Extractions
These protocols are primarily involved with the DNA extraction step.
Purpose
To lyse open cells, to lyse open the nucleus and release the DNA, to prevent naturally occurring enzymes from degrading the DNA, to get rid of cell and nuclear debris, and to acquire only “pure” nucleic acids in aqueous form.
There are many types of extractions. We have included two common filter-based methods (QIAGEN DNeasy & QIAGEN QIAamp Micro). For recommendations of other methods of extraction, such as precipitation based extraction, bead based extraction, High Molecular Weight (HMW) extraction, or extraction for low-input material, please reach out to Athena or other CCG staff.
When extracting bird and mammal tissues, you are required to dispose of all supplies that come into contact with your tissue sample in a special biohazard waste receptacle. This includes disposal of gloves, tips, tubes, glass slides, and anything else (other than sharps) that has come into contact with the tissue sample during any step of the extraction procedure. Dispose of sharps in the red sharps container also marked as biohazard. When extracting DNA from any bird and mammal samples, you are also required to:
- Work in a Biological Safety Cabinet with cleaning protocol (located in Micro Lab)
- Wear a lab coat (along with gloves)
- Wear a disposable face shield
The reason for these extra safety measures when handling bird and mammal tissues is to prevent the possibility of contracting and/or spreading a disease (such as avian influenza). Remember, these precautions are for your safety and thus should be a priority to you.
For any Qiagen kit DNA extraction (Animal & Plant):
Buffer AL and Buffer AW1 contain guanidine hydrochloride, which is harmful if swallowed and irritating to eyes, respiratory system and skin. Wear appropriate clothing and gloves when handling. In the event of skin contact with Buffer AL and Buffer AW1, immediately wash the affected area with water.
Guanidine hydrochloride can form highly reactive compounds when combined with bleach (sodium hypochlorite). If liquid with Buffer AL or Buffer AW1 is spilled, clean with suitable lab detergent and water. If the spilled liquid contains potentially infectious agents, clean first with lab detergent and water, and then with 1% (v/v) bleach. (DNeasy Blood & Tissue and DNeasy Plant Handbook 7/2006)
Extraction Room Hazardous Waste Disposal
The container labeled Guanidine Hydrochloride/Ethanol (EtOH) Waste includes: EtOH, AL, mixed AW1/EtOH, and mixed AW2/EtOH waste. Reminder: only water, soap and diluted bleach may go down the sink. Please reach out to Grace if you have any questions regarding hazardous waste disposal.
CCG Google Calendar Reminder
Due to its small size, please note your plans to use the CCG’s DNA EXT Lab on the CCG Google Calendar prior to working there.