ISSN:
1573-5028
Keywords:
glycine-rich proteins (GRPs)
;
subtractive hybridization
;
somatic embryogenesis
Source:
Springer Online Journal Archives 1860-2000
Topics:
Biology
Notes:
Abstract We report the first successful isolation by subtractive hybridization of a gene expressed specifically during somatic embryogenesis. Embryogenic cell clusters, 32–50 μm in diameter, were isolated by sieving and density-gradient centrifugation. The cDNA library was constructed from proglobulars which were formed from embryogenic cell clusters 3 days after transfer to auxin-free modified Lin and Staba's medium. For use as probe in screening, the same cDNA used for library construction was enriched for specific sequences using subtractive hybridization. The cDNA used for subtraction was prepared from suspension cultures 5 days after subculturing in auxin-containing medium. Nine independent differentially expressed cDNA clones were obtained from a screen of 150 000 recombinant phages. Northern analysis indicated one of these, CFM6, to be expressed specifically during somatic embryogenesis. In addition, one hybridizing transcript was detected in plantlet cotyledons, and two transcripts were detected in hypocotyls. Two separate and distinct hybridizing transcripts are expressed specifically in hypocotyl tissue. The amino acid sequence deduced from the nucleotide sequence of the CEM6 cDNA indicates that it encodes a glycine-rich protein containing a hydrophobic signal-sequence like domain. Its early embryo-specific expression and sequence characteristics suggest an important role as a cell wall protein in embryogenesis.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1007/BF00042036
Permalink