Microinjection of BAC DNA into the Pronuclei of Fertilized Mouse Oocytes
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Last Update: 2021-02-17
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Source: Internet
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Author: User
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Microinjection of
DNA
into the pronuclei of fertilized oocytes is one of the two most commonly used methods for gene transfer into the mouse genome (
1
,
2
). The first successful attempt to perform this technique was carried out by Lin in 1966 (
3
), who could show that the early fertilized embryo could survive the mechanical damage of inserting a glass needle into the pronucleus. However, it was not until 1981 that small DNA fragments were integrated into the genome (
4
). This technique is well described, and has now become a standard procedure (
5
,
6
). More recently, the use of larger DNA fragments has been established. Yeast artificial chromosome (YAC) (
7
,
8
), P1 artificial chromosome (PAC) (
9
,
10
), and bacterial artificial chromosome (BAC) DNA (
11
,
12
) can all be used for the generation of transgenic mice. A comprehensive review of BAC and YAC transgenesis is given by Giraldo and Montoliu (
13
) extensively comparing these applications. Although the basic technique for microinjection of large constructs are similar to those used for shorter DNA segments, there are some special requirements (
14
). In this chapter, we describe relevant steps for microinjection using large DNA constructs.
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