Application of the nonradioactive biotin–streptavidin system to visualize AFLP fragments

Abstract

Amplified fragment length polymorphism (AFLP) fingerprinting is one of the most widely used molecular techniques in plant biology today. In this paper, we describe the application of the extremely sensitive nonradioactive biotin–streptavidin system to visualize AFLP fragments blotted onto nylon membranes. The protocol is tested for different plant species (from bryophytes, gymnosperms and angiosperms) and primer combinations. Advantages of this protocol over other nonradioactive detection methods are discussed, and the suitability of the method for laboratories without automated sequencing facilities are emphasized.

Clonal reproduction, vegetative multiplication and habitat colonisation in Tussilago farfara (Asteraceae): A combined morpho-ecological and molecular study

“…AFLP analyses were conducted using the S2 sequencing gel apparatus (GIBCO BRL by Life Technologies; Whatman Biometra). The protocols followed the methods described in Pfeiffer et al (2005) with the exception of running the pre-amplification reactions in smaller volumes (12.5 ml).…”

Vegetative multiplication and patch colonisation of Asarum europaeum subsp. europaeum L. (Aristolochiaceae) inferred by a combined morphological and molecular study

“…The AFLP-analyses were conducted according to the protocol of Pfeiffer et al (2005), using an S2 sequencing gel apparatus (GIBCO BRL by Life Technologies, Whatman Biometra). Before final analysis, ten combinations of selective PCR-primers with three or four additional bases compared to the adapter-sequences (and either the MseI-or EcoRI-primers biotinylated) were tested on a selection of five samples, including presumably identical and different samples.…”

Vegetative reproduction and clonal diversity in Rhytidium rugosum (Rhytidiaceae, Bryopsida) inferred by morpho-anatomical and molecular analyses

“…For the selective amplification, 11 primer combinations were tested and two combinations selected, which revealed high levels of intraspecific polymorphism in R. rugosum: 5' biotinylated ing was carried out using a S2 sequencing system (GIBCO BRL, Life Technologies, Gaithersburg, MD, USA) according to the protocol of Pfeiffer et al (2005).…”