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When followed exactly as written, this protocol is simple and requires very few steps. Whatman sells a proprietary card-washing reagent which is quite expensive, and the literature shows mixed success using SDS as a substitute. Whatever you choose, it dissertation on the media be able to wash away proteins and other chemicals and denature nucleases.
For very concentrated samples, elution is not much more difficult than a direct-to-PCR protocol. Things like cheek swabs, blood spots, and other sample types with large quantities of DNA can be extracted using commercial kits or Whatman protocols.
However, for projects where the samples are very dilute and the entire sample area needs to be processed instead of just a few punchesprotocol optimization becomes more complicated. Consider the challenges in processing large quantities of paper:.
The amount of elution buffer required to apparel buyer resume submerge the paper can be at odds with the size of the tube, especially if you plan on ethanol precipitating and concentrating the nucleic acid later. That requires double the sample volume of ethanol achieve, so make sure you leave enough room!
Some researchers solve this by using concentrator tubes or processing the paper directly in kits that employ the bind-wash-elute column method, but that can be expensive. It also helps to reduce the number of steps, including tube transfers, to minimize loss of product. Many kits with built-in card protocols allow you to put the paper directly into the elution medium usually a buffer, a detergent, and proteinase K rather than washing and drying it first.
Our lab prefers kits that allow elution in very small volumes buy economics essay samples of low someone to write my essay paper. Incubating the tubes on ice and avoiding agitating the tubes during the wash steps can help reduce product loss. In non-column protocols, sometimes the vortexing or shaking necessary to elute the nucleic acid can also start to break down the paper, leaving behind paper fuzz in the tube.
The paper fuzz usually does not end up in the final eluate in kits with micro-column methods, but it can be frustrating when attempting to purify nucleic acid by ethanol precipitation after using a non-kit elution protocol. Some paper fuzz is usually left behind, even if you try to spin it out and transfer the supernatant to a new tube, not to mention that transferring to a new tube introduces an opportunity for product loss.
Leaving the fuzz behind tends not to interfere with PCR reactions, but it does make the ethanol wash take longer to evaporate from the tube, increasing the risk that you will either contaminate your samples or incompletely dry them.
Another downside to the FTA cards is that they are easy to contaminate, especially with sensitive projects involving deep sequencing and metagenomics, which are already very sensitive to small amounts of contamination. FTA cards might not be ideal for these types of projects.
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Collecting biological samples in the field can be difficult, since storage conditions outside of the lab are often less than optimal.
The Whatman FTA Carda filter paper product manufactured by GE Health Care, is a paper matrix laced with a proprietary mixture of chemicals that lyse cells and stabilize nucleic acids on contact for long term storage at room temperature. FTA cards are typically used when storage space and freezing capacity essay on gandhi the sample collection end of a given project is limited or non-existent, because they can be stored in boxes, bags, drawers, or even sent through the mail without ice.