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Hi all. I have some RNA-Seq data from the sequencing of a green algae in co- culture with a bacterium. The bacterial cell numbers were reported to be in a 50:1 ratio with the green algae. However, only ~7% of the reads are ascribed to bacteria, the rest are algae/ plant. I have argued myself silly around this matter but this seems a low ratio of bacterial to algal signal but I can think of no way of knowing. Ultimately the aim is to publish a comparison of functional activity amongst based on metatranscriptomic reads from both algae and bacteria. I'm concerned about what would be an expected (publishable) signal from the bacterial population for this kind of study.
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How was the RNA prepared? -
Total RNA was prepared by grinding a frozen pellet followed by the Ambion ToTALLY RNA kit. cDNA libraries were prepared after double depletion of rRNA using the plant, then bacterial RiboZero kits.Last edited by chc*; 05-25-2016, 11:13 PM.Comment
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I wouldn't expect the cells to lyse at exactly the same rates. Also, algal genomes are far bigger than bacterial genomes, and the cells are bigger as well. So I would expect plant reads to be much more abundant than the cell count ratio would suggest. Maybe you could find out the expected amount of RNA in a plant vs bacterial cell and see if that makes the math work out?Comment
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Love this - good data definitely starts from good input, and poor input can only give relatively poor data. I particularly like the mention of Nanodrop/absorbance based methods for quantification. It's such a toss up if you'll get an accurate reading or what amounts to a randomly generated number, and a lot of library/sequencing related issues can be traced back to poor quant.07-01-2026, 11:43 AM
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