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Multiplex polymerase chain reaction analysis of Glu-1 high-molecular-mass glutenin genes from wheat by capillary electrophoresis with laser-induced fluorescence detection.
J Chromatogr A. 2004 Apr 02; 1032(1-2):313-8.JC

Abstract

The unique bread-making properties of wheat are closely correlated with composition and quantity of high-molecular-mass (HMW) glutenin subunits encoded by the Glu-1 genes. We report the development of a multiplex polymerase chain reaction (PCR) method to identify bread wheat genotypes carrying HMW glutenin allele composition of Glu-1 complex loci (Glu-A1, Glu-B1 and Glu-D1) by capillary electrophoresis(CE) with laser-induced fluorescence (LIF) detection. Two triplex primer sets of HMW glutenin subunit genes were examined. An automated and rapid CE-LIF technique is helpful in the multiplex PCR optimization process. Two fluorescent intercalating dyes (EnhanCE, and YO-PRO-1) are compared for detection of DNA fragments. Amplified DNA fragments of HMW glutenin Glu-1 genes were well separated both by agarose slab-gel electrophoresis and CE, and revealed minor differences between the sequences of 1Ax2*, 1Axnull, 1Bx6, 1Bx7, 1Bx17 and 1Dx5 genes. Moreover, CE technique requires samples of smaller volumes in comparison to slab-gel electrophoresis, and data can be obtained in less than 20 min. There was a very high concordance in the assessment of the molecular size of PCR-generated DNA markers. Fast and accurate identification of molecular markers of Glu-1 genes by CE-LIF can be an efficient alternative to standard procedure separation for early selection of useful wheat genotypes with good bread-making quality.

Authors+Show Affiliations

Salmanowicz BP
Institute of Plant Genetics, Polish Academy of Sciences, ul. Strzeszynska 34, PL 60-479 Poznan, Poland. bsal@igr.poznan.pl
Moczulski M
No affiliation info available

MeSH

Base SequenceDNA PrimersElectrophoresis, Agar GelElectrophoresis, CapillaryEthidiumGlutensLasersMolecular WeightPolymerase Chain ReactionSpectrometry, FluorescenceTriticum

Pub Type(s)

Journal Article

Language

eng

PubMed ID

15065810

Citation

Salmanowicz, Boleslaw P., and Marcin Moczulski. "Multiplex Polymerase Chain Reaction Analysis of Glu-1 High-molecular-mass Glutenin Genes From Wheat By Capillary Electrophoresis With Laser-induced Fluorescence Detection." Journal of Chromatography. A, vol. 1032, no. 1-2, 2004, pp. 313-8.
Salmanowicz BP, Moczulski M. Multiplex polymerase chain reaction analysis of Glu-1 high-molecular-mass glutenin genes from wheat by capillary electrophoresis with laser-induced fluorescence detection. J Chromatogr A. 2004;1032(1-2):313-8.
Salmanowicz, B. P., & Moczulski, M. (2004). Multiplex polymerase chain reaction analysis of Glu-1 high-molecular-mass glutenin genes from wheat by capillary electrophoresis with laser-induced fluorescence detection. Journal of Chromatography. A, 1032(1-2), 313-8.
Salmanowicz BP, Moczulski M. Multiplex Polymerase Chain Reaction Analysis of Glu-1 High-molecular-mass Glutenin Genes From Wheat By Capillary Electrophoresis With Laser-induced Fluorescence Detection. J Chromatogr A. 2004 Apr 2;1032(1-2):313-8. PubMed PMID: 15065810.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Multiplex polymerase chain reaction analysis of Glu-1 high-molecular-mass glutenin genes from wheat by capillary electrophoresis with laser-induced fluorescence detection. AU - Salmanowicz,Boleslaw P, AU - Moczulski,Marcin, PY - 2004/4/7/pubmed PY - 2004/12/16/medline PY - 2004/4/7/entrez SP - 313 EP - 8 JF - Journal of chromatography. A JO - J Chromatogr A VL - 1032 IS - 1-2 N2 - The unique bread-making properties of wheat are closely correlated with composition and quantity of high-molecular-mass (HMW) glutenin subunits encoded by the Glu-1 genes. We report the development of a multiplex polymerase chain reaction (PCR) method to identify bread wheat genotypes carrying HMW glutenin allele composition of Glu-1 complex loci (Glu-A1, Glu-B1 and Glu-D1) by capillary electrophoresis(CE) with laser-induced fluorescence (LIF) detection. Two triplex primer sets of HMW glutenin subunit genes were examined. An automated and rapid CE-LIF technique is helpful in the multiplex PCR optimization process. Two fluorescent intercalating dyes (EnhanCE, and YO-PRO-1) are compared for detection of DNA fragments. Amplified DNA fragments of HMW glutenin Glu-1 genes were well separated both by agarose slab-gel electrophoresis and CE, and revealed minor differences between the sequences of 1Ax2*, 1Axnull, 1Bx6, 1Bx7, 1Bx17 and 1Dx5 genes. Moreover, CE technique requires samples of smaller volumes in comparison to slab-gel electrophoresis, and data can be obtained in less than 20 min. There was a very high concordance in the assessment of the molecular size of PCR-generated DNA markers. Fast and accurate identification of molecular markers of Glu-1 genes by CE-LIF can be an efficient alternative to standard procedure separation for early selection of useful wheat genotypes with good bread-making quality. SN - 0021-9673 UR - https://www.unboundmedicine.com/medline/citation/15065810/Multiplex_polymerase_chain_reaction_analysis_of_Glu_1_high_molecular_mass_glutenin_genes_from_wheat_by_capillary_electrophoresis_with_laser_induced_fluorescence_detection_ DB - PRIME DP - Unbound Medicine ER -