Standard Agarose Gel Electrophoresis Research Articles

Streamlining the Detection of FLT3 Internal Tandem Duplication (FLT3-ITD) Using the Cepheid Genexpert® Automated System

Objectives: The development of targeted therapies for Acute Myeloid leukemia (AML) demands fast and reliable diagnostic tests. Specifically, FMS-like tyrosine kinase 3 (FLT3-ITD), comprising nearly 25% of all AML cases, is the most common targetable mutation in AML. Current diagnosis of FLT3-ITD by PCR involves numerous laborious steps, with bench-top steps involving time-consuming whole blood processing, DNA isolation, plate-based PCR and subsequent fragment analysis by capillary electrophoresis. Consequently this adds significant overhead, time-consuming personnel training, and numerous steps which could lead to errors, all of which increase the resource burden and time to result. The objective of this study was to create an automated FLT3-ITD diagnostic prototype that is faster, requires less hands-on time that can detect all clinically relevant ITD lengths. Methods: We have developed a simplified FLT3-ITD prototype assay using the Cepheid GeneXpert® automated platform for FLT3-ITD detection. The prototype substantially streamlines the process, using a less than 30 minutes off-board time to prepare a universal blood lysate that is compatible with all Cepheid Hematology Oncology products. Once the sample is loaded into the cartridge and inserted into the instrument, the cartridge performs automated DNA isolation, purification, and qPCR. A Taqman-based probe for ABL is utilized in the qPCR step as a PCR control to ensure that the blood processing and qPCR was successful and avoid wasted downstream steps. Quantitative fragment analysis can easily be performed on the PCR product using capillary electrophoresis (CE) for precise allelic ratio (AR) callout. Additionally, the PCR product is compatible be run on a bioanalyzer or standard Agarose Gel-Electrophoresis if available. Results: The developed automated FLT3-ITD diagnostic prototype demonstrated significant improvements over the existing methodologies with substantially reduced time to result and simplicity in use. To assess the sensitivity of this prototype, we used MV4-11 cancer cells harboring a 30 bp ITD and CRISPR-CAS engineered human cell lines with 3 bp, 21 bp, and 75 bp FLT3 ITD insertions, comprising the most commonly detected ITD lengths spiked into EDTA blood as a test material. For these ITD's, we observed sensitivity and linearity down to 0.01-0.02 by CE for the ITDs tested. Importantly, the prototype detected all lengths of ITD mutations with inserts from 3bp to 300 bp using plasmid test materials. Conclusion: We have successfully developed a novel FLT3-ITD diagnostic prototype that has several major advantages over existing methodologies. The prototype significantly reduces the time and hands-on effort required for analysis, streamlining the diagnostic process for same day results. The larger sample volume used allows improved sensitivity to ensure accurate detection, even at low allelic frequencies. Additionally, the prototype requires fewer reagents in that sample prep and qPCR are performed in a single cartridge and run hands-free. Downstream analysis of amplified products can be easily performed and validated using capillary electrophoresis, bioanalyzer, or gel electrophoresis. Finally, because this prototype uses the same universal blood lysate as other AML tests, it has the potential to improve future diagnostic therapeutic interventions in the clinic.

Appearance of ghost bands in gel electrophoresis depending on agarose concentration.

Standard agarose gel electrophoresis is a widely used method to analyse diversity of nucleic acids. Certain conditions, however, may give rise to artefactual bands. We report on artefactual bands frequently occurring, especially when partially homologous nucleic acids, such as splicing variants of DNA transcripts, are analysed simultaneously. Interestingly, to some extent agarose concentration may influence the occurrence of artefactual bands.

Tetra-Primer Amplification Refractory Mutation System (T-ARMS).

Single-nucleotide polymorphisms (SNPs), the most abundant genetic variation in the population, have become the molecular marker of choice. Generally, the efficient detection of SNPs requires specialized costly equipment. Although there are a few strategies for detecting SNPs through polymerase chain reaction, followed by restriction enzyme digestion and agarose gel electrophoresis, these methods are time-consuming and might be less diagnostic. Interestingly, the tetra primer amplification refractory mutation system (T-ARMS) strategy utilizes a pair of allele-specific primers in a single PCR for the diagnostic detection of SNPs in a codominant manner through standard agarose gel electrophoresis. The simplicity and robustness of the strategy have inspired the researchers to adopt this low-cost method of SNP detection in different crop plants. Here, we have described the principle, methods, and conditions for the T-ARMS strategy. The described methodology starts from the isolation of genomic DNA and ends with the post-PCR analysis of refractory amplicons in standard agarose gel electrophoresis. The limitations and future perspectives are also discussed. Taken together, T-ARMS evolves as a method of choice for low-cost SNP detection in plants.

