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Paired end clusters

Comments

Illumina have developed technology for chenging of DNA strands within clusters. Below is a POSSIBLE scheme of cluster inversion.



PCR primers for one-read clusters (left) and for PE-clusters (right). Two types of PCR primers are attached to the surface of flowcell. One type of primers (red) has cleavable site in the 5' part (cross on red primer) and another primer (green) has cleavable part in the 3' part (green line);



After 30 cycles cluster will consist on a number of double-stranded bridges (befor (left) and after (right) last denaturation).


Cleavage and termination (cleavage leave phosphate group on 3' end) of green primer generate single-stranded templite of the first orientation. First starnd sequencing reaction may be done.



Primer terminator should be removed (phosphatase treatment (left)). Several cycles of two-dimensional PCR restore cluster (right).


Red primer should be cleaved as for the single-read sequencing (left) and sequencing of reverse strand (right).