Hello All,
We are trying to perform single cell RNA-seq in our facility. We would like to do this in a high-throughput manner on our robotic system and are working off of this published protocol:
We wanted to and have done the protocol in hand first to work out any issues before conducting the experiment on the robot. We are using a Universal RNA Standard diluted down to a single cell concentration (10pg/ul) for testing.
We have not been able to get past the cDNA stage as we are seeing a strange BioA trace when comparing to that in the troubleshooting table of the Picelli Paper: http://www.nature.com/nprot/journal/....2014.006.html.
Attached are our BioA traces as well as TapeStation results of the same sample that also yield inconsistent results.
We have played with concentration as well as PCR cycles and still see this "double-hump" in the traces.
Any advice or thoughts would be greatly appreciated!
We are trying to perform single cell RNA-seq in our facility. We would like to do this in a high-throughput manner on our robotic system and are working off of this published protocol:
We wanted to and have done the protocol in hand first to work out any issues before conducting the experiment on the robot. We are using a Universal RNA Standard diluted down to a single cell concentration (10pg/ul) for testing.
We have not been able to get past the cDNA stage as we are seeing a strange BioA trace when comparing to that in the troubleshooting table of the Picelli Paper: http://www.nature.com/nprot/journal/....2014.006.html.
Attached are our BioA traces as well as TapeStation results of the same sample that also yield inconsistent results.
We have played with concentration as well as PCR cycles and still see this "double-hump" in the traces.
Any advice or thoughts would be greatly appreciated!
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