Gel purification of gDNA for next-generation sequencing applications.

We demonstrate thatgDNA can be conveniently and efficiently isolated and purified using standard agarose gel electrophoresis, band excision and gel purification. This method yields a substantial amount at microgram levels of gDNA per gel cleanup with high purity. An RNaseA treatment step can be omitted. The quality of gDNA is suitable for next-generation sequencing, resulting in >10Mb reads and high-quality read data (Phred score>28 up to 100 of 150 base reads). Furthermore, the gDNA can be kept intact in a gel slice for several days. This method has been tested for dictyostelids, bacteria and plants.

ANALYSIS OF SERUM PROTEINS IN HEALTHY GIANT PANDAS (AILUROPODA MELANOLEUCA) UNDER MANAGED CARE.

Electrophoresis is a practical diagnostic tool for the identification of changes in serum protein fractions, which can be associated with a variety of diseases. Protein electrophoresis studies in Ursidae are limited, and currently no published fraction values are available for the giant panda (Ailuropoda melanoleuca). The aim of this study was to describe the serum protein fractions in the giant panda using both capillary zone electrophoresis (CZE) and standard agarose gel electrophoresis (AGE) techniques. Serum samples from nine healthy giant pandas (n = 19) were used for this study. Samples were evaluated using CZE and standard AGE. The CZE procedure successfully resolved serum proteins into seven fractions: prealbumin; albumin; and α1-, α2-, β1-, β2-, and γ-globulin; while AGE separated serum into only six protein fractions: prealbumin; albumin; α1-, α2-, and β-globulins; and γ-globulin. These data will serve as a preliminary baseline for further studies and provide insight for the medical management of giant pandas.

Comparative Evaluation of Apoptotic Effects of Vincristine, Cisplatin and Their Hydrogel Scaffolds on HeLa Cell Line

The present study has been undertaken for comparative evaluation of apoptotic effects of vincristine, cisplatin and their hydrogel scaffolds on HeLa cell line. Apoptosis can be visualized as a ladder pattern due to DNA cleavage by the activation of a nuclear endonuclease by standard agarose gel electrophoresis. Results showed the formation of the DNA ladder in gel electrophoresis by induction of apoptosis in HeLa cell line treated with 20 µg/ml of vincristine alone, 20 µg/ml cisplatin alone and their scaffolds, a typical ladder pattern of internucleosomal fragmentation was observed in cells after 24 hrs treatment. DNA ladder assay of genomic DNA from HeLa cells was isolated and subjected to electrophoresis in agarose gel (1.8%) and 1kb DNA ladder. DNA fragments were visualized under a UV trans-illuminator and compared with a standard marker. On the basis of the parameters studied, it was concluded that the formation of the DNA ladder in gel electrophoresis by induction of apoptosis in HeLa cell line treated with vincristine, cisplatin alone and their scaffolds showed best fragmentation in vincristine and cisplatin, however, their scaffolds are moderately effective when used in appropriate dose on HeLa cell line.

Development and validation of the OVATE gene-based functional marker to assist fruit shape selection in tomato.

The online version contains supplementary material available at 10.1007/s13205-021-03029-7.

A genotyping method combining primer competition PCR with HRM analysis to identify point mutations in Duchenne animal models

Dystrophin-null sapje zebrafish is an excellent model for better understanding the pathological mechanisms underlying Duchenne muscular dystrophy, and it has recently arisen as a powerful tool for high-throughput screening of therapeutic candidates for this disease. While dystrophic phenotype in sapje larvae can be easily detected by birefringence, zebrafish genotyping is necessary for drug screening experiments, where the potential rescue of larvae phenotype is the primary outcome. Genotyping is also desirable during colony husbandry since heterozygous progenitors need to be selected. Currently, sapje zebrafish are genotyped through techniques involving sequencing or multi-step PCR, which are often costly, tedious, or require special equipment. Here we report a simple, precise, cost-effective, and versatile PCR genotyping method based on primer competition. Genotypes can be resolved by standard agarose gel electrophoresis and high-resolution melt assay, the latter being especially useful for genotyping a large number of samples. Our approach has shown high sensitivity, specificity, and reproducibility in detecting the A/T point mutation in sapje zebrafish and the C/T mutation in the mdx mouse model of Duchenne. Hence, this method can be applied to other single nucleotide substitutions and may be further optimized to detect small insertions and deletions. Given its robust performance with crude DNA extracts, our strategy may be particularly well-suited for detecting single nucleotide variants in poor-quality samples such as ancient DNA or DNA from formalin-fixed, paraffin-embedded material.

