Screening for the Presence of a Transgene in Genetically Altered Mice
Location
CSU Ballroom
Start Date
21-4-2008 1:00 PM
End Date
21-4-2008 3:00 PM
Student's Major
Biological Sciences
Student's College
Science, Engineering and Technology
Mentor's Name
Marilyn C. Hart
Mentor's Department
Biological Sciences
Mentor's College
Science, Engineering and Technology
Description
In previous studies, genetically altered mice were produced which contained a transgene, providing additional copies of the beta 1 isoform of actin capping protein under the control of the myosin heavy chain promoter. To ensure the integrity of members of the established mouse colony, transgenic and wild type genomic DNA were isolated by digestion of a tail clip. Protease K digestion lysed the cells, liberating the nucleic acid. The DNA was purified by phenol/chloroform.extraction and precipitated using 100% ethanol. After centrifugation, the DNA pellet was washed with 70% ethanol to remove salts. We are using Polymerase Chain Reaction (PCR) to amplify the transgene. We have optimized the buffer, annealing temperature, and other PCR variables. We have amplified the genomic DNA from both wild type and transgenic mice. The transgenic product is approximately 1-kilobase (kb). An internal actin control generated a PCR product of 450 base pairs. Agarose gel electrophoresis confirmed the identification of the anticipated products.
Screening for the Presence of a Transgene in Genetically Altered Mice
CSU Ballroom
In previous studies, genetically altered mice were produced which contained a transgene, providing additional copies of the beta 1 isoform of actin capping protein under the control of the myosin heavy chain promoter. To ensure the integrity of members of the established mouse colony, transgenic and wild type genomic DNA were isolated by digestion of a tail clip. Protease K digestion lysed the cells, liberating the nucleic acid. The DNA was purified by phenol/chloroform.extraction and precipitated using 100% ethanol. After centrifugation, the DNA pellet was washed with 70% ethanol to remove salts. We are using Polymerase Chain Reaction (PCR) to amplify the transgene. We have optimized the buffer, annealing temperature, and other PCR variables. We have amplified the genomic DNA from both wild type and transgenic mice. The transgenic product is approximately 1-kilobase (kb). An internal actin control generated a PCR product of 450 base pairs. Agarose gel electrophoresis confirmed the identification of the anticipated products.
Recommended Citation
Huang, Der-How and Brad Skrukrud. "Screening for the Presence of a Transgene in Genetically Altered Mice." Undergraduate Research Symposium, Mankato, MN, April 21, 2008.
https://cornerstone.lib.mnsu.edu/urs/2008/poster-session-B/8
