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Genome Annotation and Assembly

Genome Annotation and Assembly

The genome refers to all of the genetic material in an organism. It can range from a few million base pairs in microbial cells to several billion base pairs in many eukaryotic organisms. Genome assembly refers to the process of taking the DNA sequencing data and putting it all back together in a correct order to create a close representation of the original genome. This is followed by the identification of functional elements on the newly assembled genome, a process called genome annotation.

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Related Experiment Video

Updated: Oct 9, 2025

Rapid Assembly of Multi-Gene Constructs using Modular Golden Gate Cloning

Rapid Assembly of Multi-Gene Constructs using Modular Golden Gate Cloning

Published on: February 5, 2021

Assembly of Multiple Full-Size Genes or Genomic DNA Fragments on Human Artificial Chromosomes Using the Iterative

Nicholas C O Lee¹, Nikolai S Petrov¹, Vladimir Larionov¹

¹Developmental Therapeutics Branch, National Cancer Institute, National Institutes of Health, Bethesda, Maryland.

Current Protocols

|December 17, 2021

Summary

This study details a new method for inserting multiple genes into human artificial chromosomes (HACs) using an iterative integration system (IIS). This advanced technique enables the creation of complex gene models for studying genetic disorders.

Keywords:

DNA assembly HAC IIS gene delivery human artificial chromosome iterative integration system

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Automated Robotic Liquid Handling Assembly of Modular DNA Devices

Automated Robotic Liquid Handling Assembly of Modular DNA Devices

Published on: December 1, 2017

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Last Updated: Oct 9, 2025

Rapid Assembly of Multi-Gene Constructs using Modular Golden Gate Cloning

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Published on: February 5, 2021

Generation of Plasmid Vectors Expressing FLAG-tagged Proteins Under the Regulation of Human Elongation Factor-1α Promoter Using Gibson Assembly

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Published on: February 9, 2015

Automated Robotic Liquid Handling Assembly of Modular DNA Devices

Automated Robotic Liquid Handling Assembly of Modular DNA Devices

Published on: December 1, 2017

Area of Science:

Genetics
Molecular Biology
Biotechnology

Background:

Human artificial chromosomes (HACs) are valuable gene delivery vectors, offering stable episomal maintenance and large DNA carrying capacity.
The previously developed alphoidtetO-HAC facilitates single-gene insertion into Chinese hamster ovary (CHO) cells for subsequent transfer via microcell-mediated chromosome transfer (MMCT).

Purpose of the Study:

To detail a protocol for loading multiple genomic DNA segments or genes into the alphoidtetO-HAC vector using an iterative integration system (IIS).
To enable serial assembly of genomic loci, large gene fragments, or insertion of multiple genes into a single artificial chromosome.

Main Methods:

Utilizes an iterative integration system (IIS) with Cre, ΦC31, and ΦBT recombinases for sequential DNA segment insertion.
Employs changes in marker fluorescent proteins for simplified screening and selection/counterselection markers for error-proofing.
Includes fluorescence in situ hybridization (FISH) for analysis of the circular IIS-alphoidtetO-HAC.

Main Results:

Successfully demonstrates iterative integration of multiple DNA segments into the HAC vector.
The system allows for error correction through selection and counterselection markers, ensuring accurate DNA incorporation.
Facilitates the elimination of the HAC from cells, enabling comparative phenotype analysis.

Conclusions:

The IIS-alphoidtetO-HAC system provides a robust method for engineering synthetic chromosomes with multiple genes.
This technology is expected to advance the study of complex multi-gene genetic disorders by creating humanized models.
Enables investigation of biomedically relevant pathways and multi-gene regulation.