Sequence diversity is important for good reads on illumina seq platforms, particularly at the beginning, however is it that important AFTER the clusters/ colour matrices have been identified/ calibrated in the first few bases
ie, if I have good sequence heterogeneity for the first part of a run that establish good clustering, followed by spans of homogeneity, is going to make the run significantly worse? Do you need good heterogeneity throughout the run?
And what is a sufficient length of the initial length of heterogenous sequence? On non-patterned flow cells (Nextseq/ Miseq), the first 7bp seem critical? ButI have also read up to both base 11 and 25bp are important for colour matrix/ filtering?
Yes, it is necessary to have sequence diversity in Illumina run to get good separation within clusters and get good quality of signal and finally the sequence. For less diverse sequences, spike-in's are used to maintain good heterogeneity in the sequencing run.
Those initial base pairs you are talking about are necessary because the adapter sequence is same for all, so the initial random bases will indicate that transition of the sequencing cycle is proceeding to the DNA fragment. If all the sequences have same sequence after initial few bases, the camera will just get blind by the signal from all cluster at the same time.
Conclusion:
Sequence diversity/heterogeneity is required for all steps in Illumina sequencing.
- Badges
- Science topic
- Similar topics
- Psychology
- Mental Processes
- Learning