Applications of DNA Microsatelite Markers Intilapia Culture

In Malaysia, the role of aquaculture in fish production is anticipated to increase as marine fish catches have already exceeded the maximum sustainable yield. In order to address this issue, aquaculture is being developed on a commercial scale. Fish farming in Malaysia is focused on providing the fish grower with the best fingerlings, with uniformity and fast growth rates. There is great necessity for genetic evaluation, monitoring of stocks and application of appropriate breeding approaches if tilapia production is to continue to meet market demands. With the aid of microsatellite markers, stock integrity and genetic variability can be examined. Three major experiments were carried out; a population genetic study on eight populations of tilapia. quantitative genetics study on two populations of O. niloticus. Taiwan A and B across two generations and heritability study on the Taiwan A population. The primary focus of the population genetic study was to determine the relationships among eight populations which was used for breeding and to determine the viability of these populations. Microsatellite markers were used to determine the relationships among tilapia populations. The technique was optimized by varying parameters including the amount of template DNA, different thermal cyclers and others. The results showed that microsatellite markers are good markers for tilapia genetic studies. For the population genetic study, the mean allele number and the mean heterozygosity level for the 40 loci were 43 and 0.5420 respectively. The FST value of 0.2401 among the populations suggested a moderate amount of genetic differentiation among the tilapia populations studied. Low heterozygosity levels suggest low effective population sizes, which may result in high levels ·of inbreeding. Most loci showed a deficiency in heterozygosity, which may be a sign of inbreeding. Most sampled populations showed significant deviations from Hardy-Weinberg equilibrium, which could result from mutation, migration or selection. This outcome could also be due to the small sample sizes examined and the high number of alleles present at individual loci. Based on 11 loci, different genetic distance measures were applied to test the difference in the topology of the five populations examined. The topology and correlation values varied using two models, IAM and SMM. The genetic distances used in this study was selected to resolve relationships among the sampled populations as a

Assessing Host-Pathogen Interaction Networks via RNA-Seq Profiling: A Systems Biology Approach

RNA sequencing is a valuable tool brought about by advances in next generation sequencing (NGS) technology. Initially used for transcriptome mapping, it has grown to become one of the ‘gold standards’ for studying molecular changes that occur in niche environments or within and across infections. It employs high-throughput sequencing with many advantages over previous methods. In this chapter, we review the experimental approaches of RNA sequencing from isolating samples all the way to data analysis methods. We focus on a number of NGS platforms that offer RNA sequencing with each having their own strengths and drawbacks. The focus will also be on how RNA sequencing has led to developments in the field of host-pathogen interactions using the dual RNA sequencing technique. Besides dual RNA sequencing, this review also explores the application of other RNA sequencing techniques such as single cell RNA sequencing as well as the potential use of newer techniques like ‘spatialomics’ and ribosome-profiling in host-pathogen interaction studies. Finally, we examine the common challenges faced when using RNA sequencing and possible ways to overcome these challenges

Single locus microsatellite development for the Malaysian giant freshwater prawns, Macrobrachium rosenbergii

Prawns of the genus Macrobrachium Bate, 1868 (Crustacea: Palaemonidae) are a highly diverse group of decopod crustaceans found in circumtropical marine-, estuarine- and freshwaters. At present, wild stocks have been used to cultivate commercial cultures in many areas in Malaysia. Such repeated harvesting from wild stocks, however, will eventually lead to extinction and loss of genetic diversity. Therefore, it is important to know the genetic make up of these wild stocks in order to manage the populations for conservation and breeding purposes. In the present study, a novel method known as Random Amplified Microsatellites (RAMS) was used to develop single locus microsatellite markers. A total of 13 microsatellite regions were detected. The findings of this study showed a successful detection of the microsatellite sequences in M. rosenbergii using RAMS. This study will be continued to identify more clones that contain microsatellite sequences and to design the forward and reverse primers for each microsatelite region. M. rosenbergii sequences were homologous with Danio rerio (Zebra fish) and Perna viridis (green lipped mussel)

