---
_id: '12720'
abstract:
- lang: eng
  text: Here we describe the in vivo DNA assembly approach, where molecular cloning
    procedures are performed using an E. coli recA-independent recombination pathway,
    which assembles linear fragments of DNA with short homologous termini. This pathway
    is present in all standard laboratory E. coli strains and, by bypassing the need
    for in vitro DNA assembly, allows simplified molecular cloning to be performed
    without the plasmid instability issues associated with specialized recombination-cloning
    bacterial strains. The methodology requires specific primer design and can perform
    all standard plasmid modifications (insertions, deletions, mutagenesis, and sub-cloning)
    in a rapid, simple, and cost-efficient manner, as it does not require commercial
    kits or specialized bacterial strains. Additionally, this approach can be used
    to perform complex procedures such as multiple modifications to a plasmid, as
    up to 6 linear fragments can be assembled in vivo by this recombination pathway.
    Procedures generally require less than 3 h, involving PCR amplification, DpnI
    digestion of template DNA, and transformation, upon which circular plasmids are
    assembled. In this chapter we describe the requirements, procedure, and potential
    pitfalls when using this technique, as well as protocol variations to overcome
    the most common issues.
alternative_title:
- Methods in Molecular Biology
article_processing_charge: No
author:
- first_name: Sandra
  full_name: Arroyo-Urea, Sandra
  last_name: Arroyo-Urea
- first_name: Jake
  full_name: Watson, Jake
  id: 63836096-4690-11EA-BD4E-32803DDC885E
  last_name: Watson
  orcid: 0000-0002-8698-3823
- first_name: Javier
  full_name: García-Nafría, Javier
  last_name: García-Nafría
citation:
  ama: 'Arroyo-Urea S, Watson J, García-Nafría J. Molecular Cloning Using In Vivo
    DNA Assembly. In: Scarlett G, ed. <i>DNA Manipulation and Analysis</i>. Vol 2633.
    MIMB. New York, NY, United States: Springer Nature; 2023:33-44. doi:<a href="https://doi.org/10.1007/978-1-0716-3004-4_3">10.1007/978-1-0716-3004-4_3</a>'
  apa: 'Arroyo-Urea, S., Watson, J., &#38; García-Nafría, J. (2023). Molecular Cloning
    Using In Vivo DNA Assembly. In G. Scarlett (Ed.), <i>DNA Manipulation and Analysis</i>
    (Vol. 2633, pp. 33–44). New York, NY, United States: Springer Nature. <a href="https://doi.org/10.1007/978-1-0716-3004-4_3">https://doi.org/10.1007/978-1-0716-3004-4_3</a>'
  chicago: 'Arroyo-Urea, Sandra, Jake Watson, and Javier García-Nafría. “Molecular
    Cloning Using In Vivo DNA Assembly.” In <i>DNA Manipulation and Analysis</i>,
    edited by Garry Scarlett, 2633:33–44. MIMB. New York, NY, United States: Springer
    Nature, 2023. <a href="https://doi.org/10.1007/978-1-0716-3004-4_3">https://doi.org/10.1007/978-1-0716-3004-4_3</a>.'
  ieee: 'S. Arroyo-Urea, J. Watson, and J. García-Nafría, “Molecular Cloning Using
    In Vivo DNA Assembly,” in <i>DNA Manipulation and Analysis</i>, vol. 2633, G.
    Scarlett, Ed. New York, NY, United States: Springer Nature, 2023, pp. 33–44.'
  ista: 'Arroyo-Urea S, Watson J, García-Nafría J. 2023.Molecular Cloning Using In
    Vivo DNA Assembly. In: DNA Manipulation and Analysis. Methods in Molecular Biology,
    vol. 2633, 33–44.'
  mla: Arroyo-Urea, Sandra, et al. “Molecular Cloning Using In Vivo DNA Assembly.”
    <i>DNA Manipulation and Analysis</i>, edited by Garry Scarlett, vol. 2633, Springer
    Nature, 2023, pp. 33–44, doi:<a href="https://doi.org/10.1007/978-1-0716-3004-4_3">10.1007/978-1-0716-3004-4_3</a>.
  short: S. Arroyo-Urea, J. Watson, J. García-Nafría, in:, G. Scarlett (Ed.), DNA
    Manipulation and Analysis, Springer Nature, New York, NY, United States, 2023,
    pp. 33–44.
date_created: 2023-03-12T23:01:02Z
date_published: 2023-03-01T00:00:00Z
date_updated: 2023-03-16T08:34:24Z
day: '01'
department:
- _id: PeJo
doi: 10.1007/978-1-0716-3004-4_3
editor:
- first_name: Garry
  full_name: Scarlett, Garry
  last_name: Scarlett
external_id:
  pmid:
  - '36853454'
intvolume: '      2633'
language:
- iso: eng
month: '03'
oa_version: None
page: 33-44
place: New York, NY, United States
pmid: 1
publication: DNA Manipulation and Analysis
publication_identifier:
  eisbn:
  - 978-1-0716-3004-4
  eissn:
  - 1940-6029
  isbn:
  - 978-1-0716-3003-7
  issn:
  - 1064-3745
publication_status: published
publisher: Springer Nature
quality_controlled: '1'
scopus_import: '1'
series_title: MIMB
status: public
title: Molecular Cloning Using In Vivo DNA Assembly
type: book_chapter
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 2633
year: '2023'
...