing, sprouts
were cut in such a way that each cutting must contains 1 or 2 buds. All cultures for in vitro micropropagation
were kept and maintained under 16 hours photoperiod at 26±1˚C.
Table 1 Micropropagation media (MPm)
MPm1 MPm2
MPm 3
MPm 4
MPm 5
MPm 6
MPm 7
MPm 8
NAA
0 mg/l
0.75 mg/l
1.75 mg/l
2.75 mg/l
0 mg/l
0 mg/l
0 mg/l
2.75 mg/l
BAP
0 mg/l
0 mg/l
0 mg/l
0 mg/l
0.75 mg/l
1.75 mg/l
2.75 mg/l
2.75 mg/l
Nutrient spp. MSN* MSN*
MSN*
MSN*
MSN*
MSN*
MSN*
MSN*
Note: MSN* (MS salt = 4.33 g/l, Sucrose = 30 g/l, Myoinositol = 0.1 g/l, Nicotinic acid = 1 g/l, Pyridoxine HCl = 1 g/l, Thymine
HCl = 2 g/l, Glycine = 4g/l, Gellan gum powder = 2.66 g/l)
Rooting and acclimatization of micropropagated shoots:
For the induction of healthy, strong and profuse rooting,
micropropagated shoots were transferred to rooting medium containing basal MS salt (M524, PhytoTechnology
Laboratories®, USA), 3% sucrose, vitamins, devoid of growth regulator (s) and 0.266 % Gellan gum powder for
solidification of medium. Rooted shoots or plantlets were shifted to pots containing Belgium compost and
wrapped with polythene bag to maintain humidifying condition at 26±1˚C. These plantlets were then slowly
acclimatized to lower humidity prior to green house transfer.
Experimental layout and statistical analysis:
The data was collected as the number of shoots per explant and the
days to growth initiation after culturing. The experiment was conducted in completely randomized design (CRD).
