Identification of Mutator insertional mutants of starch-branching enzyme 1 (sbe1) in Zea mays L.

被引：136

作者：

Blauth S.L. ^{[1
]}

Kim K.-N. ^{[1
]}

Klucinec J. ^{[2
]}

Shannon J.C. ^{[3
]}

Thompson D. ^{[2
]}

Guiltinan M. ^{[1
,3
]}

机构：

[1] Department of Food Science, 3113 Tyson Hall, Pennsylvania State University, University Park

[2] Department of Horticul-ture, Pennsylvania State University, 113 Tyson Hall, University Park

来源：

Plant Molecular Biology | 2002年 / 48卷 / 03期

关键词：

Amylopectin; Amylose; Reverse genetics; SBE; Starch biosynthesis;

D O I：

10.1023/A:1013335217744

中图分类号：

学科分类号：

摘要：

Starch-branching enzymes (SBE) alter starch structure by breaking an α-1,4 linkage and attaching the reducing end of the new chain to a glucan chain by an α 1,6 bond. In maize, three isoforms of SBE have been identified. In order to examine the function of the SBEI isoform, a reverse-genetics PCR-based screen was used to identify a mutant line segregating for a Mutator transposon within Sbe1. Compared to wild-type controls, Sbe1 transcripts accumulate at extremely low levels in leaves of the homozygous mutant. Antibodies failed to detect SBEI in leaf tissue of mutants or wild-type controls. In contrast, the level of SBEI in endosperm is undetectable in homozygous mutants while easily detected in wild-type controls. Starches extracted from mutant leaves and endosperm have structures indistinguishable from starches of wild-type controls as determined by size-exclusion chromatography (SEC) of intact starch and high-performance SEC of debranched starch. To investigate the possibility of compensation for the lack of SBEI by expression of the homologous sequence reported by Kim et al. (1998), a genomic fragment (Sbe1b) of this sequence was cloned. Northern hybridizations of mutant leaf, root, tassel, endosperm and embryo tissues with non-specific Sbe1b probes failed to reveal expression of the homologous sequence. These results suggest that the homologous sequence is not compensating for a lack of SBEI in sbe1::Mu mutants. Further study of this sbe1 mutation in the presence of other genetic mutations may help to understand the role of SBEI in determining starch structure in leaves and endosperm.

引用

页码：287 / 297

页数：10

共 29 条

[21]

Klucinec J., Thompson D., Fractionation of high-amylose maize starches by differential alcohol precipitation and chromatography of the fractions, Cereal Chem., 75, pp. 887-896, (1998)

[22]

Mena M., Ambrose B., Meeley R., Briggs S., Yanofsky M., Schmidt R., Diversification of C-function activity in maize flower development, Science, 274, pp. 1537-1540, (1996)

[23]

Sambrook J., Fritsch E., Maniatis T., Molecular Cloning: A Laboratory Manual, 2nd ed., (1989)

[24]

Smith A., Denyer K., Martin C., The synthesis of the starch granule, Annu. Rev. Plant Physiol. Plant Mol. Biol., 48, pp. 67-87, (1997)

[25]

Stinard P., Robertson D., Schnable P., Genetic isolation, cloning and analysis of Mutator-induced, dominant antimorph of the maize amylose-extender1 locus, Plant Cell, 5, pp. 1555-1566, (1993)

[26]

Stoscheck C., Quantitation of protein, Meth. Enzymol., 182, pp. 50-69, (1990)

[27]

Takeda Y., Guan H.-P., Preiss J., Branching of amylose by the branching isoenzymes of maize endosperm, Carbohydrate Res., 240, pp. 253-263, (1993)

[28]

Towbin H., Staehelin T., Gordon J., Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: Procedure and some applications, Proc. Natl. Acad. Sci. USA, 76, pp. 4350-4354, (1979)

[29]

Vineyard M., Bear R., Amylose content, Maize Genet. Coop. Newsl., 26, (1952)

← 1 2 3 →