Identification of short-length oligonucleotides biomarker for canine species detection using mitochondrial cytochrome b gene

Introduction: Stray dogs are still available in certain countries without any offered price and made it as a potential source for adulteration with costly meats for more benefit. Furthermore, human forensic evidences from crime scenes were often integrated with biomaterial of canine origin. Most of the DNA based assay for canine species detection used longer amplicon size (>150 bp) which are not suitable for highly degraded food or forensic sample analysis. Therefore, in this study for development of short length canine specific biomarker, mitochondrial cytochrome b (cytb) gene was targeted using simple PCR assay. Objective: Detection of canine species using short length DNA biomarker targeting cytb gene. Methods: The assay targeted a 100-bp fragment of cytochrome b gene using a pair of canine specific primers. The primers specificity were tested under Insilco, as well as in real PCR assay using dog and eight other species DNAs. The consensus 100 bp canine specific site along with cytb sequences of 14 species including dog and human were used for analysis of pair wise distances, construct dendogram and primers mismatch calculation. The stability of the biomarker was tested under commonly used cooking condition and extensive autoclaving state which was known for degradation of target DNA. The sensitivity of the assay was tested using binary admixture composed of dog and most consumed chicken DNA pool. Results & Discussion: The biomarker was 100% canine specific and successfully amplified 100 bp region of canine cytb gene specific target. It was highly stable and sensitive enough to detect as low as 0.1% (0.02 ng) of canine specific target from admixed DNAs. Conclusion: The primers provided the shortest DNA biomarker for canine species detection. The shortest amplicon length, high stability and sensitivity offered its potentiality for canine biomaterials determination from food as well as from degraded samples

Review: Potential of Using Seaweed Silage Inoculated with Lactic Acid Bacteria and Its Impact Towards Sustainable Animal Feed

Since ancient times, seaweed has been used to supplement animal feed in coastal areas. In recent years, there has been a growing demand for seaweed as an animal feed. Effective preservation methods are required because of annual variations in seaweed availability and biochemical composition. Ensiling could be an effective method to preserve seaweeds for animal feed applications. Using lactic acid bacteria is a substitute biological technique for keeping and restoring the usual physiological state of the animal and increasing efficiency. The aim of this review is the potential of using lactic acid bacteria as an inoculant for seaweed silage for the development of sustainable animal feed. According to the microbiological point of view, the results in this area are weak, and limited information is available. Due to its high nutritional content, seaweed silage is a promising animal feed ingredient and is getting acceptance as an alternative animal feed. Therefore, seaweeds contain valuable metabolites such as polyunsaturated fatty acids, carotenoids, phlorotannins, carrageenan, alginate pigments, agar, and minerals (manganese, iodine, calcium, iron, selenium, sodium, zinc) are used as a natural antibiotic source in animal feed. In conclusion, increasing the use of effective lactic acid bacteria as an inoculant in animal feed can make the livestock sector more productive, safer, and friendly to humans and the environment, contributing to animal feed\u27s long-term development

A checklist of the bats of Peninsular Malaysia and progress towards a DNA barcode reference library

Several published checklists of bat species have covered Peninsular Malaysia as part of a broader region and/or in combination with other mammal groups. Other researchers have produced comprehensive checklists for specific localities within the peninsula. To our knowledge, a comprehensive checklist of bats specifically for the entire geopolitical region of Peninsular Malaysia has never been published, yet knowing which species are present in Peninsular Malaysia and their distributions across the region are crucial in developing suitable conservation plans. Our literature search revealed that 110 bat species have been documented in Peninsular Malaysia; 105 species have precise locality records while five species lack recent and/or precise locality records. We retrieved 18 species from records dated before the year 2000 and seven species have only ever been recorded once. Our search of Barcode of Life Datasystems (BOLD) found that 86 (of the 110) species have public records of which 48 species have public DNA barcodes available from bats sampled in Peninsular Malaysia. Based on Neighbour-Joining tree analyses and the allocation of DNA barcodes to Barcode Index Number system (BINs) by BOLD, several DNA barcodes recorded under the same species name are likely to represent distinct taxa. We discuss these cases in detail and highlight the importance of further surveys to determine the occurences and resolve the taxonomy of particular bat species in Peninsular Malaysia, with implications for conservation priorities

