Fig 1.
Photomicrographs of leaf transverse sections of cassava cultivars and wild relatives.
(a1), (b1) and (c1), leave midrib of W14, SC205 and SC8, respectively; (a2), (b2) and (c2), Amplification of leave midrib of W14, SC205 and SC8, respectively; Primary xylem (PX); Primary Phloem (PP); Spongy mesophyll (SM); Collenchyma (CC); (a3), (b3) and (c3), leaf transverse sections of W14, SC205 and SC8, respectively. Note the long single palisade layer (PL) and the conspicuous green vascular bundle sheath (VBS) cells situated beneath the palisade layer. Scale bar = 40 μm.
Fig 2.
Imaging pulse amplitude modulation of W14 (a), SC205 (b) and SC8 (c) leaves.
Parameters were Fv/Fm [maximal photosystem II (PSII) quantum yield], ΦPSII (effective PSII quantum yield) (at 185μE m-2 s-1), and NPQ/4 (nonphotochemical quenching) (at 185μE m-2s-1). The color gradient provided a scale from 0 to 100% for assessing the magnitude of the parameters.
Table 1.
Photosynthetic parameters collected from cassava leaves of W14, SC205 and SC8.
Values were means ± SE. Different capital letters in the same column indicated statistically significant differences according to Duncan test (P<0.01).
Table 2.
Dry matter content, starch content, amylose and amylopectin content in storage roots of W14, SC205 and SC8.
Values were means ± SE. Different capital letters in the same column indicated statistically significant differences according to Duncan test(P<0.01).
Fig 3.
Starch staining with KI in storage roots and SEM pictures of starch granules incubated with cell-free supernatants, and paraffin section of transverse and longitude of storage roots of SC205, SC8 and W14.
Paraffin section of transverse and longitude of storage roots of SC205, SC8 and W14, stained with Safranin O/Fast green and viewed under light microscope X20. (a1), (b1) and (c1), transverse sections of cassava genotype W14, SC205 and SC8, respectively; (a2), (b2) and (c2), starch staining with KI of cassava genotype W14, SC205 and SC8, respectively; (a3), (b3) and (c3), SEM of starch granules of cassava W14, SC205 and SC8, respectively. SEM magnification time was 1000; scale bar = 50 μm. (a4)-(c4), transverse sections of storage roots from W14, SC205 and SC8; (a5)-(c5), longitude sections of storage roots from W14, SC205and SC8. Scale bar = 100 μm. It shows more vessel grouping and tyloses with starch granules. SC205 and SC8 showing parenchyma cells with more starch granules, while W14 showing parenchyma cells with a little starch, these cells are bigger. Red arrows indicate xylem vessel, black arrow shows parenchyma cell, and blue arrow presents starch granules.
Fig 4.
148, 157 and 152 proteins identified by MALDI-TOF-TOF-MS/MS in 2-D gel protein profiles of W14(a), SC205(b) and SC8(c) leaves, respectively.
The pink numbers are common proteins to W14 and SC205, the yellow numbers are common proteins to W14 and SC8, and the orange numbers are common proteins to SC205 and SC8.
Fig 5.
Venn diagrams of 175 proteins identified (a) and their functional classification (b) in leaves of SC205, SC8 and W14.
Functional categorization was performed according to the MIPS database (http://mips.gsf.de).
Table 3.
Identification of 122 common proteins in leaves of SC205, SC8 and W14.
The spots showing the similar proteins from 2-DE images of cassava SC205, SC8 and W14 leaves, and the number were counted after gel analysis and manual editing with Delta 2D software.
Table 4.
Identification of the unique protein spots in leaves detected by pairwise comparison of W14/SC205, W14/SC8 and SC205/SC8.
Fig 6.
196, 228 and 232 proteins identified by MALDI-TOF-TOF-MS/MS in 2-D gel protein profiles of W14(a), SC205(b) and SC8(c) storage roots, respectively.
The pink numbers are common proteins to W14 and SC205, the yellow numbers are common proteins to W14 and SC8, and the orange numbers are common proteins to SC205 and SC8.
Fig 7.
Venn diagrams of 308 proteins identified (a) and their functional classification (b) in storage roots of SC205, SC8 and W14.
Functional categorization was performed according to the MIPS database (http://mips.gsf.de).
Table 5.
Identification of 127 common proteins from storage roots of SC205, SC8 and W14.
The spots showing the same proteins in storage roots of W14, SC205 and SC8, and the number were counted after gel analysis and manual editing with Delta 2D software.
Table 6.
Identification of the unique proteins in storage roots detected by pairwise comparison of W14/SC205, W14/SC8 and SC205/SC8.
Fig 8.
Western blotting of Rubisco (a), OEC (b) and D1 (c).
The expression of Rubisco, OEC and D1 in leaves of cassava W14, SC205 and SC8 were detected by western blotting using anti-Rubisco-polyclonal antibody (AS07218), anti-OEC antibody (AS 05092) and anti-D1 antibody (AS05084) from Agrisera, respectively.
Fig 9.
Western blotting of linamarase (a), GBSS1 (b) and beta-amylase (c).
The expression of linamarase, GBSS1and beta-amylase in storage root of cassava W14, SC205 and SC8 genotypes were detected by western blotting using anti- linamarase antibody anti- GBSS1 antibody, produced by GenScript, and anti-beta-amylase antibody (AS09379) from Agrisera.
Fig 10.
Biological networks generated from a combination of 19 differential proteins involved with photosynthesis (a) in cassava leaves and 11 differential proteins related with starch accumulation (b) in storage roots.
Nineteen differentially up-(a red and upward arrow) and down-(a blue and downward arrow) regulated proteins including ribulose-bisphosphate carboxylase, phosphoribulokinase, ribulose- phosphate-3-epimerase, ribose-5-phosphate isomerase, RCA, transketolase, ATP synthase subunit beta, phosphoglycerate kinase, malate dehydrogenase, alcohol dehydrogenase and enoyl-ACP reductase, ethylene receptor, peroxiredoxin, heat shock protein, glucokinase, glutaredoxin, superoxide dismutase, beta-glucosidase and APX2 in cassava cultivars were used to generate a protein-protein interaction network about photosynthesis through Pathway Studio analysis. Eleven differentially up-(a red and upward arrow) and down-(a blue and downward arrow) regulated proteins including succinate dehydrogenase, dihydrolipoyllysine-residue succinyltransferase, UDP- glucosyltrans-ferase, transaldolase, uroporphyrinogen decarboxylase, pectinesterase, triosephosphate isomerase, N-acetyltransferase, aldo-keto reductase, annexin and pyruvate dehydrogenase in cassava cultivars were used to generate a protein-protein interaction network regarding starch accumulation through Pathway Studio analysis. Regulation is marked as an arrow with R, Chemical Reaction as an arrow with C, MolTransport as an arrow with M, Expression as an arrow with E and Binding as an arrow without any marks. The entity table and relation table were presented in S1 and S2 Tables.