Why does variant detection only support human and not other species, and what are the limitations?

PacBio HiFi sequencing variant detection supports all species. The reason why the current results are currently still mainly human, this is mainly because the current human reference genome is currently the most accurate.

Is the advantage of PacBio compared to Nanopore in HiFi reads?

The main advantage of PacBio SMRT sequencing is that there is no systematic error, which is why HiFi reads are so accurate.

What is the minimum sequencing depth required for a standard HiFi assembly? How much sequencing depth would be better?

Generally speaking, it depends on how many ploidy the specific assembled genome is. Take the human genome for example, we recommend a minimum of 15X-20X, if the genome is more than one haplotype need to increase the data of 10X HiFi. But it also depends on the complexity of the genome, if it is a complex genome may require an additional 20X HiFi data for one more haplotype.

Ultra-low starting volume library building requires genomes below 500M, can we do other species besides arthropods such as insects?

We have only tested insects and arthropods for the time being, because the DNA content of individual individuals of these samples are very small. Microorganisms we have not tested for now. To clarify, the genome is less than 500M, this is for De Novo application, if it is resequencing there is no genome size limit.

HiFi data is more accurate, can you estimate the genome size?

Actually, all of them can be estimated by k-mer.

The genome size of sandfly estimated by GenomeScope is 306 Mb, and the assembly obtained is 370 Mb, is it underestimated by GenomeScope or overestimated by assembly?

This is mainly related to genomic heterozygosity. When assembling, if the difference between two haplotypes is large, then the difference will be reflected in the assembly result, and it will look larger.

What is the process and enzyme of PCR for ultra-low starting volume library building?

We use long range PCR. PacBio has done a lot of tests to select the two PCR enzymes that can better cover the genomic regions with high GC and low GC as much as possible. After sample interruption, we split the sample into two parts, one for PCR A and the other for PCR B. The instrument is able to obtain the most uniform genomic coverage.

PacBio SMRT Sequencing Q&A

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PacBio SMRT Sequencing Q&A

General Questions