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Related Concept Videos

Characteristics of Practical Op Amps01:16

Characteristics of Practical Op Amps

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A difference amplifier, a crucial component in numerous electronic devices, ideally amplifies only the difference-mode signal, which is the difference between two input signals. However, in practical circuits, the output voltage depends on both the differential gain and the common-mode gain.
The ratio of differential gain to the common-mode gain is defined as the common-mode rejection ratio (CMRR). This ratio quantifies the ability of operational amplifiers (op-amps) to reject common-mode...
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Theoretical Foundations of Nursing Practice

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Theories play an essential role in organizing patient care. Theories refer to a proposed or followed belief, policy, or procedure that is the basis for action. Nursing theories are knowledge-based concepts that guide nurses' actions, influence nursing education and practice, and allow nurses to care for their patients.
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Equivalent Circuits for Practical Transformers01:28

Equivalent Circuits for Practical Transformers

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The practical equivalent circuits of single-phase two-winding transformers exhibit significant deviations from their idealized versions due to the inherent properties of winding resistance and finite core permeability. These properties result in real and reactive power losses, affecting the transformer's performance. Understanding these deviations is crucial for designing more efficient transformers.
In a practical transformer, each winding exhibits resistance and leakage reactance. The...
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Preparation of Amides01:29

Preparation of Amides

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Amides are synthesized by treating carboxylic acids with amines in the presence of dehydrating agents like dicyclohexylcarbodiimide (DCC).
The DCC-promoted synthesis of amides begins with the protonation of DCC by carboxylic acid. The protonation makes it a better acceptor. Next, the addition of carboxylate to the protonated carbodiimide gives a reactive acylating agent.
Subsequently, the amine acts as a nucleophile that attacks the acylating agent to form a tetrahedral intermediate. In the...
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Preparation of Nitriles01:12

Preparation of Nitriles

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One of the common methods to prepare nitriles is the dehydration of amides. This method requires strong dehydrating agents like phosphorous pentoxide or boiling acetic anhydride for converting amides to nitriles. Another reagent namely, thionyl chloride also accomplishes the dehydration of amides, where amide acts as a nucleophile. The first step of the mechanism involves the nucleophilic attack by the amide on the thionyl chloride to form an intermediate. In the next step, the electron pairs...
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Preparation of Epoxides03:00

Preparation of Epoxides

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Overview
Epoxides result from alkene oxidation, which can be achieved by a) air, b) peroxy acids, c) hypochlorous acids, and d) halohydrin cyclization.
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Epoxidation of alkenes via oxidation with peroxy acids involves the conversion of a carbon–carbon double bond to an epoxide using the oxidizing agent meta-chloroperoxybenzoic acid, commonly known as MCPBA. Since the O–O bond of peroxy acids is very weak, the addition of electrophilic oxygen of peroxy acids to...
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Related Experiment Video

Updated: Jan 23, 2026

Single Read and Paired End mRNA-Seq Illumina Libraries from 10 Nanograms Total RNA
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Best Practices for Illumina Library Preparation.

Iraad F Bronner1, Michael A Quail1

  • 1Wellcome Trust Sanger Institute, Wellcome Trust Genome Campus, Hinxton, Cambridgeshire, United Kingdom.

Current Protocols in Human Genetics
|June 20, 2019
PubMed
Summary

This unit details best practices for Illumina library preparation, covering quantitation, fragmentation, cleanup, and indexing. Following these protocols enhances cost-effectiveness and library yields for both high-throughput and low-throughput applications.

Keywords:
IlluminaPCRbiasindexinglibrarynext-generation sequencingsample preparation

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Last Updated: Jan 23, 2026

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Area of Science:

  • Molecular Biology
  • Genomics
  • Biotechnology

Background:

  • Illumina sequencing is a cornerstone of modern genomics.
  • Optimizing library preparation is crucial for accurate and efficient sequencing outcomes.
  • Previous protocols require updates to incorporate current best practices and technologies.

Purpose of the Study:

  • To provide comprehensive protocols and recommendations for Illumina library preparation.
  • To guide researchers in optimizing workflows for reduced bias and increased cost-effectiveness.
  • To serve as an extensive update to existing library preparation guidelines.

Main Methods:

  • Review of template quantitation and fragmentation methods.
  • Detailed protocols for solid-phase reverse-immobilization cleanup, including buffer exchange and size selection.
  • Guidelines for end repair, A-tailing, adapter ligation, and indexing strategies.
  • Considerations for polymerase chain reaction (PCR) use and normalization/pooling strategies.
  • Final library quantification methodologies.

Main Results:

  • Established best practices for various stages of Illumina library preparation.
  • Workflows applicable to both high-throughput and low-throughput sequencing.
  • Demonstrated potential for reduced bias and increased cost-effectiveness.
  • Aimed at achieving high library yields.

Conclusions:

  • The described protocols offer a robust framework for Illumina library preparation.
  • Adherence to these recommendations can significantly improve sequencing data quality and experimental efficiency.
  • This updated unit serves as a vital resource for researchers utilizing Illumina sequencing technology.