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Validation of micropropagation protocols of pineapple (Ananas comosus L.) with clonal fidelity analysis
Pineapple (Ananas comosus L.) is the third most important tropical fruit in the world with a production share of 20 per cent (Kole, 2007). It is a commercial fruit crop of India and is cultivated in Assam, Meghalaya, Kerala, Tripura, Manipur, West Bengal, Karnataka and Goa. In commercial cultivation...
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| Format: | Ph.D Thesis |
| Jezik: | English |
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Vellanikkara
Centre for plant biotechnology and molecular biology, College of horticulture
2015
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| Teme: |
| Sažetak: | Pineapple (Ananas comosus L.) is the third most important tropical fruit in
the world with a production share of 20 per cent (Kole, 2007). It is a commercial
fruit crop of India and is cultivated in Assam, Meghalaya, Kerala, Tripura,
Manipur, West Bengal, Karnataka and Goa. In commercial cultivation per hectare,
number of planting material requirement is 40,400. Conventionally, pineapple is
propagated by slips, suckers and crowns and the propagules available from these
materials are limited especially when new varieties are released.
Micropropagation technique offers an opportunity for large scale production of
uniform pineapple planting material in a relatively short period of time (Escalona
et al., 1999; Ika and Mariska, 2003). Micropropagation protocols in pineapple
have been reported; and suitability of the same for clonal propagation has to be
ascertained. So, the present investigation on “Validation of micropropagation
protocols of pineapple (Ananas comosus L.) with clonal fidelity analysis” was
taken up at the Centre for Plant Biotechnology and Molecular Biology, College of
Horticulture from 2013 to 2015.
Commercial variety of Kerala, Mauritius, was used for the study. Four
micropropagation protocols: P1 (Valsala et al. 2010), P2 (protocol available at
CPBMB), P3 (Jose, 1996), and P4 (Mhatre, 2007) were selected to regenerate
plants and for clonal fidelity analysis. The molecular marker used for clonal
fidelity analysis was Inter Simple Sequence Repeats (ISSR).
Explants used for the study were stem segment/shoot tip. The treatment,
T1 involving 30 minutes carbendazim, 5 minutes 70 per cent ethanol and 10
minutes 0.1 per cent mercury chloride was found to be the best recording
maximum survival of 40 per cent and minimum contamination rate of 32.22 per
cent. Cultures were in vitro established adopting the procedures in three protocols
(P1, P2, and P3) and the ideal medium was MS supplemented with 4 mg/l BA+ 3
per cent sucrose+ 0.7 per cent agar + 250mg/l cefotaxime reported by Jose (1996)
which recorded minimum days of 14 for shoot initiation and maximum shoot per
culture of 2.25.
Shoot proliferation, elongation and rooting of plantlets were also observed
in P1, P2, and P3 protocols. The P3 protocol recorded highest shoot proliferation
rate of 51.25 at SC5 passage in the medium MS+1mg/l BA+ 0.5mg/l NAA. In P1
and P2 protocols, callusing was observed during shoot proliferation. Shoot
elongation and rooting were found best in P1 protocol in the medium MS+2mg/l
NAA recording maximum mean length of shoots of 8.97cm and mean number of
roots of 4.95. Potting medium vermiculite: coir peat: soil rite at the ratio of 2:1:1
was found to be the best for ex vitro establishment.
Clonal fidelity studies using ISSR assay were carried out with the mother
plants along with 14 regenerants. Eight pineapples specific ISSR primers tested.
In ISSR assay, P1 protocol recorded the highest average polymorphism of 30.14
per cent followed by P2 with 23.20 per cent and P3 recorded the least average
polymorphism of 12.64 per cent. Using NTSYS software, similarity coefficients
between mother plants and 14 regenerants of P1, P2 and P3 protocols were 0.73,
0.83 and 0.90 respectively.
P3 protocol (Jose, 1996) is identified as the ideal for micropropagation.
The P1 protocol (Valsala et al. 2010) can be used for exploitation of somaclonal
variation for crop improvement. For conclusive results, the regenerants have to be
evaluated along with mother plants for phenotypic expression. P3 protocol
regenerants could be advanced to SC8 passage and clonal fidelity should be
checked. |
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| Opis: | 91 Pages |