Page 9 - Hooked on Herefords Oct 2022

PBB Update Megan Ellett | PBB Manager

What's behind the N4 DNA error?

PBB Data and Genetics Manager Megan Ellett says some as that means the sample isn’t sealed correctly, allowing
PBB Neogen DNA clients have been receiving N4 errors the buffer to leak and the sample to dry out.”
following diagnostics of their SNP tested animals and she’s
outlined how breeders can best reduce that result. When it comes to hair samples, she says it’s best practice
to pluck the hair sample from the animal and not cut it.
An N4 error means that the sample submitted had “DNA is extracted from a hair sample via the root or bulb
insufficient DNA material for an appropriate SNP. at the end of the hair follicle.”
Megan said the main reasons for an “N4” result are
generally insufficient sample material or a contaminated To ensure a result can be obtained and the sample reaches
sample. an acceptable call rate, more than 30 hairs with roots
attached need to be collected. The ideal place to take a
There are two key steps to helping reduce the chances of sample is the tail switch. The more hair follicles supplied
returning an N4 error when DNA testing. the better. A Neogen approved hair card should be used
Those steps are choosing to test at a high level, such (available from the PBB office).
as the 100K SNP test and taking additional care when
harvesting a DNA sample. Samples can be cross-contaminated if they get wet or
dirty. Contamination can also be caused from dyes such as
Analysis on the N4 results has found that samples with those found in animal markers, animal health treatments
less than a 97% call rate (that means the percentage of and cleaning agents etc. Heat exposure can also lead to
SNP markers that could be taken from the DNA sample) degradation of the sample if stored in direct sunlight or a
will not always pass the AGBU quality control measures for hot position.
inclusion in evaluations.
The correct storage of a sample is critical – see the PBB
Using the higher-level testing, such as 100K is a strong website: www.pbbnz.com services/genomics/faqs/
step towards helping eliminate an N4 result, as it means • Hair Collection Guide: https://bit.ly/3E6ibie
an improved chance of reaching the required 97% call rate • Reasons for Sample Fails: https://bit.ly/3fvTEsL
needed for AGBUs genomic evaluation. • TSU Applicator Instructions: https://bit.ly/3y2UZxC

Megan says DNA sample quality has a significant role to In cattle, DNA can cross the placenta, which can mean
play in acceptable call rates. A lower call rate typically mixed genotypes in non-identical/fraternal twins –
means poorer DNA quality. The simplest option when otherwise known as blood chimeras.
genotypes are flagged with an N4 error is resampling (by
collecting a new sample). “A blood chimera will generally fail it’s DNA quality control
and have lower call rates,” says Megan.
“The easiest way to ensure less N4 errors is to submit
quality DNA samples, that means taking particular care Identical twins have identical DNA, so that means going
when collecting samples,” says Megan. forward any DNA sample that fails to reach the 97% call
rate threshold will first be identified to see if it’s a twin. If
When using the Allflex TSU system it’s vital that the ear that’s the case, a resample using hair should be submitted
punch is sitting in the buffer liquid in the vile as this helps as it’s been found this type of sample results in higher
it retain viability. percentage call rates for identical twins.

TSUs should be kept at room temperature or in the
fridge, they should not be placed in the freezer. Allflex Key Points
recommends DNA testing occurs within 12 months of
sample collection. • There are ways to prevent N4 errors.
• Follow the correct sampling process to
She also urges all breeders to ensure the tissue sample ensure good quality samples.
makes it into the vile and isn’t trapped in the TSU cap • Test at highest level possible e.g. 100K.
accidentally. • Test identical twins using hair sample.

“There also shouldn’t be any hair sticking out of the tube

Hooked on Herefords NZ Herefords Newsletter. October, 2022 | 9