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SNP genotyping and its applications
Advancements in the field of SNP genotyping and its applications
a) SNP genotyping has been used in identifying useful agricultural traits often controlled by multiple genes
b) SNP genotyping to identify gene variants for herbicide resistance
1) SNP analysis combined with Simple sequence repeats (SSR) to identify QTLs
A group of scientists combined simple sequence repeat (SSR) markers along with the bulked segregant and SNP analyses (BSA and SNP), to identify QTL by inclusive composite interval mapping analysis. these QTLs can be further used for fine mapping and marker-assisted selection (MAS) to achieve long lasting resistance to certain diseases like Leaf rust (Puccinia recondite f. sp. Tritici).
2) SNP genotyping combined with quantitative genotyping PCR (qgPCR) to help CRISPR editors
Did you know there have been reports of accidental CRISPR-induced on-target effects (OnTEs) leading to Loss of heterozygosity (LOH)? OnTEs are often mischaracterized by standard Sanger sequencing as correctly edited. OnTEs can severely affect the phenotype of the edited cells or organisms. This may challenge the reliability of the whole technique and thus a scientific group has come up with a protocol to detect these OnTEs using SNP genotyping that can help detect the occurrence of LOH around the edited locus. The protocol begins with exclusion of clones with monoallelic OnTEs and hemizygous editing by determining the number of intact alleles at the target site. This is done by quantitative genotyping PCR (qgPCR) of the genomic DNA. The LOH are then identified by genotyping neighboring single-nucleotide polymorphisms (SNPs).
3) Enhanced SNP genotyping platforms with hs-CRISPR system
In a recent research study, the CRISPR/Cas13a system was enhanced with a strategy called hs-CRISPR. The system is comparable to ordinary crRNA:
a) it shows high on-target affinity of Cas13a, while suppressing off-target activity, using CRISPR RNA (crRNA) with hairpin spacer.
b) It has at least 3-fold higher sensitivity and discriminatory ability of Cas13a system.
c) It can be performed within 1 hour.
d) It is compatible with the traditional Cas13a system.
The increased specificity of CRISPR/Cas13a can be used to extend SNP genotyping. With genotyping based on the improved hs-CRISPR (hairpin spacer-CRISPR) system, the techniques are simpler to use, easier to program, and thus simply better than it is today.
4) SNP-seq genotyping technology for vegetable variety identification using DNA fingerprinting
In 2020, a study of 261 cucumber varieties was conducted in China to evaluate the application of SNP-seq genotyping technology in variety identification. This was done by constructing a target SNP-seq library.
GENAXXON not only offers SNP Polymerase MasterMix but also SNP Polymerase and SNP Taq Polymerase to compliment aforementioned technologies to help take your precision to another level. Genaxxon products can be combined with similar protocols to achieve more specific results.
Not only this, what if we told you that it is accurately possible to distinguish mutant alleles from wild type alleles and that to, without sequencing?
Yes, it is possible using GENAXXON SNP Polymerases.
The GENAXXON SNP Pol DNA Polymerase, GENAXXON SNP PolTaq DNA-Polymerase and our GENAXXON SNP Pol DNA-Polymerase MasterMix distinguishes with high specificity, whether a mismatch of the primer template complex is present or not. The mismatch (point mutation) must be at the 3 'end of the primer. And the trick here is that the Polymerase simply does not amplify in the case of a mismatch. How perfect is that? But this was just one of the several applications of our SNP polymerase.
The other applications of GENAXXON SNP polymerases are:
- Monitoring, verification and detection of point mutations
- Validation of CRISPR/Cas9 products
- Quantification of mutations (e.g. NGS results)
- Allele-specific amplification (ASA)
- Methylation specific PCRs (MSP) after bisulfite treated DNA (CpG methylation sides)
- HLA genotyping Micro sequencing
- Realtime PCR with hydrolysis probes
- Realtime multiplex PCRs
http://www.plantbreedbio.org/journal/view.html?doi=10.9787/PBB.2014.2.3.195
https://link.springer.com/article/10.1186/s12870-015-0689-9
https://onlinelibrary.wiley.com/doi/full/10.1002/advs.202003611
https://www.nature.com/articles/s41598-020-62518-6
Methods to study allele frequencies - and distribution in the acetolactate synthase (ALS) gene in target site resistance in Echinochloa crus-galli.
Fabian Runge et al. (2020), IdentXX GmbH, Stuttgart, Germany.
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