The common methods that are presently used to identify Vibrio harveyi include microscopic examination and biochemical, immunological and PCR-based assays. These methods require technical expertise, and can be time-consuming. A rapid method is required for the high-throughput screening of large number of samples. As such, we have developed a rapid, simple yet sensitive and specific detection method based on the use of the loop-mediated isothermal amplification (LAMP) of DNA. A set of six primers, i.e., two outer, two inner and two loop primers, was designed based on the in silico analysis of a large pool of 39 strains of the toxR gene sequence of V. harveyi. The addition of the loop primers decreased the reaction time of the LAMP by more than half. Furthermore, with the application of SYBR Green, the result can be obtained as quickly as in 10 to 15 min without the need of gel electrophoresis. The specificity of the method primers was then determined by performing LAMP with Vibrio and non-Vibrio samples. LAMP has a greater sensitivity than PCR reaction. The sensitivity of PCR was at 0.6 pg concentration of V. harveyi recombinant plasmid DNA standard, while LAMP was able to detect lower amounts even at 0.6 fg. The development of the LAMP assay will provide a valuable tool for the high-throughput rapid detection of V. harveyi contamination both in laboratories and in the field.
Harumanis' (Mangifera indica L) is one of the mango cultivars which has high market value because of the excellent quality of the fruit which has attractive color, good aroma, delicious taste and high nutritive values. In this study, fifty accessions from five different collection sites which belonging to North (
For the last twenty years, many newly described spiders were collected from Malaysia and in fact, more than 11,000 species were recorded in Peninsular Malaysia as well as in Sabah and Sarawak states. Scientists have put an immense effort on untangling the spider biology from its physical structure and behavior to silks and venoms. However, working with spiders is impeded by the difficulties in species identification via solely morphological methods. Thus, DNA barcoding is an alternative technique that employs standard fragment to facilitate species identification. Isolation of genomic DNA from three Malaysian spiders were performed using NucleoSpin® DNA insect extraction kit. Amplification of reference mitochondrial cytochrome oxidase I (COI) gene employing PCR with two set of primers followed by the DNA sequencing and validation through phylogenetic analysis were carried out. The commercial extraction kit was effective for the recovery of good quality of intact genomic DNA band as indicated by the integrity analysis. Both set of primers successfully amplified 100% of the samples with approximately 600 – 700 bp of PCR products. The obtained sequences (610 bp to 692 bp) were compared with the sequences available in Gene Bank. BLAST and phylogenetic analysis revealed that the analyzed individual samples belong to Nephila pilipes, Neoscona nautica and Crossopriza lyoni, respectively. Phylogenetic analysis provided unique insight into the evolutionary relationship of each analyzed sample. This study aids in an accurate identification of the selected local spider species at molecular level using the COI gene.
Harumanis is one of the main signatures of Perlis with regards to its delightful taste, pleasant aroma and expensive price. Harumanis authenticity and productivity had become the remarks among the farmers, entrepreneurs, consumers and plant breeders due to the existence of morphological characteristics variation among the fruits and high production cost. Assessment of Harumanis morphological characteristics of natural population and different tree ages may represent a possible source of important characteristics for development and breeding purposes of Harumanis . The aim of this study is to evaluate the morphological variation of Harumanis collected from different location in Perlis and tree age. A total of 150 Harumanis fruits from 50 trees with three different stages of development (young, middle-aged and old) were characterised using 11 traits; 10 quantitative and one qualitative morphological trait. The ANOVA analyses in combination with Dunn’s pairwise and Kruskal-Wallis multiple comparison test able to point out the existence of environmental factor and age influence towards the significant different of identified morphological traits except for Total Soluble Solid (TSS) and pulp percentage. Five clusters of 50 Harumanis accessions reflect a grouping pattern which not according to neither geographical region nor age. The result of Principal Component Analysis (PCA) using the first two principal components (PCs) provided a good approximation of the data explaining 84.09% of the total variance which majorly contributed by parameters of weight, fruit dimensional characteristics, peel percentage and hue angle, h . Preliminary screening of important morphological characteristics which contribute to the phenotypic diversity of Harumanis is successfully achieved. The findings can be employed by the plant breeders and farmers for the establishment of standard grading of Harumanis and advancement of breeding crop of Harumanis in future.
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