Monoclonal antibodies used to characterize cDNA clones expressing specific wheat endosperm proteins

Donovan, Gregory R.; Skerritt, John H.; Castle, SL

doi:10.1016/s0733-5210(89)80011-6

Cited by 13 publications

(9 citation statements)

References 19 publications

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“…This might result from the different specificity of two antibodies, which only bound to LMW-GS, without binding to HMW-GS. However, the antibodies reported before, which showed significantly positive correlations with wheat quality properties, bound only to some HMW-GS or not only to LMW-GS and gliadins but to HMW-GS (Skerritt and Underwood 1986;Donovan et al 1989;Skerritt and Robson 1990;Song et al 1997Song et al , 1998c. In this study, the antibody mAb 14588, binding not only to HMW-GS, but to some LMW-GS, also had significantly positive correlations with both wheat quality parameters development time and sedimentation volume.…”

Section: Quality-antibody Binding Correlation Analysismentioning

confidence: 99%

Characterization of monoclonal antibodies specific to wheat glutenin subunits and their correlation with quality parameters

Li¹,

Ji²,

Zhang³

et al. 2009

Can. J. Plant Sci.

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2009. Characterization of monoclonal antibodies specific to wheat glutenin subunits and their correlation with quality parameters. Can. J. Plant Sci. 89: 11Á19. Immunochemical methods are very useful in predicting the quality of wheat and differentiating alleles. In order to prepare appropriate monoclonal antibodies, HMW-GS 1Bx13 and 1By16 from spelt wheat were used as antigens to immunize BALB/C mice. Four monoclonal antibodies (mAbs) were obtained and designated 24231, 24245, 14588 and 14587, respectively. Results of Western blot showed that mAbs 24231 and 24245 prepared against 1Bx13 bound only to LMW-GS. The mAb 14588 prepared against 1By16 bound strongly to LMW-GS, but weakly to 1By and some 1Dy type HMW-GS. The mAb 14587 prepared against 1By16 bound only to 1Dx HMW-GS. The results of indirect ELISA and statistical analysis showed that correlations between mAb 24231 and development time and stability were significantly (P B0.05) and highly significantly (PB0.01) negative, respectively, whereas those of mAb 24245 with development time and extensibility were highly significantly (PB0.01) and significantly (PB0.05) negative, respectively. Significantly (PB0.05) and highly significantly (PB0.01) positive correlations were observed between mAb 14588 and stability and development time. Mean differential binding of mAb 14587 with 1Dx5 t and 1Dx2 t subunits from Aegilops tauschii was highly significant (PB0.01), suggesting that ELISA could potentially be used as an effective screening tool during direct genetic transfer of desirable glutenin subunits from Aegilops tauschii to hexaploid wheat.. 2009. Caracte´risationCaracte´risation des anticorps monoclonaux spe´cifiquesspe´cifiques aux sous-unite´sunite´s de la glute´nineglute´nine du bleét correíation avec lesparamè tres de la qualite´.qualite´. Can. J. Plant Sci. 89: 11Á19. L'immunochimie s'est ave´reéave´reéparticuliè rement utile pour pre´direpre´dire la qualite´duqualite´du bleét la diffe´renciationdiffe´renciation desallè les. Pour pre´parerpre´parer les anticorps monoclonaux ne´cessairesne´cessaires, les auteurs ont pris les sous-unite´sunite´s de la glute´nineglute´nine a` haut poids mole´culairemole´culaire 1Bx13 et 1By16 de l'e´peautree´peautre commeantigè nes et s'en sont servi pour immuniser des souris BALB/C. Ils ont ainsi obtenu quatre anticorps monoclonaux (mAb) qu'ils ont baptise´sbaptise´s respectivement 24231, 24245, 14588 et 14587. Les re´sultatsre´sultats de la me´thodeme´thode de Western indiquent que les mAb 24231 et 24245 pre´pare´spre´pare´pre´pare´s avec 1Bx13 ne se lient qu'aux sous-unite´sunite´s de la glute´nineglute´nine a` faible poids mole´culairemole´culaire. Le mAb 14588 pre´pareávecpre´pareávec 1By16 se lie fortement aux sous-unite´sunite´s de la glute´nineglute´nine a` faible poids mole´culairemole´culaire, mais se lie peu aux sous-unite´sunite´s a` haut poids mole´culairemole´culaire 1By et a` certaines du type 1Dy. Le mAb 14587 pre´pareávecpre´pareávec 1By16 ne se lie qu'aux sous-unite´sunite´s de la glute´nineglute´nine a` haut...

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Section: Quality-antibody Binding Correlation Analysismentioning

confidence: 99%

Characterization of monoclonal antibodies specific to wheat glutenin subunits and their correlation with quality parameters

Li¹,

Ji²,

Zhang³

et al. 2009

Can. J. Plant Sci.

