Genetics Exam 3

Specific sequencing techniques (for memorization purposes).

Sanger Sequencing: Read Length

Medium reads typically ranging from 800 to 1000 base pairs

Sanger Sequencing: Accuracy

The "Gold Standard" for accuracy at approximately 99.99%

Sanger Sequencing: Efficiency

Low throughput; it sequences only one DNA fragment per capillary run

Sanger Sequencing: Primary Use Case

Best for validating single genes

Illumina: Read Length

Short reads usually between 50 and 300 base pairs

Illumina: Accuracy

High accuracy (~99.9%) making it the standard for finding small variants

Illumina: Efficiency

Massive throughput; generates billions of reads in a single run

Illumina: Primary Use Case

Best for Whole Genome Sequencing (WGS)

PacBio SMRT: Read Length

Long reads typically averaging 10kb to 20kb but can exceed 100kb

PacBio SMRT: HiFi Mode

A circular consensus method that provides both long reads and high accuracy (>99%)

PacBio SMRT: Primary Use Case

Best for de novo genome assembly and detecting large structural variants

Oxford Nanopore (ONT): Read Length

Ultra-long reads with the potential to reach over 1 million base pairs (1 Mb+)

Oxford Nanopore (ONT): Portability

Unique for its MinION device which allows for sequencing in the field or remote locations

Oxford Nanopore (ONT): Accuracy

Historically lower raw accuracy than Illumina

Oxford Nanopore (ONT): Primary Use Case

Best for closing gaps in assemblies

SNP Genotyping: Mechanism

Uses a microarray (chip) with fixed probes that hybridize to specific DNA sequences

SNP Genotyping: Discovery Power

Zero; it cannot find new or "private" mutations because it only tests for known variants

SNP Genotyping: Cost/Throughput

Extremely cheap and efficient for screening hundreds of thousands of SNPs across large populations

SNP Genotyping: Primary Use Case

Best for Genome-Wide Association Studies (GWAS) to link known variants to common diseases

PCR: Function in Sequencing

A preparatory step used to amplify or "enrich" a specific DNA region before the actual sequencing run

Sequencing "Read": Definition

The discrete string of nucleotides (A, T, C, G) generated from a single DNA fragment.

Sequencing "Run": Definition

One complete execution of the sequencing process on a machine

"Contig": Definition

A continuous sequence formed by overlapping multiple reads; represents a physical portion of the genome.

Assembly "Scaffold": Definition

A series of contigs oriented in the correct order but separated by gaps of unknown sequence