Related Experiment Video
Updated: Dec 22, 2025

Experimental Design for Laser Microdissection RNA-Seq: Lessons from an Analysis of Maize Leaf Development
Published on: March 5, 2017
Effect of sequence depth and length in long-read assembly of the maize inbred NC358
Shujun Ou1, Jianing Liu2, Kapeel M Chougule3
1Department of Ecology, Evolution, and Organismal Biology, Iowa State University, Ames, Iowa, 50011, USA.
Abstract:
Improvements in long-read data and scaffolding technologies have enabled rapid generation of reference-quality assemblies for complex genomes. Still, an assessment of critical sequence depth and read length is important for allocating limited resources. To this end, we have generated eight assemblies for the complex genome of the maize inbred line NC358 using PacBio datasets ranging from 20 to 75 × genomic depth and with N50 subread lengths of 11-21 kb. Assemblies with ≤30 × depth and N50 subread length of 11 kb are highly fragmented, with even low-copy genic regions showing degradation at 20 × depth. Distinct sequence-quality thresholds are observed for complete assembly of genes, transposable elements, and highly repetitive genomic features such as telomeres, heterochromatic knobs, and centromeres. In addition, we show high-quality optical maps can dramatically improve contiguity in even our most fragmented base assembly. This study provides a useful resource allocation reference to the community as long-read technologies continue to mature.
Related Concept Videos
Maxam-Gilbert Sequencing
Challenges of the Maxam-Gilbert Method
The...
Next-generation Sequencing
Next-Generation Sequencing Methods
Although all next-generation methods use different technologies, they all share a set of standard features....
RNA-seq
Before the discovery of RNA-seq, microarray-based methods and Sanger sequencing were used for transcriptome analysis. However, while...
Genome Annotation and Assembly
Sanger Sequencing

