Transgene Excision Protocol
Transgene Excision Protocol - Related Videos
03:37
Porcine Heart Excision: A Protocol to Extract Porcine Heart for Cardiac Transplant
0 Views •
2025
In this video, we describe a step-by-step protocol to extract the heart from a porcine model for cardiac transplantation.
01:54
Base Excision Repair
0 Views •
2020
One of the common DNA damages is the chemical alteration of single bases by alkylation, oxidation, or deamination. The altered bases cause mispairing and strand breakage during replication. This type of damage causes minimal change to the DNA double helix structure and can be repaired by the base excision repair (BER) pathways. BER corrects damaged DNA sequences by removing the damaged base and restoring the original base sequence using the complementary strand as a template. The first step of...
08:56
Terahertz Imaging and Characterization Protocol for Freshly Excised Breast Cancer Tumors
0 Views •
Cited by 18 •
2020
Freshly excised human breast cancer tumors are characterized with terahertz spectroscopy and imaging following fresh tissue handling protocols. Tissue positioning is taken into consideration to enable effective characterization while providing analysis in a timely manner for future intraoperative applications.
02:16
Differentiating Non-Transgenic and Transgenic Human Pluripotent Stem Cells into Neural Progenitor Cells
0 Views •
2025
This video demonstrates the generation of neural progenitor cells from non-transgenic and transgenic human pluripotent stem cells (hPSCs). hPSCs are differentiated into neural progenitor cells using a neuronal differentiation medium. While differentiation is induced in non-transgenic cells using small molecules, the transgenic hPSCs are differentiated by adding an antibiotic that induces the promoter for a neuronal differentiation-specific transcription factor.
10:15
Generation of Transgenic C. elegans by Biolistic Transformation
0 Views •
Cited by 27 •
2010
Transgenic worms are commonly used in C. elegans research. Described is a simple, yet effective, protocol to introduce transgenes into worms using biolistic bombardment with DNA-coated gold particles. The effort involved and results of bombardment compare favorably with microinjection for the generation of transgenic animals.
FAQs
Recognition sites define the boundaries of the DNA segment targeted for removal. The site-specific recombinase acts at those matching positions, cutting and then rejoining the surrounding DNA so the intervening transgene is released. Their placement therefore determines what is excised, whether that is the transgene alone or a larger cassette containing markers and regulatory elements.
After recombination, the original insertion is not expected to remain as an intact functional cassette. The intervening DNA is released, while the recombined recognition-site arrangement leaves minimal genetic material behind. This matters because residual selectable markers, regulatory elements, or other construct components could otherwise complicate interpretation of the engineered system.
Selectable markers and regulatory elements can add unwanted construct material to an engineered locus. Removing them produces a cleaner genetic model and reduces components that could complicate analysis of the intended design. This makes the resulting cells or organisms more suitable for examining the biological effects associated with the transgene itself rather than the complete engineering cassette.
A basic workflow begins by designing the transgene so the sequence targeted for removal is flanked by the appropriate recognition sites. The system then uses the corresponding site-specific recombinase to cut and rejoin those sites. Researchers assess the resulting genetic arrangement to confirm release of the intervening DNA and retention of the desired host sequence.
It is useful when an engineered sequence has completed its purpose but its continued presence could obscure later experiments. Excision supports cleaner genetic models, conditional studies, and development of engineered cells or organisms with fewer unwanted sequences. It can also help separate effects attributable to the transgene from effects associated with where the construct entered the genome.
Comparing a system before and after removal can help distinguish effects caused by the transgene itself from effects caused by its genomic insertion. That distinction is important because an engineered sequence may be accompanied by markers or regulatory elements, and its insertion site can contribute to observed results. Removing the cassette provides a cleaner basis for interpretation.