PLANT TISSUE CULTURE

MADE BY VISWANATHAN ROHIT MSc BIOTECHNOLOGY

CONTENTS
INTRODUCTION

PREPARATION OF MEDIA
SUB-CULTURED SEED-CULTURED

HARDENING
CONCLUSION

INTRODUCTION
Broadly refers to technique of growing plant cells,

tissues,organs, seeds or other plant parts in a sterile environment on a nutrient medium The first commercial use of plant propagation on artificial media was in the germination and growth of orchid plants in the 1920s It was only after the development of a reliable artificial medium by Murashige & Skoog in 1962 that plant tissue culture really took off commercially

MURASHIGE & SKOOG MEDIA

COMPONENT

AMOUNT mg/L 1650 1900 440 370 170 0.83 6.2 22.3 8.6 0.25 0.025 0.025 27.8 37.3 100 0.5 0.5 0.1 2

STOCK SOLUTION mg/L

NHNO KNO CaCl.2HO MgSO.7HO KHPO KI HBO MnSO.4HO ZnSO.7HO NaMoO.2HO CuSO.5HO CoCl.5HO FeSO.7HO Na.EDTA MESO-INOSITOL NICOTINIC ACID PYRIDOXINE HCl THIAMINE HCl GLYCINE

STOCK SOLUTION mg/500ml MACRO SALTS 33000 38000 8800 7400 3400 MICRO SALTS 166 1240 4460 1720 50 5 5 Na SALTS (III) 5560 7460 VITAMINS 20000 100 100 20 400

WORKING SOLUTION ml/L 500ml 16500 19000 4400 50ml 3700 1700 83 620 2230 860 5ml 25 2.5 2.5 2780 5ml 3730 10000 50 50 5ml 10 200

250ml

25ml

12.5ml

2.5ml

1.25ml

2.5ml

1.25ml

2.5ml

1.25ml

SUCROSE AGAR pH

30g 8g 5.8

15g 4g 5.8

7.5g 2g 5.8

Preparation of media
MS basal + 0.1% A.C. ii. MS basal iii. MS + 0.25 mg /l NAA (alpha- naphthalene acetic acid) + 0.2 mg/l BAP (benzyl amino purine).
i.

Sub-cultured
Two bottles of Cymbidium Soul hunt and one bottle of Sleeping nymph is cultured in MS basal + 0.1% A.C. ii. One bottle each of Lunavian atlas and Pine clash in MS basal + 0.1% A.C. iii. one bottle each of Kalanchoe pinnata and Anoectochilus are subculture in MS basal. iv. One bottle of Citrus medica is sub-cultured in MS basal + 0.25 mg/l NAA + 0.2 mg/l BA.
i.

Seed-cultured
Cymbidium mastersii is cultured in MS basal.

Citrus marcoptera is cultured in MS basal .

Hardening
Taking out the plant from the bottle

by forceps. Cleaning the plants from media with tap water and then deep into Bavistin for about 15 mins. 1:1 of charcoal and brick are collected in a pot with small pores in it. Plant is then taken out for Bavistin after 15 mins and planted . Then plastic is covered over a plant and after that it is taken to hardening room. Sleeping nymph.

CONCLUSION
Plant tissue culture is a successful technique for

growing endangered species & hybrids so that in the future there will be enough food for the generations to come & saving the ornamental plants.