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Genomics is the science of genomes: it is the study of all the genetic material of an organism. In humans, the genome consists of information carried in 23 pairs of chromosomes in the nucleus, as well as mitochondrial DNA. In genomics, both coding and non-coding DNA is sequenced and analyzed. Genomics allows a better understanding of all living things, their evolution, and their diversity. It has a myriad of uses: for example, to build phylogenetic trees, to improve productivity and...
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Diploid organisms inherit genetic material through chromosomes from both parents. Copies of the same gene are known as alleles. In most cases, both alleles are simultaneously expressed and allow various cellular processes to function optimally. If one of the alleles is missing or mutated, the expression of the other allele can compensate; however, this is not true for all genes.
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While every living organism has a genome of some kind (be it RNA, or DNA), there is considerable variation in the sizes of these blueprints. One major factor that impacts genome size is whether the organism is prokaryotic or eukaryotic. In prokaryotes, the genome contains little to no non-coding sequence, such that genes are tightly clustered in groups or operons sequentially along the chromosome. Conversely, the genes in eukaryotes are punctuated by long stretches of non-coding sequence.
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Cis-regulatory sequences are short fragments of non-coding DNA that are present on the same chromosomes as the genes that they regulate. These fragments serve as binding sites for transcriptional regulators, proteins that are responsible for controlling gene transcription and differential gene expression across cell types in eukaryotes. Cis-regulatory sequences can be close to the gene of interest or thousands of bases away in the DNA sequence; however, those sequences that are further away are...
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The present-day mitochondrial and chloroplast genomes have retained some of the characteristics of their ancestral prokaryotes and also have acquired new attributes during their evolution within eukaryotic cells. Like prokaryotic genomes, mitochondrial and chloroplast genomes neither bind with histone-like proteins nor show complex packaging into chromosome-like structures, as observed in eukaryotes. Unlike mitotic cell divisions observed in eukaryotic cells, mitochondria and chloroplasts...
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Sequences are fundamental mathematical objects consisting of ordered lists of numbers that follow a specific rule or pattern. Sequences are critical in various mathematical concepts, including calculus, series, and number theory. They can model real-world phenomena such as population growth, financial investments, and physical processes like the diminishing height of a bouncing ball.Each number in a sequence is referred to as a term. Typically, the terms are denoted as a1, a2, a3,…, where...
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Ultra-long Read Sequencing for Whole Genomic DNA Analysis
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Genome Sequencing.

Yuko Yoshinaga1, Christopher Daum1, Guifen He1

  • 1United States Department of Energy Joint Genome Institute, Walnut Creek, CA, USA.

Methods in Molecular Biology (Clifton, N.J.)
|June 8, 2018
PubMed
Summary

Choosing between Illumina short-read and PacBio long-read sequencing for fungal genomes depends on DNA quality, cost, and genome complexity. Both next-generation sequencing platforms offer distinct advantages for genomic research.

Keywords:
HiSeq 2500IlluminaNext-generation sequencing (NGS)Pacific BiosciencesRS IISequencing by synthesis (SBS)Single molecule real-time (SMRT) sequencing

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Area of Science:

  • Genomics
  • Bioinformatics
  • Molecular Biology

Background:

  • Next-generation sequencing (NGS) strategies for fungal genomes are influenced by species-specific genomic characteristics, DNA quality/quantity, and budget constraints.
  • Short-read sequencing by Illumina (Sequencing by Synthesis - SBS) is cost-effective but can struggle with repetitive genomic regions due to limited read length (~300 bp).
  • Long-read sequencing by PacBio (Single Molecule, Real-Time - SMRT) aids de novo assembly of complex genomes with long repeats, but higher cost and lower throughput present challenges.

Purpose of the Study:

  • To compare and contrast Illumina SBS and PacBio SMRT sequencing platforms for fungal genome sequencing.
  • To outline key decision-making factors when selecting an appropriate NGS platform for fungal genomics projects.

Main Methods:

  • Discussion of Illumina SBS technology, highlighting its speed, cost-effectiveness, and suitability for low-quality DNA.
  • Analysis of PacBio SMRT technology, emphasizing its long-read capability for resolving complex genomic structures.
  • Consideration of DNA input requirements for both platforms, with SMRT demanding high-quality, long DNA fragments.

Main Results:

  • Illumina SBS excels in throughput and cost-efficiency, suitable for well-characterized genomes or when DNA quality is compromised.
  • PacBio SMRT provides superior data for assembling genomes with extensive repetitive elements, despite higher costs.
  • Platform choice necessitates balancing read length, cost, DNA quality, and the specific challenges of the target fungal genome.

Conclusions:

  • The optimal NGS platform for fungal genome sequencing is context-dependent, requiring careful evaluation of project-specific needs.
  • Understanding the trade-offs between short-read and long-read sequencing is crucial for successful fungal genome assembly and analysis.