High-throughput sequencing can help researchers skip library construction, avoiding the bias introduced during the subcloning process. Relying on a powerful bioinformatics analysis capability in the later stages, genome resequencing can be done very easily with a reference genome high-throughput sequencing technology.

In 2007, van Orsouw et al. combined improved AFLP technology and 454. Sequencing technology to re-sequence the maize genome. More than 75% of the SNPs found in this resequencing experiment can be verified by SNPWave technology, providing a polymorphism analysis of complex genomes, especially plant genomes containing highly repetitive sequences.

In 2008, Hillier re-sequenced Solexa on the C. elegans CB4858 line to find deletions or amplifications of SNPs and single-sites in the nematode genome. However, it should also be noted that due to the limitations of high-throughput sequencing read lengths, the use of novo sequencing for unknown genomes is limited, and this work still requires assistance of traditional sequencing (read lengths up to 850 bases). However, this does not affect the application of high-throughput sequencing technology in genome-wide mRNA expression profiling, microRNA expression profiling, ChIP-chip and DNA methylation.

In 2008, Mortazavi et al. performed RNA deep sequencing of mouse brain, liver and skeletal muscles. This work demonstrates two major advances in transcriptome research, expression counting and sequence analysis.

Counting each sequence measured to obtain the expression level of each specific transcript is a digital expression profiling that detects very low abundance transcripts. By analyzing the measured sequences, more than 90% of the data showed to fall in known exons, and those outside the known sequence showed by data analysis is the unreported RNA slice form. The 3' untranslated region, the altered promoter region, and the potential small RNA precursors found that at least 3,500 genes have more than one splicing form. And this information can not be found whether using chip technology or SAGE library sequencing.

Another widely used field of high-throughput sequencing is small-molecule RNA or non-coding RNA (ncRNA) research. Sequencing methods can easily solve the technical problems encountered by chip technology in detecting small molecules (short sequence, highly homologous), and the short sequence of small RNA matches the length of high-throughput sequencing, making the data "not wasted". At the same time, the sequencing method can also find new small RNAs in the experiment. For example, new RNAs have been found in Chlamydomonas, zebrafish, fruit flies, nematodes, humans and chimpanzees. 400,000 sequences were obtained from the nematode, and 18 new small RNA molecules and a new class of small RNAs were discovered through analysis.

In the study of DNA-protein interactions, chromatin immunoprecipitation-depth sequencing (ChIP-seq) experiments have also demonstrated their great potential. The DNA after chromatin immunoprecipitation is directly sequenced. Compared with ref seq, the binding information of protein to DNA can be directly obtained. Compared with ChIP-chip, ChIP-seq can detect smaller binding segments, unknown binding sites, mutations within the binding site and segments with lower protein affinity.

Author's Bio: 

CD Genomics was established in 2004, the company is aiming at providing the research community with high quality Next Generation Sequencing, PacBio SMRT sequencing, and microarray services. Due to the demand for sequencing services has being increased, CD Genomics has already updated its technology platform to mainstream NGS and microarray instruments. At present, the company’s senior bioinformaticians have ever viewed more than ten thousands of trace files and accumulated abundant experience with its Illumina HiSeq 2500, HiSeq 4000, Miseq Benchtop Sequencer, PacBio Sequel, PacBio RS II, Ion Torrent PGM, and ABI 3730/3730XL analyzer, etc. The company continues to work hard to offer researchers the same dependable services to pharmaceutical and biotech companies, as well as academia and government agencies for the purpose of satisfying all sequencing or array needs.