PacBio Flashcards
1
Q
Describe a PacBio SMRT cell
A
Nano-scale chambers each containing a tethered DNA polyerase, which allow a CONTINUOUS DNA synthesis reaction to be imaged in real time.
2
Q
Describe how PacBio and Illumina sequencing are different
A
Illumina uses DISCONTINUOUS DNA synthesis where the 3’ Dye acts as a block to synthesis
PacBio sequencing is a CONTINUOUS DNA synthesis reaction with DNA sequence being imaged in real time
3
Q
where is the dye positioned in PacBio dNTPs?
A
the dye is on the triphosphate of the dNTP
4
Q
Describe the adaptors used in PacBio sequencing
A
- Template molecules are ligated to hairpin adaptors
- hairpin adaptors are syntehtic ssDNAs that form a region of complementary dsDNA
- hairpin adaptors allow the dsDNA to be opened into a large ssDNA circle
- Hairpin adaptors provide sites for sequencing primers to bind
5
Q
How many reads per run can be achieved in PacBio sequencing
A
350,000
6
Q
What is a typical read length for PacBio sequencing?
A
20,000
7
Q
How accurate is PacBio sequencing?
A
99.999%
