INVITRO CULTURE OF MENTHA PIPERITA L. THROUGH SHOOT TIP AND SINGLE NODE CULTURE

Roshani Rajbanshi

OUTLINE

Geographical location of Nepal Vegetation of Nepal Introduction of Mentha piperita L. Uses of Mentha piperita L. Objectives Methods Material Result Conclusion Acknowledgement References

INTRODUCTION OF NEPAL

INTRODUCTION

Arid Tibetan highland in the north (towards China)

Gangetic plains in the south (towards India)

Elevation 75m-8,848m Climate Subtropical to

Arctic

VEGETATION OF NEPAL

6500 species flowering plants 4% of flowering plants Endemic to

country 4000 species non-flowering plants 450 spp. Pteridophytes 853 spp. Bryophytes 460 spp. Lichens 687 spp. Algae 1600 spp. Fungi(Source: Bentham & Hooker (1973)

MEDICINAL PLANTS OF NEPAL

700 species are recorded as medicinal plants (Chopra et.al. 1885).

In 1999, Tiwari reported 1463 species of medicinal plants in Nepal.

50 items of crude herbs and aromatic plants are sold to India and China (HMG, 2000).

INTRODUCTION OF MENTHA L.

Nepali name is “Pudina” Found in northern hemisphere Flowering plant Family Labiatae Herbaceous, aromatic and medicinal plant Mentha L. 40 species. (HMG, 2000) M. piperita hybrid of M. spicata and M.

aquatica.

USES

Peppermint Aromatic, stimulant Used to treat stomachache, allaying nausea,

flatulence and vomiting. A good source of peppermint oil Peppermint oil is used in pharmaceuticals,

dental preparation, mouthwashes, cough drops, soaps, chewing gums and candies.

Also used in “Chatney” in South Asia.

OBJECTIVES

To establish in-vitro culture in different hormone concentration.

To observe the response in different hormone concentration.

To find the variation in single node culture and shoot tip culture.

METHODOLOGY Murashinge and Skoog (1962) medium was prepared

with the following nutrients Macro nutrients MgSO4.7H2O, KH2PO4, KNO3,

NH4NO3, CaCl2.2H2O

Micro nutrients H3BO3, MnSO4.4H2O, ZnSO4.7H2O, NaMoO4.2H2O, CuSO4.5H2O, COCl2.6H2O, KI

Iron Source FeSO4.7H2O, NaEDTA.2H2O Vitamins Thiamin HCl, Pyridoxin HCl, Nicotinic

acid, Myoinositol, Glycin Carbon source Sucrose Hormones Naphthalene acetic acid(NAA) and Benzyl

amino purine (BAP) 0.8 % Difco-facto agar solidification

MATERIAL—MENTHA PIPERITA L.

Sterilized in running water for 1 hour, 1% sodium hypochloride for 5 minutes, 70% ethanol for 1 minute, rinsed in distilled water for 3 times

Single nodes and shoot tip of 4-5 mm pieces were cut

Explants were inoculated in culture tubes inside laminar air flow, sealed and transferred to incubation room

Temperature +/- 25oC Observation 4-8 weeks Subculture of plantlets were done in different

hormones concentration.

RESULT

Invitro Culture of Single Node

MS = Multiple Shoot RMS = Rooted Multiple Shoot

NAA (ppm)\BAP (ppm) 0 1 2

04/4-RMS

2/2-Single Shoot 2/2-Single Shoot

11/4-plantlet 3/4-RMS 2/2-MS

1/2 -shoot 1/2 -poor growth

RESULT

Invitro culture of Shoot tip

MS = Multiple Shoot RMS = Rooted Multiple Shoot

NAA (ppm)\BAP (ppm)

0 1 2

0 2/2-Plantlet 1/2 – MS 1/2 - MS1

1/2-plantlet1/2-Callus1/2 –Plantlet 2/2- MS

RESULT

Subculture

RMS= Rooted Multiple Shoot MS= Multiple Shoot

NAA (ppm)\BAP (ppm)

O ppmNode

0 ppmShoot Tip

1ppmNode

1ppmShoot Tip

2 ppmNode

2 ppmShoot Tip

0 2/2-RMS

2/2-Plantlet

2/2-Plantlet

2/2 RMS

2/2-MS 1/2-RMS1/2-MS

1 1/2-Plantlet

1/2-Plantlet

1/2 – RMS

1/2 –MS

2/2-MS 1/2-Callus1/2-Plantlet

CALLUS FORMATION

Callus obtained after 4 weeks from Shoot tip culture

8 WEEKS OLD PLANTLET FROM SHOOT TIP CULTURE

Plantlet

4 WEEKS OLD MULTIPLE SHOOT FROM NODE CULTURE

VIGOROUS GROWTH OF MULTIPLE SHOOT AFTER 8 WEEKS OF SHOOT TIP IN MS+BAP+NAA

CONCLUSION

Both single node and shoot tip can be cultured artificially.

Basal media is the best media to obtain plantlet and rooted multiple shoot.

In few cases, addition of NAA suppressed the proliferation of multiple shoot.

The basal media supplemented with BAP (2ppm) and NAA (1ppm) gave high number of multiple shoot.

ACKNOWLEDGEMENT

Mukti Ram Aryal Sanjeev Kumar Rai Shankar Pahari

REFERENCES

Bentham, G. & Hooker, J.D. (1973). “Genera Plantarum” Vol II. Reeve & Co. London.

Anonymus. (2000). National Register of Medicinal Plants. HMG Nepal/ IUCN

THANK YOU