rna sequencing

rna sequencing

Transcriptome Analysis Takes Predictive Breeding a Step Ahead
With the advent of genetic engineering, there has been a drastic change in the agricultural techniques. Gone are those days when man used to depend solely on nature for rainfall, soil fertility etc. Modern-man can toy with the genetic sequence and identify the part of genome which is beneficial for him.

An attempt to improve the crop varieties began almost simultaneously with the agricultural practices. Around ten thousand years ago, man started to alter the genetic make up of crops without his knowledge. The conventional plant breeding practices began when early farmers selected the plant with desirable traits and saved its seeds for the next generation. Later with the knowledge of genetics, plant breeders were able to select the crops with desirable traits like faster growth, pest and disease resistance, sweeter fruits, larger seeds etc. Two plants having two different desirable characters were cross-pollinated artificially to obtain an offspring containing a mixture of these characters (hybrid variety). As a net result of these techniques new varieties of crops have developed which are significantly different from their wild relatives. Conventional breeding techniques largely depend on selection of superior varieties of crops and incorporation of desirable traits into future generations.

Genetic engineering has brought quantum change from conventional breeding practices and has introduced combination of genes which do not exist naturally. Sequencing of genomes of various plants has now become feasible with the faster progression in genetic engineering. Predictive breeding is an advanced technique which comes up with a promise of identifying the beneficial genes out of the entire genome. Genetic markers like SNPs (single nucleotide polymorphism) or minisatellites are used to construct linkage maps. The linkage maps help to identify the traits which are inherited together. The knowledge of genome sequence along with the linkage maps helps to associate the beneficial traits of the plant with specific parts of genome sequence.

Complicated genomes of polyploid crops pose a serious problem to predictive breeding. A cell of a polyploidy organism has more than two sets of homologous chromosomes and each set contains all the genes responsible for the biological traits of that organism. Professor Ian Bancroft took this as a challenge and identified the sequence of rapeseed (Brassica napus). This crop has been found to originate from two related species that is Brassica rapa and Brassica oleracea. Rapeseed (Brassica napus) was declared as a third leading source of vegetable oil in the world in the year 2000. Canola is the name given to Canadian oil that is extracted from certain varieties of Rapeseed. This oil is a rich source of essential fatty acids (omega-6 and omega-3 fatty acids in the ratio of 2:1). Apart from these it is also an important source of Biodiesel which can power motor vehicles. This oil is generally combined with fossil-fuel diesel in ratios varying from 2% to 20% of biodiesel. This does not damage the engine. Isn't it an eco-friendly plant?

Sequencing the DNA of this plant directly was difficult because of its complicated genome. Hence they adopted a different strategy. Transcriptomes were sequenced from young juvenile leaves. Transcriptome is a set of all RNA molecules, including mRNA, rRNA, tRNA, and other non-coding RNA. It represents a small percentage of the genome. The sequence of a transcriptome mirrors the sequence of the DNA from which it was transcribed. This study gives an idea about all the genes which get expressed into proteins in that tissue. Sequence of transcriptome will act like a raw material for scientists struggling to locate the human friendly genes within the entire genome of the polyploid plants. Based on this improved varieties of seeds can be produced.

A large number of crops like potato, wheat, oat, sugarcane, cotton etc are polyploid and few crops amongst them are staple crops in certain parts of the world. Transcriptome sequencing will help the scientists to identify the desirable genes even in the absence of complete genome sequence. This new technique will make predictive breeding of crops with complicated genomes feasible.

Article Source: rna sequencing

single molecule dna sequencing

single molecule dna sequencing

The polymerase chain reaction (PCR) is a process for amplifying very tiny amounts of DNA such that they can be visualized and assessed or used in further scientific processes. PCR is widely used in almost all branches of biology including molecular biology, microbiology, genetics, environmental science, food science, biotechnology, forensic science, and clinical diagnostics. The PCR technique involves using an enzyme called DNA polymerase to amplify (duplicate many times) a piece of DNA. The original molecule of DNA is duplicated by the DNA polymerase enzyme, thus doubling the number of DNA molecules.

Then each of these molecules is duplicated in a second cycle of replication, resulting in four molecules. Again, each of these molecules is duplicated by the enzyme in a third cycle of replication. The process is referred to as a chain reaction - hence the name "polymerase chain reaction". The original single molecule piece of DNA is amplified over many cycles, generating millions of copies of the original DNA molecule. Often the PCR experiment is performed simply to verify the presence or absence of a particular type of DNA but sometimes PCR is used to generate sufficient DNA to be used in a subsequent experiment, for example cloning or DNA sequencing. The original PCR process has been extensively modified to perform a wide array of genetic manipulations, diagnostic tests, and for many other uses. (Reference: Real-Time PCR: Current Technology and Applications ISBN: 978-1-904455-39-4)

Modern variations of PCR include Real-time PCR. This technology allows the newly generated DNA molecules to be analysed immediately they are produced. This ability to monitor the PCR process in real time, while the reaction is still occuring, is of enormous benefit to scientists as it reduces the amount of time required to generate a result (especially important in clinical diagnostics) and also enables the quantitation of the DNA. Real-time PCR can not only answer the question "what DNA is present?" but also "how much DNA is present?".

The polymerase chain reaction is used by a huge number of scientists in a vast range of scientific disciplines. In microbiology and molecular biology laboratories, for example, PCR is used in DNA cloning experiments, genetic manipulation, DNA sequencing, genetic modification, and many other procedures. In clinical diagnostic laboratories PCR is used for the diagnosis of microbial infections, the detection of hereditary diseases and in epidemiological studies. In medicine PCR has led to significant improvements in the diagnosis of infectious disease. PCR is also used for genetic fingerprinting in forensic and paternity testing laboratories. (Reference: Real-Time PCR in Microbiology: From Diagnosis to Characterization ISBN: 978-1-904455-18-9)

Article Source: single molecule dna sequencing