Bulking generated considerable bias in detection of amplified fragment length polymorphism variations in oat, fringed brome and smooth bromegrass

Abstract

Amplified fragment length polymorphism (AFLP) variations in bulk and plant‐by‐plant (PBP) samples of five oat (Avena sativa L.) accessions, one fringed brome (Bromus ciliatus L.) accession and two smooth bromegrass (B. inermis Leyss.) accessions were compared. The proportions of AFLP bands detected in PBP, but lost in bulk, samples of oat, fringed brome, and smooth bromegrass ranged from 19 to 31%, 40 to 44%, and 22 to 33% of the total bands scored, respectively. These lost bands had occurred at frequencies ranging from 0.1 to 1 in the PBP samples. These findings demonstrate bulking can generate substantial bias in detection of AFLP variations.

Genetic Variation and Relationships of Pedigree-Known Oat, Wheat, and Barley Cultivars Releaved by Bulking and Single-Plant Sampling

“…Some novel bands, that were fully absent in the eight single-plant samples but detected in their bulk sample, were also observed in two oat cultivars (Glen and AC Mustang) and one barley cultivar Betzes. The proportions of such novel bands were up to 2.2% of the scored bands and were relatively low when compared with those AFLP bands reported elsewhere (up to 8.7%; Fu et al 2003b). These results clearly demonstrate the upward and downward biases of bulking in detecting AFLP variation in these three crop species.…”

“…Applications of bulking strategies have played an increasingly important role in molecular characterization of plant germplasm (Yu and Pauls 1993;Divaret et al 1999;Gilbert et al 1999;Fu 2003;Fu et al 2003b). A recent literature survey showed 45% of the 181 articles of molecular germplasm characterization published in five leading scientific journals in crop science and breeding from 1997 to 2002 applied bulking strategies (Fu 2003).…”

“…Such an advantage would be more appealing when considering the large number of germplasm accessions widely held in national genebanks worldwide (FAO 1998). However, bulking homogenizes the within-accession variations, if any, and can introduce biases in detecting alleles occurring at low frequency either due to technical limitation or the use of small sample size (Michelmore et al 1991;Divaret et al 1999;van Treuren 2001;Fu et al 2003b). Biased estimates of genetic variation in bulk samples would reduce the accuracy of genetic distance estimates among germplasm accessions and would consequently affect the inferences of genetic relationship that are based on the estimated genetic distances (Yu and Pauls 1993;Divaret et al 1999;Mellish et al 2002;Fu et al 2003a).…”

“…While the accuracy of detecting DNA polymorphism in bulk samples has been fairly evaluated for random amplified polymorphic DNA (RAPD) markers (Williams et al 1990), few in-depth studies (van Treuren 2001;Fu et al 2003b) have been made for other marker techniques such as amplified fragment length polymorphism (AFLP) markers (Vos et al 1995). Also, the robustness of genetic relationship inferences against bulking biases has not been adequately addressed (Fu 2003).…”

Assessment of the congruity of genetic relationships and variation revealed by individual- and bulked-samples-based approaches using RAPD and ISSR markers in Japanese turnip (Brassica rapa ssp. rapa) cultivars

“…In this study, scoring only of the clear band of the method according to individual and each bulk method was carried out independently, and high coincidence was obtained between genetic-relatedness. However, in the bulk method, use of a small sample size produces distortion in the detection of rare alleles (Divaret et al 1999, Fu et al 2003b, Mailer and May 1999. On the other hand, if sample size of bulk becomes large, a large difference in banding pattern will no longer be found (Divaret et al 1999).…”