NPH_2694_sm_FigS3

advertisement

(a)

Col: TcNRAMP4

4-1 4-2 WT

Col: TcNRAMP4

4-1 4-2 WT

NRAMP4 actin

(b)

Ws

_ev

Ws

_

TcNRAMP4

-6

(c)

80 tubulin

3221

4522

NRAMP4

2511

954700

NRAMP4/ tubulin

0,78

211

Col

_WT 4-1 4-2

Ws

_ev 4-6

60

40

20

(d)

0 10 30 300

CdCl

2

(µM)

ZnSO

4

(µM)

Col

_WT 4-1 4-2

Ws

_ev 4-6

300

200

100

90

60

30

Zn Cd

Supporting Information

Fig. S3 Expression of

TcNRAMP4

in

A. thaliana

accessions

Columbia

(

Col

) and

Wassilewskija

(

Ws

) does not modify Zn or Cd sensitivity nor their content.

(a) Semi-quantitative RT-PCR of

TcNRAMP4

and Actin (control) expression in the

Col 35S::TcNRAMP4

lines (

4-1 and 4-2

) and WT. (b) Quantitative RT-PCR analysis of

TcNRAMP4

35S::TcNRAMP4

and

tubulin

(control) expression in

Ws

line 4-6 and empty vector control. (c) Root length of the

35S::TcNRAMP4

lines and the control lines grown vertically for 11 d on ABIS medium containing 50

µM FeHBED. The medium was supplemented with 0,

10 and 10

µM CdCl

2

, or 300

µM ZnSO

4

. Values are the average of measurements on 10-12 roots and bars represent SD. (d) Zn and Cd contents in shoots of the

35S::TcNRAMP4

lines and the control lines grown on control

ABIS medium (Zn content) and ABIS with 3

µM CdCl

2

(Cd content).

Metal content determination (different from the method used in the principle manuscript)

Seedlings were grown vertically for 12 d on plates containing ABIS medium (1% agar), supplemented or not with 30

µM CdCl

2 or 300

µM ZnCl

2

. Roots and shoots were harvested separately and their metal content was measured as described in Lanquar measured after drying at 60

et al.

(2005). The dry weight (DW) of the samples was

°C for 3 d. The dried samples were digested in a mixture of 1 ml of 70% nitric acid and 5 ml ultrapure H

2

O in a MARS X microwave (CEMCorporation, Matthews, NC, USA)) at

210

°C at 24 bar for 20 min. After dilution in trace-metal-free water, the metal content of the samples was determined by inductively coupled plasma optical emission spectroscopy (ICP-OES) using a Vista-MPX,

(Varian, Australia)).

Download