scholarly journals Identification of Phytophthora alni subspecies in riparian stands in the Czech Republic

2013 ◽  
Vol 49 (Special Issue) ◽  
pp. S3-S10 ◽  
Author(s):  
P. Štěpánková ◽  
K. Černý ◽  
V. Strnadová ◽  
P. Hanáček ◽  
M. Tomšovský
Keyword(s):  
Czech Republic ◽  
Alnus Glutinosa ◽  
Pcr Primers ◽  
The Czech Republic ◽  
Specific Pcr ◽  
Related Taxon ◽  
Allele Specific ◽  
Allele Specific Pcr ◽  
Subspecific Level

In the Czech Republic, Phytophthora alni was first confirmed in 2001 and the pathogen has been quickly spreading and occupying almost the whole area of the country. The pathogen attacks Alnus glutinosa or A. incana to a lesser extent and causes considerable losses of alder trees along hundreds of kilometres of riverbanks. The aim of our work was to perform the identification of P. alni isolates at the subspecific level using PCR and to determine the frequencies and distribution of particular subspecies. The allele-specific PCR primers focused on allele diversity of orthologs of ASF-like, TRP1, RAS-Ypt, and GPA1 genes were selected for identification. Eighty-eight per cent of the 59 analysed isolates belonged to P. alni ssp. alni while 12% were P. alni ssp. uniformis. P. alni ssp. multiformis has not been recorded in the country till now. The two subspecies differed in distribution. P. alni ssp. alni dominated in riparian stands along broader rivers in lowlands and the results confirmed the more effective spreading of P. alni ssp. alni based on its higher aggressiveness and ecological advantage. P. alni ssp. uniformis was acquired rather from riparian stands of small watercourses at higher altitudes. The insular distribution of P. alni ssp. uniformis may represent the remains of its former occurrence. Therefore, P. alni ssp. uniformis may be an indigenous subspecies suppressed by the more aggressive related taxon.

scholarly journals Use of site-directed mutagenesis of allele-specific PCR primers to identify the GSTM1 A, GSTM1 B, GSTM1 A,B and GSTM1 null polymorphisms at the glutathione S-transferase, GSTM1 locus

1993 ◽  
Vol 295 (1) ◽  
pp. 313-315 ◽  
Author(s):  
A A Fryer ◽  
L Zhao ◽  
J Alldersea ◽  
W R Pearson ◽  
R C Strange
Keyword(s):  
Pcr Primers ◽  
Single Step ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Glutathione S Transferase ◽  
Specific Pcr ◽  
Target Dna ◽  
Allele Specific ◽  
Pcr Method ◽  
Allele Specific Pcr

We describe the identification of the GSTM1 null, GSTM1 A, GSTM1 B and GSTM1 A,B polymorphisms at the glutathione S-transferase GSTM1 locus using a single-step PCR method. Target DNA was amplified using primers to intron 6 and exon 7 with site-directed mutagenesis being used to introduce a restriction site in DNA amplified from GSTM1 *A, thereby allowing differentiation of this allele and GSTM1 *B. The accuracy of this approach in identifying the GSTM1 A, GSTM1 B, GSTM1 A,B and GSTM1 null polymorphisms was confirmed by comparison with, firstly, an established PCR method that distinguishes GSTM1 *0 homozygotes from individuals with the other GSTM1 genotypes and, secondly, GSTM1 phenotypes determined using chromatofocusing.

scholarly journals Detection of mutation delta F508 in the cystic fibrosis gene using allele-specific PCR primers and time-resolved fluorometry.

1992 ◽  
Vol 2 (2) ◽  
pp. 157-162 ◽  
Author(s):  
A Iitia ◽  
E Hogdall ◽  
P Dahlen ◽  
P Hurskainen ◽  
J Vuust ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Pcr Primers ◽  
Cystic Fibrosis Gene ◽  
Time Resolved ◽  
Specific Pcr ◽  
Allele Specific ◽  
Allele Specific Pcr

scholarly journals Nested Allele-Specific PCR Primers Distinguish Genetic Groups of Uncinula necator

1999 ◽  
Vol 65 (9) ◽  
pp. 3950-3954 ◽  
Author(s):  
Christophe Délye ◽  
Valérie Ronchi ◽  
Frédéric Laigret ◽  
Marie-France Corio-Costet
Keyword(s):  
Point Mutations ◽  
Pcr Primers ◽  
Uncinula Necator ◽  
Single Spore ◽  
Group Iii ◽  
Specific Pcr ◽  
Genetic Groups ◽  
Group I ◽  
Allele Specific ◽  
Allele Specific Pcr

ABSTRACT Isolates of the obligately biotrophic fungus Uncinula necator cluster in three distinct genetic groups (groups I, II, and III). We designed PCR primers specific for these groups in order to monitor field populations of U. necator. We used the nucleotide sequences of the gene that encodes eburicol 14α-demethylase (CYP51) and of the ribosomal DNA internal transcribed spacer 1 (ITS1), ITS2, and 5.8S regions. We identified four point mutations (three in CYP51 and one in ITS1) that distinguished groups I and II from group III based on a sample of 132 single-spore isolates originating from Europe, Tunisia, Israel, India, and Australia. We developed a nested allele-specific PCR assay in which the CYP51 point mutations were used to detect and distinguish groups I and II from group III in crude mildewed samples from vineyards. In a preliminary study performed with samples from French vineyards in which isolates belonging to genetic groups I and III were present, we found that a shift from a population composed primarily of group I isolates to a population composed primarily of group III isolates occurred during the grapevine growing season.

