John Hay
John Hay Biomedical
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Human Anatomy & Physiology (Human Body Systems)
Unit 4: Unit 4: Patient PerspectivesHBS 4.1Molecular and Genetic Technology

Reading a DNA gel to decide who carries a variant

Separate DNA fragments by size, then use the band pattern in each family member's lane to say who carries the variant and who does not.

Builds on (2 levels back)inferred · high confidence
  • DNA carries a negative charge: The whole separation depends on DNA being pulled toward the positive electrode, so the direction of travel is not arbitrary.
  • Reading a value off a labeled scale: A ladder lane is a ruler. Sizing an unknown band means reading it against the nearest known rungs, the same skill as reading a thermometer.

Prerequisites are inferred: pending teacher review.

Re-learn the skill with worked practice and clear examples.

Now size a band against the ladder, and read a family's lanes as genotypes rather than as a picture.

Step 1: Size against the two nearest rungs
Find the ladder rungs immediately above and below the unknown band. The band's size is between those two numbers, nearer the rung it sits closer to. Report it as an estimate, because a ladder fixes only the values it carries.
Step 2: Count the bands in one person's lane
Each person has two copies of the gene. If the two copies are the same length, the lane shows ____ band. If they differ in length, the lane shows two bands, one for each copy.
Step 3: Compare lanes to compare people
Lanes are only comparable when they were run on the same gel for the same time. Matching a lane to the affected control tells you that person carries the same variant length, and nothing yet about whether they will show symptoms.
Practice

A ladder lane shows rungs at 1000, 500 and 250 base pairs. A patient's band sits between the 500 and 250 rungs, a little closer to the 500 rung. What is the best estimate of that fragment's size?

Reviewed
  1. A.About 150 base pairs
  2. B.Exactly 375 base pairs
  3. C.About 750 base pairs
  4. D.About 400 base pairs
Show the worked solution ▾

Answer: D. About 400 base pairs

  1. Step 1: Bracket it: The band is between the 250 and 500 rungs, so the answer must be between 250 and 500.
  2. Step 2: Use the position inside the bracket: It sits nearer the 500 rung, so estimate above the midpoint of 375. About 400 bp fits.

Why it's right: The band lies between the 250 and 500 rungs and closer to 500, so an estimate a little above the 375 midpoint is right. About 400 bp is the only option inside that bracket.

Why the others miss:
  • A: 150 bp is below the 250 rung, which would put the band FARTHER down the gel than the 250 rung. It is not.
  • B: The arithmetic midpoint is right but the certainty is wrong. A ladder brackets a value; it cannot fix it to the base pair, and the stem says the band is nearer 500 than the midpoint anyway.
  • C: 750 bp is above the 500 rung, which would put the band closer to the well than the 500 rung. It is not.

Aligned to Molecular and Genetic Technology · reading level ~grade 9

On a family gel, the affected-control lane shows a single band at 400 bp and the unaffected-control lane shows a single band at 600 bp. A family member's lane shows TWO bands, one at 400 bp and one at 600 bp. What does that lane show?

Reviewed
  1. A.The gel ran too long, so one band split into two
  2. B.This person carries one copy of each version of the gene
  3. C.The sample was contaminated, because a person cannot have two sizes
  4. D.This person carries two copies of the 400 bp version
Show the worked solution ▾

Answer: B. This person carries one copy of each version of the gene

  1. Step 1: Count the copies: Every person carries two copies of the gene, one from each biological parent.
  2. Step 2: Match each band to a copy: One band matches the 400 bp control and one matches the 600 bp control, so the two copies are different lengths.

Why it's right: Two bands in one lane means the person's two copies of the gene are different lengths: one matches the 400 bp version and one matches the 600 bp version. That is one copy of each.

Why the others miss:
  • A: Running longer moves bands farther down the gel; it does not split one fragment into two sizes.
  • C: Two bands is the expected result for a person carrying two different versions, not evidence of contamination. Contamination usually adds bands that match NEITHER control.
  • D: Two copies of the same version would give ONE band at 400 bp, because both copies would travel the same distance.

Aligned to Molecular and Genetic Technology · reading level ~grade 9

Guided notes

Fill these in as you work through the lesson.

Big idea: DNA is negatively charged, so an electric field pulls every fragment toward the positive end, and smaller fragments slip through the gel faster: position on the gel is a measure of size, and the pattern of sizes in a lane is the person's genotype.
Key terms: write the meaning
  • Electrophoresis (separating by charge and size):  
  • Ladder (the known-size ruler lane):  
  • Base pair (bp) (the unit of DNA length):  
  • Allele (one version of a gene):  
The rule

Smaller fragments travel   through the gel, so a band far from the well is   than a band that stayed near it.

Check yourself
  1. Why does every DNA fragment move the same direction, no matter its size? 
  2. If a lane shows two bands instead of one, what does that say about the two copies of the gene that person carries? 
  3. What would you have to know before you could call a band 'the disease allele'? 
Work one example

A ladder shows rungs at 1000, 500 and 250 bp. An unknown band sits between the 500 and 250 rungs, closer to 500. Estimate: roughly 400 bp. You cannot call it exactly 400, because the ladder only fixes the two values you read between.