[PDF][PDF] ARTISAN PCR: rapid identification of full-length immunoglobulin rearrangements without primer binding bias

MT Koning, SM Kielbasa, V Boersma… - British journal of …, 2017 - research.rug.nl
MT Koning, SM Kielbasa, V Boersma, HPJ Buermans, SAJ van der Zeeuw, CAM van Bergen
British journal of haematology, 2017research.rug.nl
B cells recognize specific antigens by their membrane-bound B-cell receptor (BCR).
Functional BCR genes are assembled in pre-B cells by recombination of the variable (V),
diversity (D) and joining (J) genes [V (D) J recombination]. When B cells participate in
germinal centre reactions, non-templated point mutations are introduced into BCR genes by
somatic hypermutation (SHM)(Rajewsky, 1996). V (D) J recombination and SHM create
virtually unlimited BCR repertoires. Many research applications require identification of …
B cells recognize specific antigens by their membrane-bound B-cell receptor (BCR). Functional BCR genes are assembled in pre-B cells by recombination of the variable (V), diversity (D) and joining (J) genes [V (D) J recombination]. When B cells participate in germinal centre reactions, non-templated point mutations are introduced into BCR genes by somatic hypermutation (SHM)(Rajewsky, 1996). V (D) J recombination and SHM create virtually unlimited BCR repertoires. Many research applications require identification of fulllength BCR sequences expressed in biological samples. BCR are commonly identified by multiplex polymerase chain reaction (PCR) with upstream primers that anneal to conserved stretches in variable or leader gene segments. BIOMED-2 is the current European standard protocol to assess BCR clonality as diagnostic criterion for lymphoma (van Dongen et al, 2003). However, a single point mutation in a primer binding site can obscure a dominant clonal BCR (Wu et al, 2009), whereas reliable identification of functional BCR genes is essential to avoid erroneous conclusions about the BCR (Koning et al, 2014). Therefore, several investigators have emphasized the need for an unbiased method of BCR amplification (Warren et al, 2013; Georgiou et al, 2014; Koning et al, 2014). We here describe ARTISAN PCR (Anchoring
research.rug.nl