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Research on the Isolation of Mutated Major Gene Controlling Wheat Grain Hardness from Aegilops

Author: HuoHuaLei
Tutor: HeGuangYuan
School: Huazhong University of Science and Technology
Course: Biochemical Engineering
Keywords: Puroindoline Mutant Grain hardness Aegilops Wheat Quality
CLC: S512.1
Type: Master's thesis
Year: 2005
Downloads: 81
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Abstract


Wheat hardness by the the wheat hardness major gene Puroindoline A (PinA) Puroindoline B (PinB) decided both 78.0 mutated or missing, both will have a different impact on grain hardness. The Aegilops be important gene source of wild relatives of wheat and wheat is a genetic relationship, but also the improvement of wheat varieties. Rich crops of wheat grain hardness of the gene pool, lay the foundation for China's wheat quality improvement work, the study of the mature seeds Puroindolines protein analysis method and Aegilops PinA homology gene were isolated and analyzed, the main contents and as follows: (1) the establishment of Puroindoline protein electrophoresis analysis system and optimization: from diploid and hexaploid wheat materials and wheat relative species Aegilops mature seeds extract Puroindoline protein experiments Puroindoline proteins by SDS-PAGE and the dyeing conditions were optimized, the establishment of a concentrated gel T (gel concentration) 4, C (crosslinking degree) of 2.6, the electrophoresis voltage is 128V; separating gel T is 13.5, C 2.6, electrophoresis voltage of 240V, stable and reproducible optimized separation gel electrophoresis time 1.5h electrophoresis results SDS gel electrophoresis conditions. Also the introduction of electrophoretic bands clearer analysis of the effect of urea gel electrophoresis and Coomassie blue staining (blue dye) and stained with silver nitrate (silver stain) stain concentration range of two staining methods and staining effect compared and obtain good experimental results, and laid the foundation for the the wheat Puroindoline further study of the gene. (2) Aegilops PinA gene mutations in body screening: reported PinADNA sequence, designed the PinA of Jane and primers, Aegilops genus (Aegilops) four groups of 34 copies of material in the genomic DNA as a template, the optimization 25 amplification products were amplified by PCR system, and isolated and sequenced, and compared with the sequence DNAstar software and GenBank, the results show that the seven sequencing sequence with sequences in GenBank exactly the same, but the materials different sources; 14 sequences determined PinA novel mutant, the homology between them is 97.8% to 100%, the sequence of this are 447 bp - length, top of the two materials in RM0070 and RM0072 PinA Since point mutations tryptophan the second W mutations in the domain (WWKWWK) point mutation in the tryptophan extraterritorial also produced a terminator for the termination of the child, and the the RM0032 RM0051 material PinA generated, other sequences encoding a rich color histidine region of 149 amino acid residues.

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CLC: > Agricultural Sciences > Crop > Cereal crops > Wheat > Wheat
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