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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 flow of genetic information in cells from DNA to mRNA to protein is described by the central dogma, which states that genes specify the sequence of mRNAs, which in turn specify the sequence of amino acids making up all proteins. The decoding of one molecule to another is performed by specific proteins and RNAs. Because the information stored in DNA is so central to cellular function, it makes intuitive sense that the cell would make mRNA copies of this information for protein synthesis...
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DNA Sequence Recognition by DNA Primase Using High-Throughput Primase Profiling
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Protocols for Investigating the Leaf Mycobiome Using High-Throughput DNA Sequencing.

Shawn P Brown1, Devin R Leopold2, Posy E Busby3

  • 1Department of Biological Sciences, University of Memphis, Memphis, TN, USA.

Methods in Molecular Biology (Clifton, N.J.)
|September 6, 2018
PubMed
Summary

DNA metabarcoding is crucial for studying plant-associated fungi. This protocol details best practices for DNA metabarcoding the leaf mycobiome to minimize errors and ensure reliable fungal community data.

Keywords:
AmpliconsFungal leaf endophytesMetabarcodingMock communityMycobiomeNext-generation sequencingPhyllospherePhytobiomePrimer biasSample bias

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Area of Science:

  • Mycology
  • Molecular Biology
  • Ecology

Background:

  • DNA metabarcoding is a powerful technique for analyzing fungal communities associated with plants.
  • Despite its utility, potential systematic errors can bias metabarcoding data, complicating the study of plant-fungal interactions.
  • Standardized protocols are needed to ensure the accuracy of leaf mycobiome investigations.

Purpose of the Study:

  • To present a standardized protocol for DNA metabarcoding of the leaf mycobiome.
  • To minimize systematic errors and biases in fungal community profiling.
  • To provide a foundation for reliable elucidation of plant-fungal interactions.

Main Methods:

  • Utilizes high-throughput sequencing of taxon-specific loci.
  • Employs careful laboratory practices to mitigate errors.
  • Incorporates validation steps to ensure data integrity.

Main Results:

  • The presented protocol aims to reduce common sources of systematic error in metabarcoding.
  • Implementation of best practices enhances the reliability of leaf mycobiome composition data.
  • The protocol facilitates more accurate studies of plant-fungal interactions.

Conclusions:

  • This protocol offers a robust approach to DNA metabarcoding of the leaf mycobiome.
  • Adherence to best practices is essential for generating high-quality fungal community data.
  • The method supports advancements in understanding plant-fungal ecological relationships.