10.1071/FP06224_ AC Accessory Publication: Functional Plant Biology, 2007, 34(4), 368-381.
© CSIRO 2007
Functional Plant Biology, 2007, 34, 368–381
ISSN 1445-4408
Isolation of dehydration-responsive genes in a drought tolerant common bean cultivar and expression of a group 3 late embryogenesis abundant mRNA in tolerant and susceptible bean cultivars A ´ M. Pena-Castro ˜ Blanca E. Barrera-FigueroaA , Julian , Jorge A. Acosta-GallegosB , A,C ´ Roberto Ruiz-MedranoA and Beatriz Xoconostle-Cazares A
´ y de Estudios Avanzados del Instituto Departamento de Biotecnolog´ıa y Bioingenier´ıa, Centro de Investigacion Polit´ecnico Nacional, Av. IPN 2508, 07360 San Pedro Zacatenco, M´exico. B Programa de Mejoramiento del Frijol, Instituto Nacional de Investigaciones Forestales, Agr´ıcolas y Pecuarias. Campo Experimental Baj´ıo. Km. 6.5 carretera Celaya – San Miguel de Allende, 38010 Celaya, Guanajuato, M´exico. C Corresponding author. Email:
[email protected]
Supplementary material
Rehydration Control
a
Drought
Drought
b
1h 2h 4 h 24 h Time
IRWC 82 83 96 84 88 89 79 70 64 55 45 47
IRWC 1.7 Kb
PvLEA3
PvLEA3 rRNA
rRNA
c IRWC
l ca l l ni e rt o inity ha ag rt o t d l ec am on on ea ol C Sa M d C H C 99 96 99 96 89 96
49 50 53 49 71 86 69 76
ht
l
tro
on
d
C
A AB
ug
ro
8h 24h 8h 24h 8h 24h
IRWC 80 80 85 80 45 30
af
Le
e
D
Time
t
oo
R
em
St
C D C D C D IRWC 84 55 84 55 84 55
PvLEA3
PvLEA3
Pv LEA3
rRNA
rRNA
rRNA
Fig. S1. Northern blot analysis showing the differential accumulation of PvLEA3 transcript in Pinto Villa roots. PvLEA3 was accumulated in roots by progressive drought stress (a) and quickly downregulated by rehydration (b). Cold (4◦ C for 2 h) and salt (250 mM NaCl for 2 h) stress, but not heat stress (42◦ C for 2 h) or mechanical damage induced the expression of PvLEA3 in Pinto Villa roots (c). Induced expression of PvLEA3 in roots by exogenous ABA treatment (100 µM ABA for 8 h and 24 h) (d). Accumulation of PvLEA3 was not restricted to roots; it was also differentially accumulated in leaves and stem, but at lower levels (e). For each case, 20 µg of total RNA were loaded per lane, blotted onto nylon membranes and hybridised with a radiolabelled PvLEA3 probe. Ethidium bromide staining of ribosomal RNA (rRNA) is shown to monitor equal loading of total RNA. lRWC, Leaf relative water content (%); C, Control; D, Drought.
f
e
d
h
c
g
Fig. S2. In situ hybridisation assays of PvLEA3 in Pinto Villa plants. Panel a shows indigo mRNA-associated signal of PvLEA3 transcript in the phloem area of stem, particularly in both developing and functional phloem cells. In roots, the signal was limited to phloem with no significant RNA accumulation in immature cells (b). In Rhizobia-infected roots the signal was highly represented in the parenchyma of the middle and inner cortex and in vascular bundles localised in proximity to the symbiosome (c). Leaves of water-deprived plants displayed the signal in both developing and functional phloem cells; parenchyma cells present in the central vein were devoid of signal (d). Panels a–d show cross sections of early drought-stressed tissues hybridised with anti-sense probe (positive signal). Stem (e), root ( f ), nodule (g) and leaf (h) cross sections hybridised with sense probe (negative signal) are shown. X, Xylem; Ph, Phloem; VB, Vascular bundles; S, Symbiosome. Scale bars = 100 µm.
b
a
