©2019 by Christopher Monaco

 
  • Christopher Monaco

DNA Necklaces: The Hottest Fashion Accessory of the Spring

A new year brings new fashion and this year's hottest new accessories are DNA necklaces. That's right! Who knew genes could be so fashionable?

Our middle school makers showing off their DNA.

Introduction

Late last year I was asked to develop a #DIYbio activity for the Middle School Girls Makers Club at Decatur Makers. I decided this would be the perfect time to finalize a project that had been on my to-do list for a while: the DNA Necklace. This simple project is great for kids of all ages and can be adapted to any grade level depending on how in-depth the biological background is presented. I'll admit that I didn't come up this idea, and you can buy kits to create the DNA necklace from various bio-ed suppliers, however, this protocol is a bit cheaper and it's easier to get the materials because everything is available from Amazon or the Science Company.


Biological Context

Before we get to the actual protocol, let's talk about the science. Extracting DNA is an important part of biological research. If you want to look at what's in an organism's DNA, you need to get it out of the cell. And while there are many methods to do this, this extraction is probably one of the most basic.


Step 1. Cell Lysis

If you've taken any introductory biology class, chances are you know that DNA is found inside the nucleus of the cell. And you probably know that the nucleus is surrounded by the cell membrane. In order to get to that DNA we need to break apart the cell membrane and the nucleus. So how can we do that?


There are two main categories of cell lysis: physical lysis, using any phyical force to break cell membranes such as grinding or chopping; and reagent-based lysis, using chemicals or enzymes to degrade a cell membrane. For this protocol we will use a reagent-based method and so for this post I will focus on that method.


A phospholipid.

But before we talk about the reagent we will use, let's first recall the structure of cell membranes. A cell membrane consists specific arrangement of molecules called phospholipids. These molecules are amphiphilic, meaning that they possess both hydrophobic (water hating) and hydrophilic (water loving) properties. In the case of the phospholipid, one end of the molecule, referred to as the head, is made up of a hydrophilic phosphate group, while the other end consists of two fatty acid, hydrophobic tails.


Phospholipid Bilayer

In the presence of water, like in our bodies, these phospholipid molecules form a special arrangement called a bi-layer. This bi-layer exists with the hydrophobic tails all oriented towards the center and the hydrophilic heads oriented out. This configuration is energetically favored since the internal area where the hydrophobic tails exist contains virtually no water. This phosopholipid bi-layer is the basis for the cell membrane.

SDS Molecule

So, now that we know what the cell membrane is made of, we can start to think about what reagent-based approach we can take to degrade this structure. For this protocol we're going to be applying the principle of "like dissolves like" and use a molecule very similar in nature to the phospholipid: an anionic detergent. In particular for this protocol, we will be using Sodium Dodecyl Sulfate, or SDS (pictured here). You can probably tell from structure how similar SDS is to a phospholipid; and because of this similarity, SDS will happily mix in the phospholipid bi-layer. When this happens, though, the bi-layer structure becomes inherently unstable and as a results degrades and forms a single layer spherical structure called a micelle; thus lysing the cell.

A couple of other chemicals go into the cell lysis solution that it's worth talking about. First, tris(hydroxymethyl)aminomethane or Tris is added to the solution. Tris is a buffer, a chemical that resists pH change, that operates in the pH range of most living organisms. It will help ensure that our lysis solution doesn't become to basic or acidic which would result in DNA degradation. Second, we will add thylenediaminetetraacetic acid or EDTA. EDTA is a chelating agent, meaning that will bind metal ions in solution. This is important because may enzymes, including the DNA destroying DNase, require metal ions to work. By removing those ions, we can inhibit the activity of enzymes that may degrade our DNA.


Step 2. Precipitation

After lysing the cells, we're left with a soupy mess containing DNA and everything else that we're not interested in. We need some way to get the DNA out while leaving all the other garbage behind. To do that, we will use ice-cold alcohol, as either ethanol or isopropanol. If you read the protocol in the next section of this post, you'll notice that one of the first step is to swish with an 8% saline solution. That salt is important in the extraction process and will work with the alcohol to pull out or precipitate out DNA from the rest of the lysis solution.

The structure of DNA. Note the negative charges on the backbone.

All nucleic acids are polar, meaning they have an uneven distribution of charge. This allows them to easily dissolve in other polar solvents like water. The negatively charge backbone of DNA will be attracted to positive poles of the water molecules found in our lysis solution and won't want to come out of solution. By adding sodium ions, which have a stronger positive charge than the water, we will have the DNA preferentially bind to the sodium over the water. This essentially neutralizes the charge in the DNA backbone and makes the molecule less soluble in water.


Now, when we add alcohol, we introduce an environment where the DNA and sodium can more strongly interact than when in water. This means the DNA become even less soluble and precipitates out of solution. At that point we'll be able to start seeing tiny strands of DNA in the alcohol layer.

The DNA Necklace

A note, this protocol is a bit more sophisticated than your typical demonstration DNA extraction using dish soap. I have read that dish soap can work, but I was unable to get enough yield with dish soap to make the DNA visible.


Materials

  • 1% SDS Lysis Buffer (see below)

  • 8% Saline Solution

  • 95% Ethanol (ice cold) (colored if desired)

  • 15mL Conical Tubes

  • Microcentrifuge Tubes

  • Transfer Pipettes

  • Yarn

  • Super Glue

Protocol

Avoid eating or drinking for at least 20 minutes prior to performing the activity.

Carefully transferring DNA to necklace tube.
  1. Measure 5mL of the 8% saline solution into a disposable cup.

  2. Gently chew on the inside of your cheeks for about 15 seconds.

  3. Swish the saline solution around in your mouth for 30 seconds then spit back into disposable cup.

  4. Pour spit solution into 15mL centrifuge tube and then dispose of cup.

  5. Add 2 mL of Lysis Buffer to tube. Cap tube and gently invert five times to mix.

  6. Allow tube to incubate at room temperature for five minutes.

  7. Slowly add 6mL of cold EtOH to tube to create a layer on top of extraction solution.

  8. Let tube sit for five minutes to allow DNA to participate.

  9. Slowly invert tube five times to allow DNA to aggregate.

  10. Carefully transfer DNA and EtOH solution to a clean microcentrifuge tube. Place yarn in centrifuge tube hinge and seal cap with super glue.

Reagents

Preparing Extraction Buffer (1L)


Prepare 1M Tris-HCl pH 7.5 (200 mL)

  1. Dissolve 24.23g Tris-Base in 100mL of dH2O (a heated stir plate may help).

  2. Adjust pH to 7.5 using concentrated HCl.

  3. Make solution up to 200mL using dH20.

Prepare 0.5M EDTA pH 8 (200 mL)

  1. Dissolve 37.22g EDTA in 150mL of dH2O.

  2. Adjust pH to 8 using NaOH pellets.

  3. Make final volume up to 200mL using dH2O.

Prepare 10% SDS (100mL)

  1. Dissolve 10g SDS in 100mL dH2O.

Add 100mL of 1M Tris-HCl, 100mL of 0.5M EDTA, 100mL of 10% SDS, and 700mL of dH2O to a container and gently shake to combine.


Prepare 8% Saline Solution (500mL)

  1. Dissolve 40g NaCl in 400mL of dH20 (heating may be required).

  2. Make up solution to 500mL using dH2O.

Prepare Colored Ethanol (500mL)

  1. Dissolve 2 drops of liquid food coloring per 250mL of EtOH.

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