410 bp peak in bioanalyzer results

[mhkiani]

We are trying to do RNA-seq for mitochondrial and nuclear RNA and we sent the sample to genomics center, they made th elibrary and they found a prominent peak in all of sample around 410bp. Any guess about that?



My samples are from plant

[thomasblomquist]

Depending on the type of library preparation method you use, it could be heterodimers between non-complementary templates within your sample preparation. These heterodimers have complementarity on either ends (adapters are homologous), but are mismatched or have non-canonical watson-crick base pairing in portions in the center. This "bulky" 3-D structure will electrophorese slower than the typical library prep, despite having insert fragment size that is the expected length.

Because of this phenomenon, becareful to assume this is your true library length. Try a denaturing chip (such as an RNA-chip on an agilent bioanalyzer), and see if the upper peak disappears. If it does, then it was a heterodimer population of partially homologous strands of nucleic acid.

There are other possible causes for this peak, but the above is far and away the most likely.

Regards,

-Tom