Easy-to-Use InDel Markers for Genetic Mapping between Col-0 and Ler-0 Accessions of Arabidopsis thaliana.

Map-based gene cloning has played a key role in many genetic studies using the model plant, Arabidopsis thaliana. In the post- next generation sequencing era, identification of point mutations and their corresponding genes is increasingly becoming a powerful and important approach to define plant gene function. To perform initial mapping experiments efficiently on Arabidopsis mutants, enrichment of easy-to-use and reliable polymorphic DNA markers would be desirable. We present here a list of InDel polymorphic markers between Col-0 and Ler-0 accessions that can be detected in standard agarose gel electrophoresis.

Detecting miRNAs Using DNA Nanoswitches

miRNAs are short, noncoding RNAs (18–25 nt) and their expression levels in tissues, cells, and bodily fluids serve as biomarkers for diagnosing cellular events or diseases. The miRNA-activated conditional looping of engineered switches (miRacles) assay is based on a conformational change of a DNA nanoswitch. Recognition and binding of a target miRNA to the detectors reconfigures the switch from the linear ‘off’ state to the looped ‘on’ state. The two states can be quantified using standard agarose gel electrophoresis and gel stains, where the detection signal arises from the intensity of the looped nanoswitch.

Extraction and purification of RNA from human carious dentine: an approach to enable bacterial gene expression studies

Background: RNA isolation from bacteria within dentine caries lesions could be difficult due to reduced amount of collectable biomass and high mRNA instability. Attempting to overcome this challenge we describe one protocol developed to extract and purify total RNA from dentine lesions. Objective: customize a bacterial RNA extraction and purification method from human carious dentine. Methods: quantity and purity of extracted RNA were measured with a microvolume UV-VIS spectrophotometer, RNA integrity was assessed by standard denaturing agarose gel electrophoresis and images were captured under ultraviolet light with camera and analyzed. DNase treatment removed genomic DNA and an additional step of purification was carried out in silica spin column. Results: final yield (ng/μl) was 67.01 ± 22.33, absorbance ratio A260/A280 = 2.0 ± 0.07 and RNA integrity were obtained. The purified samples were reversely transcribed and the expression of atpD and fabM gene from Streptococcus mutans analyzed by quantitative real-time PCR. Conclusion: the extraction methodology developed produced high-quality RNA from dentine microbiota for transcriptional analysis.

InDel markers for monitoring the introgression of downy mildew resistance from wild relatives into grape varieties

We identified haplotype-tagging insertion/deletions (InDels) for downy mildew resistance (Rpv3-1) in grapevine and converted them into InDel markers. InDel-25,941 and InDel-26,032 were validated by fragment analysis via capillary electrophoresis in 174 varieties of Vitis vinifera, 50 resistant varieties of the ‘Seibel 4614’ lineage that share Rpv3-1 by descent, and in 83 Vitis accessions. Amplicon sequencing of ancestral and derived alleles revealed that both mutations were caused by deletions. The 25,941-deletion is most likely recent. The derived allele is present only in resistant varieties obtained from ‘Seibel 4614’ and has originated in North American populations through two successive deletions within a predicted multiple stem-loop ssDNA structure, consisting of three nearby short inverted repeats, which shortened the ancestral DNA stepwise. The 26,032-deletion is more ancient. The derived allele is always present in resistant varieties of the ‘Seibel 4614’ lineage, completely absent from V. vinifera, not found in other North American accessions, and rarely present in Asian species. It may have originated in a common ancestral population before the continental disjunction, followed by incomplete lineage sorting, or in either lineage followed by introgression via secondary contacts. Genotyping with these markers does not require special instruments or chemistry for routine screening in breeding practice. Differences in amplicon size between grapes that carry or do not carry Rpv3-1 are detectable via standard agarose gel electrophoresis, or classical melting curve analysis using nonsaturating fluorescent dyes. The recombination rate between each marker and the trait locus is 0.118% for InDel-25,941 and 0.071% for InDel-26,032.

