ЭлБиб

PLANT GENETIC TRANSFORMATION, CELL SELECTION AND SOMATIC HYBRIDIZATION

DEVELOPMENTOFMETHODSFORSHOOTREGENERATIONAND

GENETICTRANSFORMATION OF

RHAPONTICUM CARTHAMOIDES

PLANTS

I.V.Orlova, *E.G.Semenujk,N.S.Zacharchenko, Ya.I.Buryanov

Branch ofShemyakin andOvchinnikovInstitute ofBioorganic Chemistry

*PushchinoState University, Pushchino, Moscow region, 142292, E-Mail:

orlova@fibkh.serpukhov.su

An efficient method for shoot regeneration from leaf disks and

Agrobacterium

mediated transformation of an important medicinal plant

Rhaponticum ca rth am o id es

, capable of producing, ecdysteroids has been

developed.

The leaf explants of

Rhaponticum ca rth am o id es

were precultivated on

Murashige and Skoog (MS) basal medium supplemented w ith

benzyladenine (BAP) (1 mg/l) and 2,4 dichlorophenoxyacetic acid (2,4 D)

(1 mg/l) and were subcultivated on MS with BAP (0,5 mg/l) and indol - 3-

acetic acid (IAA) (0,5 mg/l).

Callus formation was observed after 17-19 days and shoot regeneration

started after 22-25 days. The efficiency of regeneration obtained was 47-

57%. Isolated shoots were proliferated and rooted on MS without hormones.

The of

Agrobacterium tumefaciens

strains employed inthe transformation

experiments were:

A. tum efacien s

LBA 4404 (pAL4404) and LBA 4301(pUCD2614)

harbouring binary vector pGA482::M.EcoRII with bacterial methyiase gene

under control of 35S CaMV-promoter.

2. A

tum efacien s

3101 (pMP90RK) harbouring binary vector pPCV002

with

rol C

gene under control of 35S CaMV- promoter.

In all cases selection for stable transformants marker gene npt II

(neomycin phosphotransferase) was used.

Kanamycin resistant shoots formed from disks growing on selected

medium containing 50 mg/l kanamycin will be analysed by Southern

hybridization.

The influence of

rol C

and bacterial methyiase genes expression on

secondary metabolism of transformed plants will be studied.

2 9 3

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