Good evening. I had recently ran a sequencing run of 24 barcodes on a flongle. Samples where amplification and barcoded utilizing the 16s barcoding kit from Nanopore. Upon loading my library and selecting the proper workflow I started to receive failed reads. The flongle hasn't stopped running but as of the last time I checked it only had 9 out 339 reads pass. What could have gone run and any recommendation on how to fix it?
Dear friend Joshua Segoviano
I'm sorry to hear that you're experiencing difficulties with your sequencing run. I understand how frustrating this can be, and I’d be glad to help you troubleshoot the issue together.
To start, let’s consider the preparation stage. Is it possible there were any challenges with sample amplification or barcoding? The concentration and purity of the library loaded onto the Flongle are also critical—do you feel confident about these parameters? Sometimes, even minor inconsistencies here can impact read quality.
Next, it’s worth reviewing the loading process. Was the library diluted according to Nanopore’s guidelines for the Flongle? Both overloading and underloading can lead to read failures, so ensuring the correct dilution is important. Additionally, have you checked whether the flow cell or the software settings might be contributing factors? If this is your first time using this particular Flongle and kit combination, that could also be relevant.
Exploring these areas may help us pinpoint the root of the problem. I also recommend checking the Nanopore community forums, as others may have encountered similar issues. Please let me know which aspects you think are most worth investigating, or if you have any additional details to share. I’m here to support you as we work through this.
Best regards,
Kaushik
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