Ethidium Bromide Modifies The Agarose Electrophoretic Mobility of CAG•CTG Alternative DNA Structures Generated by PCR.

The abnormal expansion of unstable simple sequence DNA repeats can cause human disease through a variety of mechanisms, including gene loss-of-function, toxic gain-of-function of the encoded protein and toxicity of the repeat-containing RNA transcript. Disease-associated unstable DNA repeats display unusual biophysical properties, including the ability to adopt non-B-DNA structures. CAG•CTG trinucleotide sequences, in particular, have been most extensively studied and they can fold into slipped-stranded DNA structures, which have been proposed as mutation intermediates in repeat size expansion. Here, we describe a simple assay to detect unusual DNA structures generated by PCR amplification, based on their slow electrophoretic migration in agarose and on the effects of ethidium bromide on the mobility of structural isoforms through agarose gels. Notably, the inclusion of ethidium bromide in agarose gels and running buffer eliminates the detection of additional slow-migrating DNA species, which are detected in the absence of the intercalating dye and may be incorrectly classified as mutant alleles with larger than actual expansion sizes. Denaturing and re-annealing experiments confirmed the slipped-stranded nature of the additional DNA species observed in agarose gels. Thus, we have shown that genuine non-B-DNA conformations are generated during standard PCR amplification of CAG•CTG sequences and detected by agarose gel electrophoresis. In contrast, ethidium bromide does not change the multi-band electrophoretic profiles of repeat-containing PCR products through native polyacrylamide gels. These data have implications for the analysis of trinucleotide repeat DNA and possibly other types of unstable repetitive DNA sequences by standard agarose gel electrophoresis in diagnostic and research protocols. We suggest that proper sizing of CAG•CTG PCR products in agarose gels should be performed in the presence of ethidium bromide.

Undergraduate physics laboratory: Electrophoresis in chromatography paper

An experiment studying the physical principles of electrophoresis in liquids was developed for an undergraduate laboratory. We have improved upon the standard agarose gel electrophoresis experimental regime with a straightforward and cost-effective procedure, in which drops of widely available black food coloring were separated by electric field into their dye components on strips of chromatography paper soaked in a baking soda/water solution. Terminal velocities of seven student-safe dyes were measured as a function of the electric potential applied along the strips. The molecular mobility was introduced and calculated by analyzing data for a single dye. Sources of systematic and random errors were investigated.

Genotyping of α-thalassemias by the colorimetric nanogold probes

BackgroundThe novel colorimetric nanogold probe was created to genotype subgroups of the mostly found α-thalassemias. They are α-thalassemia 1 (SEA and THAI deletion) and α-thalassemia 2 (3.7-kb and 4.2-kb deletion). MethodsThe genotyping was performed by two-steps hybridizations. First step was hybridization of target DNA with the nanogold mixed probes of either α-thalassemia 1 or α-thalassemia 2. No hybridization in both reactions showing blue color indicated absence of abnormal genes causing these α-thalassemias. Positive reaction showing either red or purple color was further analyzed in second hybridization with the nanogold single probe. Positive of α-thalassemia 1 was genotyped with the single probes of both SEA and THAI deletion while those of α-thalassemia 2 were genotyped with both 3.7-kb and 4.2-kb deletion. ResultsGenotypic potency of the nanogold mixed and single probes was evaluated using both known diagnosed and suspected clinical samples. The results by naked eye were consistence with those analyzed by standard agarose gel electrophoresis. ConclusionsPotency of the colorimetric nanogold α-thalassemia probes was accurate, precise, sensitive, specific, simple, cheap and field applicable. Color reaction was simply visualized by naked eye. This development is an example of colorimetric molecular diagnosis which can be applied in any genetic detection.

