Can I do paired-end sequencing with single-indexed libraries?

Yes, paired-end sequencing can be done with single-indexed libraries. This is because single vs. dual-indexed adapters have nothing to do with sequencing due to how the sequencing is performed. Sequencing is done using Read Primer 1 or Read Primer 2, both of which are present in all libraries. You can also do single reads even if you use dual-indexed kits.Unique Dual Indexing [...]

What is the difference between single and paired-end reads?

For single reads, only one end of the nucleic acid fragment is sequenced at your indicated length (e.g., 50-, 75-, or 100-bp reads) For paired-end reads, nucleic acid fragments are sequenced at the indicated length first from one end, then from the other, allowing the detection of insertions, deletions, and genomic rearrangementsUnique Dual Indexing [...]

How can I mitigate index crosstalk or ""sample bleeding"" issues?

The following steps may help you mitigate these issues: *Use dual-indexing kits, rather than single-indexing kits, in library preparation *Increase the percentage of PhiX spike-in, which will reduce the number of adjacent clusters with indexes *Lower the amount of library loaded to decrease the cluster densityUnique Dual Indexing [...]

What is index hopping?

Index hopping (or index switching) is a known phenomenon where sequencing reads are assigned to incorrect indexes, potentially leading to misalignment of reads or incorrect assumptions in downstream analyses. Using unique dual indexes minimizes index hopping and is recommended for sensitive applications.Unique Dual Indexing [...]

What are the advantages of using dual- versus single-indexed libraries?

There are several advantages of using dual indexes: Dual indexes increase the accuracy of sample identification by adding another identifier to minimize issues caused by index-hopping events Dual-indexed libraries allow higher multiplexing and the ability to run more samples per lane Dual-indexed libraries are sequenced with 2 additional reads, termed Index 1 (i7) Read and Index 2 (i5) read [...]