Short Read Sequencing
Short Read Sequencing - Related Videos
08:57
Amplicon Sequencing using the Long-Read Sequencing Technologies
0 Views •
2025
This protocol was optimized for targeted deep sequencing of 18 drug-resistance regions in Mycobacterium tuberculosis using a long-read sequencing platform, followed by analysis with a tuberculosis-specific bioinformatics pipeline designed for long-read data.
12:08
Hybrid De Novo Genome Assembly for the Generation of Complete Genomes of Urinary Bacteria using Short- and Long-read Sequencing Technologies
0 Views •
Cited by 9 •
2021
This protocol details a comprehensive approach for the culturing, sequencing, and de novo hybrid genome assembly of urinary bacteria. It provides a reproducible procedure for the generation of complete, circular genome sequences useful in studying both chromosomal and extrachromosomal genetic elements contributing to urinary colonization, pathogenesis, and antimicrobial resistance dissemination.
06:56
Purification of High Molecular Weight Genomic DNA from Powdery Mildew for Long-Read Sequencing
0 Views •
Cited by 17 •
2017
Described here is a method for the extraction, purification, and quality control of genomic DNA from the obligate biotrophic fungal pathogen, powdery mildew, for use in long-read genome sequencing.
10:34
Ultra-long Read Sequencing for Whole Genomic DNA Analysis
0 Views •
Cited by 28 •
2019
Long-read sequences greatly facilitate the assembly of complex genomes and characterization of structural variation. We describe a method to generate ultra-long sequences by nanopore-based sequencing platforms. The approach adopts an optimized DNA extraction followed by modified library preparations to generate hundreds of kilobase reads with moderate coverage from human cells.
08:48
Stretching Short Sequences of DNA with Constant Force Axial Optical Tweezers
0 Views •
Cited by 5 •
2011
We illustrate the use of a constant force axial optical tweezers to explore the mechanical properties of short DNA molecules. By stretching DNA axially, we minimize steric hindrances and artifacts arising in conventional lateral manipulation, allowing us to study DNA molecules as short as ~100 nm.
FAQs
Each cycle records nucleotide incorporation through signal detection across library molecules. Repeating the cycles generates read sequences, which computational tools then analyze. This repeated measurement allows very large numbers of reads to be produced from one experiment, while the resulting sequence information provides the input for alignment to a reference genome or for de novo assembly.
Fragmentation determines the pieces of DNA or RNA entering the workflow, while adapter ligation and amplification convert those pieces into library molecules for sequencing. These preparation steps shape the material represented in the final reads. As a result, library construction is not merely preliminary handling; it establishes the sequence material available for downstream genetic analysis.
Alignment places reads against an existing reference genome, whereas de novo assembly reconstructs sequence from the reads without relying on that reference. The two routes support different analyses: alignment facilitates comparison with known genomic sequence, while assembly helps reconstruct sequence directly. Repetitive regions can complicate either approach and make interpretation less certain.
Short read length limits how much surrounding sequence each read can represent. Reads originating from repetitive regions may therefore be difficult to place, while complex structural changes can be difficult to reconstruct from fragmented sequence information. Consequently, high accuracy and extensive read production do not ensure complete resolution of every genomic feature.
A typical workflow begins with DNA or RNA fragmentation, followed by adapter ligation and amplification to produce library molecules. The library then undergoes repeated nucleotide-incorporation and signal-detection cycles. Finally, computational analysis aligns reads to a reference genome or assembles them de novo. This sequence connects laboratory preparation, signal generation, and genetic interpretation.
Short Read Sequencing can identify genetic variants, quantify gene expression, characterize microbial and human genomes, and investigate disease-associated mutations. These uses make the method relevant to both genome-focused and transcript-focused studies. Its high-throughput nature also supports large studies in which many sequence measurements must be generated and analyzed consistently.