Studies of rooting ability in the tobacco rac mutant impaired in rooting

• The rolB gene from Agrobacterium rhizogenes has proven to significantly stimulate adventitious rooting in several species. The rac mutant of Nicotiana tabacum L. ‘Xanthii’ is impaired in rooting. The objective of the present study was to explore if the rolB gene introduced into rac mutant could restore the rooting ability. Leaves of the rac and wild-type (WT) tobacco shoots were transformed by Agrobacterium strain C58C1 holding the binary vector pCMB-B:GUS harboring nptII, rolB and gus genes was used for transformation. Four transformed lines were obtained expressing the rolB gene. However, the rooting ability was not restored. The rac can only be propagated in vitro due to lack of roots. To obtain seeds from rac for further studies on rooting we grafted rac shoots onto WT seedlings in vitro. The successful grafted plants were then transferred to pots in the greenhouse to obtained seeds by selfing. The grafted plants flowered normally but were weaker with smaller leaves. Fruit setting and seed production were poor in the rac mutans. In vitro rooting showed that the roots from WT seedlings penetrated the seed coat after 3 days. After 6 days elongated roots were covered with root hair. Roots from the grafted rac seeds penetrated seed coat first after 6 days and at day eight root growth ceased. The root tip was swollen and covered with minor root hair. Since it has been shown that the root tip in the rac contains 14 times more auxin than WT we were interested to investigate if the rac was impaired in some PIN genes responsible for lateral auxin transport from the root meristem. We performed Western blot analysis with various PIN antibodies using the protein extracts from petioles and leaves of WT and rac. Both PIN 2 and 3 gave good signals. Next will be immunolocalization of PINs in the root.

Ämnesord

LANTBRUKSVETENSKAPER  -- Bioteknologi med applikationer på växter och djur -- Växtbioteknologi (hsv//swe)

AGRICULTURAL SCIENCES  -- Agricultural Biotechnology -- Plant Biotechnology (hsv//eng)

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