Cloning of DNA fragments: Ligation reactions in agarose gel

Furtado, Agnelo (2014). Cloning of DNA fragments: Ligation reactions in agarose gel. In Robert J. Henry and Agnelo Furtado (Ed.), Cereal genomics: methods and protocols (pp. 117-121) New York, United States: Humana Press. doi:10.1007/978-1-62703-715-0_10

Author Furtado, Agnelo
Title of chapter Cloning of DNA fragments: Ligation reactions in agarose gel
Title of book Cereal genomics: methods and protocols
Place of Publication New York, United States
Publisher Humana Press
Publication Year 2014
Sub-type Chapter in reference work, encyclopaedia, manual or handbook
DOI 10.1007/978-1-62703-715-0_10
Open Access Status Not Open Access
Series Methods in Molecular Biology
ISBN 9781627037143
ISSN 1064-3745
Editor Robert J. Henry
Agnelo Furtado
Volume number 1099
Chapter number 10
Start page 117
End page 121
Total pages 5
Total chapters 24
Language eng
Subjects 1312 Molecular Biology
1311 Genetics
Abstract/Summary Ligation reactions to ligate a desired DNA fragment into a vector can be challenging to beginners and especially if the amount of the insert is limiting. Although additives known as crowding agents, such as PEG 8000, added to the ligation mixes can increase the success one has with ligation reactions, in practice the amount of insert used in the ligation can determine the success or the failure of the ligation reaction. The method described here, which uses insert DNA in gel slice added directly into the ligation reaction, has two benefits: (a) using agarose as the crowding agent and (b) reducing steps of insert purification. The use of rapid ligation buffer and incubation of the ligation reaction at room temperature greatly increase the efficiency of the ligation reaction even for blunt-ended ligation.
Keyword Agarose
Gel slice
Rapid ligation
Q-Index Code BX
Q-Index Status Confirmed Code
Institutional Status UQ

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Created: Tue, 28 Jan 2014, 10:22:51 EST by System User on behalf of Centre for Nutrition and Food Sciences