Morphometric analysis of craniodental characters of the House Rat, Rattus rattus (Rodentia: Muridae) in Peninsular Malaysia

Rattus rattus which is believed to be native to India is easily found in all continents of the world. With the absence of R. rattus from lineage I and the presence of lineage IV in Southeast Asia, little is known about how rodent morphology varies biogeographically. We evaluate the skull morphometrics in the population of R. rattus in Peninsular Malaysia and examine the intraspecific variation of the skull by assessing 20 craniodental characters of 130 R. rattus specimens. After observing the correlation matrix, highly correlated characters were removed and the remaining 14 craniodental characters were analysed using the conventional morphometrics method via univariate and multivariate statistics. We assessed the impact of age, sex and geographical factors (latitude, longitude and precipitation) on craniodental size. Male and female specimens were analysed separately since statistically significant sexual dimorphism was present. Males showed greater craniodental measurements compared to females. Three distinct age groups (C2, C3 and C4) were visible in the principal component analysis (PCA) and canonical variate analysis (CVA) plots for male and female specimens. Ages C2 and C3 shared similarities in craniodental measurements while C4 showed variability compared to C2 and C3. None of the geographical factors studied showed statistical significance for both male and female R. rattus. The results obtained from this study could potentially be a yardstick to observe if similar variations in craniodental traits are present in R. rattus from other countries in the Southeast Asian region

Isolation and characterization of microsatellite loci in the Malaysian giant freshwater prawn, Macrobrachium rosenbergii

Eight single locus microsatellite markers were developed to characterize the Malaysian giant freshwater prawn, Macrobrachium rosenbergii. These microsatellites were isolated from an enriched genomic library contained by using a 5′‐anchored polymerase chain reaction technique. Primers were designed to flank the repeat sequences and subsequently used to characterize 30 unrelated individuals of the giant freshwater prawn. The polymerase chain reaction amplification products of these eight microsatellite loci were polymorphic with the number of alleles ranging from two to 10 alleles per locus while the levels of heterozygosity ranged from 0.6333 to 0.8667

Genetic diversity analysis in Malaysian giant prawns using expressed sequence tag microsatellite markers for stock improvement program

The Malaysian giant prawn is among the most commonly cultured species of the genus Macrobrachium. Stocks of giant prawns from four rivers in Peninsular Malaysia have been used for aquaculture over the past 25 years, which has led to repeated harvesting, restocking, and transplantation between rivers. Consequently, a stock improvement program is now important to avoid the depletion of wild stocks and the loss of genetic diversity. However, the success of such an improvement program depends on our knowledge of the genetic variation of these base populations. The aim of the current study was to estimate genetic variation and differentiation of these riverine sources using novel expressed sequence tag-microsatellite (EST-SSR) markers, which not only are informative on genetic diversity but also provide information on immune and metabolic traits. Our findings indicated that the tested stocks have inbreeding depression due to a significant deficiency in heterozygotes, and FIS was estimated as 0.15538 to 0.31938. An F-statistics analysis suggested that the stocks are composed of one large panmictic population. Among the four locations, stocks from Johor, in the southern region of the peninsular, showed higher allelic and genetic diversity than the other stocks. To overcome inbreeding problems, the Johor population could be used as a base population in a stock improvement program by crossing to the other populations. The study demonstrated that EST-SSR markers can be incorporated in future marker assisted breeding to aid the proper management of the stocks by breeders and stakeholders in Malaysia