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Molecular cloning and characterization of an endo-β-mannanase gene expressed in the lettuce endosperm following radicle emergence

Published online by Cambridge University Press:  22 February 2007

Aoxue Wang ,
Jieran Li  and
J. Derek Bewley
Aoxue Wang
Affiliation:
Department of Botany, University of Guelph, Guelph, Ontario, N1G 2W1, Canada
Jieran Li
Affiliation:
Department of Botany, University of Guelph, Guelph, Ontario, N1G 2W1, Canada
J. Derek Bewley*
Affiliation:
Department of Botany, University of Guelph, Guelph, Ontario, N1G 2W1, Canada
*
*Correspondence, Fax: +1 519 767 1991 Email:, [email protected]
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Abstract

Endo-β-mannanase (EC 3.2.1.78), an enzyme that mobilizes the endosperm cell walls of the lettuce (Lactuca sativa L.) seed, increases in activity in the micropylar and lateral regions of this tissue following the completion of germination. Its complementary DNA (cDNA) sequence (LsMan1) was determined using the polymerase chain reaction (PCR) with degenerate primers. The 3‘-end of the cDNA sequence was obtained by 3‘-end rapid amplification of cDNA ends (RACE), and the 5‘-end sequence by genome walking and 5‘-end RACE. The predicted amino acid sequence from the cDNA has a high identity with endo-β-mannanases present in other species (e.g. 67% identity with coffee β-1,4-mannan endohydrolase, 62% identity with tomato fruit endo-β-mannanase). Southern blot analysis suggests the presence of several members of an endo-β-mannanase gene family in the lettuce genome. Several isoforms of the enzyme, including three major ones, were detected by isoelectric focusing. Based on Northern blot analysis, accumulation of the endo-β-mannanase mRNA occurred only after lettuce seeds had germinated, and increased thereafter, although enzyme activity persisted after transcription declined.

Keywords

endo-β-mannanaseendospermgalactomannanlettuce seed germinationLactuca sativa
Type
Research Article
Information
Seed Science Research , Volume 14 , Issue 3 , September 2004 , pp. 267 - 276
Copyright
Copyright © Cambridge University Press 2004

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