Characterization and expression analysis of genes directing galactomannan synthesis in coffee.
OBJECTIVE
Galactomannans act as storage reserves for the seeds in some plants, such as guar (Cyamopsis tetragonoloba) and coffee (Coffea arabica and Coffea canephora). In coffee, the galactomannans can represent up to 25 % of the mass of the mature green coffee grain, and they exert a significant influence on the production of different types of coffee products. The objective of the current work was to isolate and characterize cDNA encoding proteins responsible for galactomannan synthesis in coffee and to study the expression of the corresponding transcripts in the developing coffee grain from C. arabica and C. canephora, which potentially exhibit slight galactomannan variations. Comparative gene expression analysis was also carried out for several other tissues of C. arabica and C. canephora.


METHODS
cDNA banks, RACE-PCR and genome walking were used to generate full-length cDNA for two putative coffee mannan synthases (ManS) and two galactomannan galactosyl transferases (GMGT). Gene-specific probe-primer sets were then generated and used to carry out comparative expression analysis of the corresponding genes in different coffee tissues using quantitative RT-PCR.


RESULTS
Two of the putative galactomannan biosynthetic genes, ManS1 and GMGT1, were demonstrated to have very high expression in the developing coffee grain of both Coffea species during endosperm development, consistent with our proposal that these two genes are responsible for the production of the majority of the galactomannans found in the grain. In contrast, the expression data presented indicates that the ManS2 gene product is probably involved in the synthesis of the galactomannans found in green tissue.


CONCLUSIONS
The identification of genes implicated in galactomannan synthesis in coffee are presented. The data obtained will enable more detailed studies on the biosynthesis of this important component of coffee grain and contribute to a better understanding of some functional differences between grain from C. arabica and C. canephora.
Centre de Recherche Nestlé, 101 Av. Gustave Eiffel, Notre Dame d'Oé, BP 49716 - 37097 Tours, France.
Martial
M
Pré
Victoria
V
Caillet
Julien
J
Sobilo
James
J
McCarthy
D000595:Amino Acid Sequence; D001483:Base Sequence; D003069:Coffee / Q000201:enzymology / Q000235:genetics*; D018076:DNA, Complementary / Q000235:genetics; D020224:Expressed Sequence Tags; D005690:Galactose / Q000031:analogs & derivatives; D005700:Galactosyltransferases / Q000235:genetics*; D018507:Gene Expression Regulation, Developmental; D018506:Gene Expression Regulation, Plant; D015723:Gene Library; D017343:Genes, Plant; D018745:Genome, Plant; D008351:Mannans / Q000096:biosynthesis*; D008364:Mannosyltransferases / Q000235:genetics*; D010802:Phylogeny; D018515:Plant Leaves / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012639:Seeds / Q000235:genetics; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA
D003069:Coffee; D018076:DNA, Complementary; D008351:Mannans; C012990:galactomannan; D005700:Galactosyltransferases; D008364:Mannosyltransferases; C021542:mannan synthetase; D005690:Galactose