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“…cDNA clones were isolated from libraries prepared from the hard Australian cv. Timgalen (Donovan et al, 1989) and from soft Falcon, which is a line near-isogenic to the hard Australian cv. Falcon and derived from a cross between Falcon and the soft Australian cv.…”

Section: Wheat Cultivars Usedmentioning

confidence: 99%

“…Timgalen endosperm cDNA libraries has been described (Donovan et al, 1989). Seed cDNA from soft Falcon was synthesised from mRNA using oligo(dT) priming and an Amersham kit according to the manufacturer's technical bulletins.…”

Section: Construction Of Librariesmentioning

confidence: 99%

“…We therefore concluded that it was justified to screen a pre-existing expression (jLgt1 1) cDNA library reverse transcribed from hard (cv. Timgalen) seeds at about 14 days after flowering (DAF), (Donovan et al, 1989), with the GSP-specific 1 2 3 4 5 6 7 8 9 1 0 1 1 l 2 1 3 l 4 asGSP 1 2 3 4 5 6 7 8 9 1 0 1 1 1 2 1 3 1 4 asp1 OA Fig. 2.…”

Section: Isolation Of Putative Gsp Clones From a Hard Cultivarmentioning

confidence: 99%

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Cloning of a wheat 15‐kDa grain softness protein (GSP)

Rahman

Jolly

Skerritt

et al. 1994

European Journal of Biochemistry

108

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The wheat starch 15-kDa protein (called grain softness protein or GSP) consists of a major polypeptide and several minor polypeptides. An antiserum raised against GSP was used to screen a wheat cDNA library. A cDNA family encoding approximately 15-kDa proteins that included a heptapeptide sequence previously isolated from protease digests of GSP was identified. A partial cDNA was used in a prokaryotic expression system to produce a fusion protein which reacted strongly against the original anti-GSP serum. A new antiserum raised against the fusion protein produced a weak reaction against a 15-kDa polypeptide extracted from wheat seeds. The results suggest that the proteins encoded by the cDNA family form a minor component of the mixture of 15-kDa polypeptides defined as GSP. RNA complementary to the cDNAs could be extracted from both soft and hard wheat grains from about half-way through grain filling. The encoded proteins are novel members of the 2s superfamily of seed proteins, a diverse family of proteins which maintain a characteristic framework of cysteine residues. The deduced proteins show the highest similarity to the oat 16-kDa avenin and to wheat puroindoline (a lipid-binding 15-kDa protein from wheat). Review of previously published data shows that puroindoline is also closely related to the major polypeptide of GSP, suggesting that the lipid-binding properties of GSP polypeptides may influence grain softness.

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“…Timgalen) as described earlier [8]. Prior to screening, the primary antibody solution (1:20 000 dilution in TBST) was pre-absorbed by incubation with nitrocellulose membrane discs soaked in E. coli extract.…”

Section: Preparation and Screening Of A Cdna Library In )~Gt L Lmentioning

confidence: 99%

Isolation and characterization of wheat triticin cDNA revealing a unique lysine-rich repetitive domain

et al. 1993

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Polyclonal antibodies were raised against a purified 22 kDa triticin polypeptide (delta) and were used to screen a wheat seed cDNA library in the Escherichia coli expression vector lambda gt11. The isolated cDNA clones were grouped into three families based on their cross-hybridization reactions in DNA dot-blot studies. Southern blots of genomic DNAs extracted from ditelocentric and nullisomic-tetrasomic lines of Chinese Spring wheat, probes with the excised cDNA inserts, indicated that one of the three families (9 clones) had triticin clones. This was finally confirmed by comparing the predicted amino acid sequences of two of these clones (lambda Tri-12, lambda Tri-25) with the published tryptic peptide sequences of triticin. The Southern blots also showed that there is at least one triticin gene located on the short arm of each of the homoeologous group 1 chromosomes (1A, 1B, 1D), although till now no triticin protein product has been identified for the chromosome 1B. The nucleotide sequence of the largest triticin cDNA clone lambda Tri-25 (1567 bp) is presented here, and its predicted amino acid sequence shows strong homology with the legumin-like proteins of oats (12S globulin), rice (glutelin) and legume seeds. A unique feature of the triticin sequence is that it contains a lysine-rich repetitive domain, inserted in the hypervariable region of the typical legumin-like genes. Northern blotting of total RNA extracted from different stages of the developing wheat seed revealed that the triticin gene expression is switched on 5-10 days after anthesis (DAA). There was a steady increase in the level of triticin mRNA until 20 DAA, after which it started decreasing. The maximum mRNA accumulation occurred between 17 and 20 DAA. These observations conform closely with the published data on triticin protein accumulation during grain development.

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Monoclonal antibodies used to characterize cDNA clones expressing specific wheat endosperm proteins

Cited by 13 publications

References 19 publications

Characterization of monoclonal antibodies specific to wheat glutenin subunits and their correlation with quality parameters

Characterization of monoclonal antibodies specific to wheat glutenin subunits and their correlation with quality parameters

Cloning of a wheat 15‐kDa grain softness protein (GSP)

Isolation and characterization of wheat triticin cDNA revealing a unique lysine-rich repetitive domain

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