esiRNA non-coding mouse

The esiRNA technology is a powerful tool for loss of function gene studies. To illustrate the potency of silencing triggers quantitative real time (qRT)-PCR is often used to measure the knock-down rates. The knock-down validation on mRNA level was here performed 24 hours post transfection of esiRNA using qRT-PCR. To be able to assess knock-down rates, the expression levels of the mRNA of interest was compared to cells (HeLa or mouse ES) simultaneously transfected with Renilla Luciferase (negative control).

However, a more valuable measure of the knock-down potency in an RNAi experiment is the reduction in protein level. We therefore also validate the knock-down rates of esiRNAs on protein level using quantitative western blot analysis (Odyssey, Li-COR). The time point for maximum knock-down rate for each protein can vary significantly, as it is depending on factors such as protein-stability, turn-over rate or cell proliferation rates. The knock-down validation on protein level was here performed at 72 hours post transfection of esiRNA in HeLa cells.  To be able to assess knock-down rates, the expression levels of the protein of interest was compared to HeLa cells simultaneously transfected with Renilla Luciferase (negative control). 

Two approaches were used to monitor knock-down at protein level: 1. Specific antibodies for the protein of interest were used for the quantitative western blot analysis. 2. Proteins of interest were GFP-tagged on a bacterial artificial chromosome (BAC) and stably integrated into the genome of HeLa cells, allowing for near physiological expression (Poser I. et al Nat Methods. 2008 May;5(5):409-15). Using a GFP antibody the detection by quantitative western blot analysis of the protein of interest is straightforward.

* Average of 2 technical replicates

esiRNA IDAccession No.LNC RNA NameLNC RNA DescriptionEnsembl IDRefSeq IDKnock-Down RatePDF
MNC-00089-1BC031453NR_046233.195,05 ± 1,66PDF
MNC-00018-1AK01835284,59 ± 2,18PDF
MNC-00010-1AF03213083,26 ± 8,18PDF
MNC-00060-1AK01232383,11 ± 0,08PDF
MNC-00063-1AK01851983,31 ± 0,91PDF
MNC-00087-1AK02035580,5 ± 12,19PDF
MNC-00173-1AK03327980,1 ± 1,17PDF
MNC-00492-1AK01699280 ± 6,6PDF
MNC-00102-1AK021247C430039J16RikRIKEN cDNA C430039J16 geneENSMUSG0000009145178,28 ± 2,97PDF
MNC-00114-1AK142231Rassf8Ras association (RalGDS/AF-6) domain family (N-terminal) member 8


78,19 ± 5,50PDF
MNC-00150-1AK02037277,46 ± 9,02PDF
MNC-00174-1AK01654476,03 ± 15,08PDF
MNC-00103-1AK0023902700038G22RikRIKEN cDNA 2700038G22 geneENSMUSG00000086802NR_045042.1
75,45 ± 7,92PDF
MNC-00127-1AK010004NR_027965.172,24 ± 1,63PDF
MNC-00139-1AK13159271,97 ± 13,21PDF
MNC-00163-1AK01462869,68 ± 6,78PDF
68,54 ± 14,96PDF
MNC-00105-1AK00870667,82 ± 5,18PDF
MNC-00190-1AK00790866,83 ± 6,30PDF
MNC-00192-1AK0185189030418K01RikRIKEN cDNA 9030418K01 geneENSMUSG0000007047666,73 ± 18,92PDF
MNC-00145-1AK004173Trmt61btRNA methyltransferase 61 homolog B (S. cerevisiae)ENSMUSG0000008549266,58 ± 20,09PDF
MNC-00169-1AK018410Cdc73cell division cycle 73, Paf1/RNA polymerase II complex component, homolog (S. cerevisiae)ENSMUSG0000002636166,42 ± 11,15PDF
MNC-00134-1AK0147144833417C18RikRIKEN cDNA 4833417C18 geneENSMUSG00000086015NR_045187.166,05 ± 16,15PDF
MNC-00386-1AK15655265,81 ± 8,13PDF
MNC-00232-1AK01554463,48 ± 7,82PDF
MNC-00210-1AK002866NR_040757.162,36 ± 4,60PDF
61,58 ± 8,96PDF