Simple and Rapid Detection of Factor V Leiden by Allele-specific PCR Amplification

1996 ◽  
Vol 75 (05) ◽  
pp. 757-759 ◽  
Author(s):  
Rainer Blasczyk ◽  
Markus Ritter ◽  
Christian Thiede ◽  
Jenny Wehling ◽  
Günter Hintz ◽  
...  
Keyword(s):  
Direct Sequencing ◽  
Activated Protein C ◽  
Factor V Leiden ◽  
Pcr Amplification ◽  
Factor V ◽  
Specific Pcr ◽  
Pcr Rflp ◽  
Allele Specific ◽  
Specific Amplification ◽  
Allele Specific Pcr

SummaryResistance to activated protein C is the most common hereditary cause for thrombosis and significantly linked to factor V Leiden. In this study, primers were designed to identify the factor V mutation by allele-specific PCR amplification. 126 patients with thromboembolic events were analysed using this technique, PCR-RFLP and direct sequencing. The concordance between these techniques was 100%. In 27 patients a heterozygous factor VGln506 mutation was detected, whereas one patient with recurrent thromboembolism was homozygous for the point mutation. Due to its time- and cost-saving features allele-specific amplification should be considered for screening of factor VGln506.

scholarly journals Development of genic KASP SNP markers from RNA-Seq data for map-based cloning and marker-assisted selection in maize

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Zhengjie Chen ◽  
Dengguo Tang ◽  
Jixing Ni ◽  
Peng Li ◽  
Le Wang ◽  
...  
Keyword(s):  
Marker Assisted Selection ◽  
Inbred Lines ◽  
Average Density ◽  
Snp Markers ◽  
Data Sets ◽  
Rna Seq ◽  
Specific Pcr ◽  
Maize Inbred Lines ◽  
Allele Specific ◽  
Allele Specific Pcr

Abstract Background Maize is one of the most important field crops in the world. Most of the key agronomic traits, including yield traits and plant architecture traits, are quantitative. Fine mapping of genes/ quantitative trait loci (QTL) influencing a key trait is essential for marker-assisted selection (MAS) in maize breeding. However, the SNP markers with high density and high polymorphism are lacking, especially kompetitive allele specific PCR (KASP) SNP markers that can be used for automatic genotyping. To date, a large volume of sequencing data has been produced by the next generation sequencing technology, which provides a good pool of SNP loci for development of SNP markers. In this study, we carried out a multi-step screening method to identify kompetitive allele specific PCR (KASP) SNP markers based on the RNA-Seq data sets of 368 maize inbred lines. Results A total of 2,948,985 SNPs were identified in the high-throughput RNA-Seq data sets with the average density of 1.4 SNP/kb. Of these, 71,311 KASP SNP markers (the average density of 34 KASP SNP/Mb) were developed based on the strict criteria: unique genomic region, bi-allelic, polymorphism information content (PIC) value ≥0.4, and conserved primer sequences, and were mapped on 16,161 genes. These 16,161 genes were annotated to 52 gene ontology (GO) terms, including most of primary and secondary metabolic pathways. Subsequently, the 50 KASP SNP markers with the PIC values ranging from 0.14 to 0.5 in 368 RNA-Seq data sets and with polymorphism between the maize inbred lines 1212 and B73 in in silico analysis were selected to experimentally validate the accuracy and polymorphism of SNPs, resulted in 46 SNPs (92.00%) showed polymorphism between the maize inbred lines 1212 and B73. Moreover, these 46 polymorphic SNPs were utilized to genotype the other 20 maize inbred lines, with all 46 SNPs showing polymorphism in the 20 maize inbred lines, and the PIC value of each SNP was 0.11 to 0.50 with an average of 0.35. The results suggested that the KASP SNP markers developed in this study were accurate and polymorphic. Conclusions These high-density polymorphic KASP SNP markers will be a valuable resource for map-based cloning of QTL/genes and marker-assisted selection in maize. Furthermore, the method used to develop SNP markers in maize can also be applied in other species.

scholarly journals Two User-Friendly Molecular Markers Developed for the Identification of Hybrid Lethality Genes in Brassica oleracea

Agronomy ◽  
2021 ◽  
Vol 11 (5) ◽  
pp. 982
Author(s):  
Zhiliang Xiao ◽  
Congcong Kong ◽  
Fengqing Han ◽  
Limei Yang ◽  
Mu Zhuang ◽  
...  
Keyword(s):  
Molecular Markers ◽  
Brassica Oleracea ◽  
Rapid Identification ◽  
Inbred Lines ◽  
Hybrid Lethality ◽  
Specific Pcr ◽  
Allele Specific ◽  
User Friendly ◽  
Allele Specific Pcr ◽  
Kasp Marker

Cabbage (Brassica oleracea) is an important vegetable crop that is cultivated worldwide. Previously, we reported the identification of two dominant complementary hybrid lethality (HL) genes in cabbage that could result in the death of hybrids. To avoid such losses in the breeding process, we attempted to develop molecular markers to identify HL lines. Among 54 previous mapping markers closely linked to BoHL1 or BoHL2, only six markers for BoHL2 were available in eight cabbage lines (two BoHL1 lines; three BoHL2 lines; three lines without BoHL); however, they were neither universal nor user-friendly in more inbred lines. To develop more accurate markers, these cabbage lines were resequenced at an ~20× depth to obtain more nucleotide variations in the mapping regions. Then, an InDel in BoHL1 and a single-nucleotide polymorphism (SNP) in BoHL2 were identified, and the corresponding InDel marker MBoHL1 and the competitive allele-specific PCR (KASP) marker KBoHL2 were developed and showed 100% accuracy in eight inbred lines. Moreover, we identified 138 cabbage lines using the two markers, among which one inbred line carried BoHL1 and 11 inbred lines carried BoHL2. All of the lethal line genotypes obtained with the two markers matched the phenotype. Two markers were highly reliable for the rapid identification of HL genes in cabbage.

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