High-throughput multiplex MIRU-VNTR typing of Mycobacterium bovis

Spoligotyping is the most widely used method for genotyping Mycobacterium bovis (M. bovis). However, its discriminatory power varies widely between countries. MIRU-VNTR typing could be a promising alternative, although it generally requires the time consuming and laborious simplex PCR assays using standard agarose gel electrophoresis. The accuracy of this approach depends on good standardization and a certain degree of expertise. This study presents a version of MIRU-VNTR based on three triplex PCRs with automatic allelic assignation of the products analyzed in capillary electrophoresis. The technique was prospectively applied to 44 M. bovis and two Mycobacterium caprae (M. caprae) isolates, and 22 different MIRU-VNTRtypes were obtained; with spoligotyping, only 14 different types were obtained. The proposal makes it possible to shorten response times, automate procedures, and increase accuracy, thus minimizing errors in assigning genotypes. It would enable the switch from a standard limited method of genotyping M. bovis to a high-throughput discriminatory fingerprinting approach.

Development and characterization of 96 microsatellite markers suitable for QTL mapping and accession control in an Arabidopsis core collection

BackgroundTo identify plant genes involved in various key traits, QTL mapping is a powerful approach. This approach is based on the use of mapped molecular markers to identify genomic regions controlling quantitative traits followed by a fine mapping and eventually positional cloning of candidate genes. Mapping technologies using SNP markers are still rather expensive and not feasible in every laboratory. In contrast, microsatellite (also called SSR for Simple Sequence Repeat) markers are technologically less demanding and less costly for any laboratory interested in genetic mapping.ResultsIn this study, we present the development and the characterization of a panel of 96 highly polymorphic SSR markers along the Arabidopsis thaliana genome allowing QTL mapping among accessions of the Versailles 24 core collection that covers a high percentage of the A. thaliana genetic diversity. These markers can be used for any QTL mapping analysis involving any of these accessions. We optimized the use of these markers in order to reveal polymorphism using standard PCR conditions and agarose gel electrophoresis. In addition, we showed that the use of only three of these markers allows differentiating all 24 accessions which makes this set of markers a powerful tool to control accession identity or any cross between any of these accessions.ConclusionThe set of SSR markers developed in this study provides a simple and efficient tool for any laboratory focusing on QTL mapping in A. thaliana and a simple means to control seed stock or crosses between accessions.

In vitro antioxidant potential and deoxyribonucleic acid protecting activity of CNB-001, a novel pyrazole derivative of curcumin

Background: Free radicals are underpinned to initiate cascade of toxic events leading to oxidative stress and resultant cell death in many neurodegenerative disorders. Now-a-days antioxidants have become mandatory in the treatment of various diseases apart from the drug's modes of action. CNB-001, a novel hybrid molecule synthesized by combining curcumin and cyclohexyl bisphenol A is known to possess various biological activities, but the antioxidative property of the compound has not yet been elucidated. Aim: The present study is aimed to analyze various free radicals scavenging by employing in vitro antioxidant assays and to evaluate the deoxyribonucleic acid (DNA) protecting the ability of CNB-001 against hydroxyl radicals. Materials and methods: The in vitro antioxidant potential of CNB-001 was evaluated by analyzing its ability to scavenge DPPH, ABTS, nitric oxide, superoxide, hydrogen peroxide, superoxide anion, hydroxyl, hydrogen peroxide radicals and reducing power using spectroscopic method. The DNA protecting activity of CNB-001 was also evaluated on pUC19 plasmid DNA subjected to hydroxyl radicals using standard agarose gel electrophoresis. Results: From the assays, it was observed that CNB-001 scavenged free radicals effectively in a dose dependent manner. CNB-001 scavenged 2,2-diphenyl-1-picrylhydrazyl (IC50 = 44.99 μg/ml), 2,2-azinobis (3-ethylbenzothiazoline-6-sulfonic acid) (IC50 = 17.99 μg/ml), nitric oxide (IC50 = 1.36 μg/ml), superoxide radical (IC50 = 77.17 μg/ml), hydrogen peroxide (IC50 = 492.7 μg/ml), superoxide (IC50 = 36.92 μg/ml) and hydroxyl (IC50 = 456.5 μg/ml) radicals effectively and the reducing power was found to be 11.53 μg/ml. CNB-001 showed considerable protecting activity against plasmid DNA (pUC19) strand scission by •OH at dose dependent manner. Conclusion: Results from these assays concluded that CNB-001 has a good antioxidant potential by reducing reactive oxygen and reactive nitrogen radicals and it showed significant protecting activity against DNA scission by hydroxyl radicals. Hence, CNB-001 can be further developed as potential drug for free radical induced neurodegenerative disorders.

Abstract 1835: A pilot study of long non-coding RNA expression profile in esophageal cancer.

