1,721,082 research outputs found
Caratterizzazione molecolare di geni per la qualità e quantità delle proteine di riserva delle cariossidi dei frumenti
No full textDevelopment of transgenic wheat plants free of herbicide marker genes and plasmid backbone sequences through “clean gene” technology and positive selection
No full textFunctional markers for glutamine synthetase and correlation with grain protein content in durum wheat
No full textDurum wheat (Triticum turgidum L. var. durum) is one of the most important cereal crops grown world-wide and provides most of the proteins in human diet, especially in the less developed countries. Seed storage proteins are directly related to the nutritional and technological value of the derived products. Several studies have attested the key-role of the glutamine synthetase enzyme in plant nitrogen metabolism. Glutamine synthetase gene encodes for an enzyme responsible of the first step of ammonium assimilation and transformation into glutamine and glutamate, essential compounds in amino acid-biosynthetic pathway. High protein content is a very important quantitative trait controlled by several genes located on wheat chromosomes. Glutamine synthetase genes are located on the homeologous chromosomes 2A, 2B, and 2D where several authors reported major QTL for protein content. The goal of the present study was to assess the linkage between GS gene and the QTL for protein content. For this purpose, the nucleotide sequence of glutamine synthetase gene acc. DQ124214 was aligned to all the wheat ESTs available in public data bases by means of BLAST tool (http://www.wheat.pw.usda.gov/GG2/blast.shtml.). The bioinformatic analysis allowed to find 40 sequences with a similarity > 94% to the GS2 gene, of which three covered the whole gene sequence (DQ124213, DQ124212 and CJ705909). For each of these sequences we designed two or three primer pairs identifying a total of 7 functional markers that were screened among the parents of three segregant populations. Mapping analysis performed by Join Map software allowed to localize the amplified polymorphic fragments and to identify 4 loci: Gs-A2, Gs-B2, Gs-A4, Gs-B4, respectively mapped on chromosome 2A, 2B, 4A and 4B. The QTL analysis for protein content was carried out in a RIL population obtained from the crossing the two durum wheat cultivars Ciccio and Svevo. Two major QTLs were identified through Composite Interval Mapping (CIM) performed by the Q-Gene software: one QTL was identified by the functional marker Gs-B2 located on chromosome 2B, and the other one was identified by the functional marker Gs-A4 located on chromosome 4A. These data were confirmed by a linkage disequilibrium analysis carried on a collection of 75 different wheat genotypes. The present study represents the first step for the identification and sequencing of GS2 gene, which could be employed in breeding programs aimed to increase grain protein content commercial cultivars. Moreover, Gs-B2 and Gs-A4 represents functional markers that could be also efficiently used in marker assisted selection (MAS) programs and map-based cloning
Detection of transgene copy number in durum wheat transgenic lines using Real-Time PCR
No full textGenetic transformation has played a key role in gaining and applying knowledge of the roles of HMW-GS in wheat end-use properties. Reliable and stable expression of transgenes as well as the characterization and field adaptation of transgenic lines are prerequisites for the successful application of gene technology. Loci that appear to be stably expressed initially can become progressively silenced over generations. The stability and the behaviour of transgenes are influenced by several factors, such as chromosomal location, transgene copy number and their interaction with the host genotype. Traditionally such factors are characterized using Southern analysis which can be time consuming and laborious. Recent results obtained in various crops indicate that Real-Time PCR could be a powerful tool for the detection and characterization of transgene locus structures. The determination of transgenic locus number through Real-Time PCR overcomes the problems linked to phenotypic segregation analysis and can analyze hundreds of samples in a day making it an efficient method for estimating copy number integrated in a transgenic line. This study was conducted to determine transgene copy number in transgenic lines and to investigate potential differences in sensitivity, resolution and variability between two different RealTime chemistries (SYBR Green dye and TaqMan probes). We have applied Real-Time PCR to a set of four transgenic durum wheat lines previously obtained. A total of six experiments (two experiments for each gene) were conducted and standard curves were obtained from serial dilutions of the plasmids containing the genes of interest. The correlation coefficients of the standard curves were rather good, being in the range between 0.95 and 0.97. By using TaqMan quantitative Realtime PCR we were able to achieve estimates of 1 to 42 copies of transgenes per haploid genome in T4 homozygous transformants. Conversely, SYBR Green dye method revealed unable to accurately quantify transgene copy number as it failed in detecting the inserted genes when integrated in few copies. In our study we assessed Real-Time PCR as a fast, sensitive and reliable method for the detection of transgene copies in durum wheat, which can be a valid alternative to Southern analysis
Identification of QTLs with a key role in resistance against Fusarium Head Blight in durum wheat
No full textDurum wheat (T. turgidum ssp. durum) is one of most susceptible cereals to
Fusarium head blight (FHB, scab) which is annually responsible for serious
economic threats due to huge losses in yield, and for decay in qualitative
characteristics of the grain (destruction of cell walls, alteration of the
lipid fraction and the reduction of the protein fraction). FHB is also
responsible to produce mycotoxins mainly deoxynivalenol (DON), a powerful
inhibitor of eukaryotic protein synthesis and very harmful to human and
animal health. The most effective strategy to manage FHB disease and gain a
more economically and ecologically sustainable wheat production is the use
of genetic resistance, which is controlled by the combined effects of
several quantitative trait loci (QTL) and environment.
Resistance to FHB is a complex and quantitative trait controlled by
multiple genes, largely influenced by plant architecture and genotypeenvironment interactions, and characterized by large genetic variation in
wheat gene pool. Resistance to FHB is of quantitative nature and its
inheritance includes many genes and is affected by environmental
conditions.
Cell wall is the first line of plants defense against fungal pathogens and
several lines of evidence indicate that structural components of the cell
wall are involved in plant resistance against such pathogens. Understanding
the biological mechanisms and associated biomarkers driving wheat
development and adaptation relies on an urgent understanding of the
continuum between structural, expressional and epigenetic variations. A
specific activity has been conducted for Fusarium Head Blight (FHB),
reported QTL identification and the map-based cloning of a new FHB QTL
located on 2A chromosome from a resistant bread wheat line deriving from
Sumai 3. New functional marker genes conferring durable plant resistance
against Fusarium were developed and efficiently used in a marker-assisted
selection program for the constitution of resistant superior durum wheat
genotypes to be used for a sustainable agriculture and food security
Il “clean gene” come metodo innovativo per la produzione di piante transgeniche prive di geni di resistenza ad antibiotici o erbicidi
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