Fig 1.
Expression of Os08g01480 in Arabidopsis confers tolerance for HMs. Seeds of WT (Col-0) and transgenic lines (L1, L2 and L3) were germinated vertically for 11 d on ½ MS medium supplemented with 5 μM As(III), 100 μM As(V), 50 μM Cd and 50 μM Cr(VI).
(A) Phenotypic changes of WT and transgenic lines grown in different HMs. (B) Relative Root length of the WT and transgenic plants during stress conditions in comparison to WT and transgenic plants grown in non treated media. All values are the mean of triplicates (±SD) Values marked with similar letters are not significantly (Duncan’s test: p<0.05) different.
Fig 2.
Expression of Os08g01480 in Arabidopsis confers tolerance for salt and osmotic stress.
(A) Relative root length of the WT (Col-0) and transgenic plants after vertical growth of 11 d on ½ MS media plates supplemented with NaCl and Mannitol in comparison to WT and transgenic lines grown in non treated media. All values are the mean of triplicates (±SD) Values marked with similar letters are not significantly (Duncan’s test: p<0.05) different. (B) Phenotypic changes of WT and transgenic lines grown in ½ MS medium supplemented with NaCl and Mannitol.
Fig 3.
Expression of Os08g01480 in Arabidopsis confers tolerance to temperature stress.
(A) Relative root length of the WT (Col-0) and transgenic plants after vertical growth of 11 d on ½ MS media plates after exposure at -20°C and 37°C for 4h in comparison to WT and transgenic lines grown in non treated media. (B) Phenotypic changes of WT and transgenic lines grown after exposure at -20°C and 37°C All values are the mean of triplicates (±SD) Values marked with similar letters are not significantly (Duncan’s test: p<0.05) different.
Fig 4.
Chlorophyll fluorescence imaging (Fv/Fm) of WT and transgenic line (L1) was performed during control and stress conditions.
Fv/Fm of non-treated WT and transgenic was carried out after 14 days of germination. Fv/Fm was carried out for WT and transgenic line after they were exposed to 37°C and 0°C for 4 h and again after10 and 15d recovery from heat and cold stress, respectively. For salt and different HM stress WT and transgenic line were watered with nutrient media containing NaCl (300 mM), 100 μM As(III), 200 μM As(V), 200 μM Cd and 400 μM Cr(VI) for two weeks and then Fv/Fm were measured. For osmotic stress WT and transgenic line were withheld from water for 14 d, and then rewatered for 7 d before Fv/Fm was measured. The false color code depicted on the right side of images ranges from 0 (black) to 1 (pink) Level of significance is considered as P≤0.05.
Fig 5.
Pathway studio analysis used for prediction of molecular network modulated by expression of CYP like gene Os08g01480 in Arabidopsis.
Pathway studio is utilized for finding common molecular connections between the proteins/transcription factors encoded by the expressed probe sets. This software analyses through the ResNet database for all known interactions between genes/proteins such as regulation and their expression. Each arrow indicates interactions between genes and a cell process pathway.
Fig 6.
Expression analysis of co-regulated genes related to network modulated by expression of Os08g01480 in Arabidopsis.
qRT-PCR analysis was carried out in two week old seedlings of WT and transgenic lines grown in ½ MS media.
Fig 7.
Promoter activity in Arabidopsis line expressing ProOs08g01480:uidA.
(A) Histochemical GUS staining (B) Relative expression level of uid-A gene of transgenic seedling (10 d old), grown in ½ MS media supplemented with 5 and 25 μM As(III), 50 and 100 μM As(V), 30 and 50 μM Cd, 50 and 100 μM Cr(VI), 150 and 300 mM mannitol, 100 and 150 mM NaCl. For temperature stress seeds of transgenic line grown in ½ MS media plate and exposed to -20°C, -80°C, 37°C and 45°C for 4 h then allowed to grow in control conditions (10 d). NT represents for non treated transgenic line.