Abstract Background LncRNAs, range from 200 nt to >100 kilobases (kb), demonstrate a wide range of structural and functional roles through the regulation of a metastable epigenome and transcriptional network. Recently, accumulating evidence indicates aberrant expression of LncRNAs may play an important role in cancer biology. Objective To study the profile of differentially expressed LncRNAs in esophageal cancer tissues and the adjacent normal tissues, and to find out the possible function of LncRNAs involved in the process of tumor development preliminarily. Methods The total RNA were extracted from esophageal cancer tissues and the adjacent normal tissues and were quantified by the NanoDrop ND-1000 and standard denaturing agarose gel electrophoresis. Each sample was amplified and transcribed into fluorescent cRNA along the entire length of the transcripts without 3’ bias utilizing a random priming method. The labeled cRNAs were hybridized onto the Human LncRNA Array v2.0 (8 x 60K, Arraystar) including the global profiling of 33,045 human LncRNAs and 30,215 coding transcripts and then were scanned by the Agilent Scanner G2505B. Quantile normalization and subsequent data processing were performed using the GeneSpring GX v11.5.1 software package (Agilent Technologies). Differentially expressed LncRNAs and mRNAs with statistical significance were identified through Volcano Plot filtering or Fold Change filtering. Pathway analysis and GO analysis were applied to determine the roles of those differentially expressed mRNAs played in biological pathways or GO terms. Finally, Hierarchical Clustering was performed to show the distinguishable LncRNAs and mRNAs expression pattern among samples. Results With a criterion of Fold Change >= 2.0 and P-value <= 0.05, 1019 of differentially expressed LncRNAs were identified in esophageal cancer tissues, including 399 of overexpressed LncRNAs and 620 of downexpressed LncRNAs. The top 10 of aberrant LncRNAs were AK022914, chr10:127700955-127703336, LOC284233, RP11-38M15.6, RNU12, FAM25E, C21orf81, AK095500, AC009336.24, and RP11-397A15.4. Using GENCODE annotation of human genes, 11 of differentially expressed enhancer-like LncRNAs and their nearby coding genes were identified. In addition, there were 25 of differentially expressed lincRNAs and nearby coding gene pairs associated with esophageal cancer according to John Rinn reported lincRNAs profiling. The differentially expressed transcripts were further identified and the matched coding genes adjacent to differentially expressed LncRNAs (distance < 300kb) showed participating in the processes of cell cycle, cell differentiation, protease activities, and et al according to GO analysis and Pathway analysis. Conclusions The results suggests a specific profile of differentially expressed LncRNAs in esophageal cancer, which function prediction implies the broad participation in the processes of oncogenesis. Citation Format: Ran Liu, Miao Yang, Juan Liao, Yi Wang, Enchun Pan, Wei Guo, Lihong Yin, Yuepu Pu. A pilot study of long non-coding RNA expression profile in esophageal cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1835. doi:10.1158/1538-7445.AM2013-1835 Note: This abstract was not presented at the AACR Annual Meeting 2013 because the presenter was unable to attend.

Development of New CAPS/dCAPS and SNAP Markers for Rice Eating Quality

Rice eating quality traits are very complex and essential to be evaluated not only through physicochemical analysis and sensory test but also by PCR-based marker approach. To date, simple markers based on single nucleotide polymorphism (SNP) discovery to evaluate eating quality of cooked rice are still limited. Thus, the aims of this study were to develop PCR-based markers, called SNAP (single nucleotide amplified polymorphism) as alternative markers of cleaved amplified polymorphic sequence/derived cleaved amplified polymorphic sequence (CAPS/dCAPS). Four primer pairs specific to targeted alleles (CAPS/dCAPS and SNAP) of four loci were successfully designed based on the discovered SNPs according to the eating quality-QTL and searching genomic database. The primer pairs were able to identify alleles corresponding loci among indica and japonica varieties with diverse palatability (overall eating quality). There was consistent allele pattern produced by SNAP and CAPS/dCAPS for the same base mutation. The SNAP marker for rice eating quality trait could be easily assayed by standard agarose gel electrophoresis, allowing to increase the advantage of genotyping methods. Moreover, the SNAP markers together with our previous developed markers which were recommended as applicable marker set for evaluation of rice eating quality, will facilitate as marker-assisted selection for rice breeding program.