Rapid typing of truffle mycorrhizal roots by PCR amplification of the ribosomal DNA spacers

Specific PCR‐primers as a reliable tool for the detection of white truffles in mycorrhizal roots

“…Molecular methods are therefore necessary. However, the use of primers designed for the amplification of the variable intergenic region (ITS) of ribosomal RNA genes could lead to amplification of the same region of the host plant (Paolocci et al, 1995 ;Mello et al, 1996), thus necessitating the designing of primers specific to T. borchii and T. magnatum.…”

“…The former is of little value and marketing of the latter is illegal. Since the morphological features of mycorrhizal roots do not allow positive differentiation of truffle species during their symbiotic phase, a range of PCR protocols, mostly based on amplification of the internal transcribed spacer (ITS) region of nuclear ribosomal RNA genes, have been developed (Henrion et al, 1994 ;Chevalier et al, 1995 ;Paolocci et al, 1995Paolocci et al, , 1997Mello et al, 1996 ;Gandeboeuf et al, 1997a ;Amicucci et al, 1998 ;Bertini et al, 1998).…”

“…The universal 18S rDNA primers NS1, NS2 (Dams et al, 1988), as well as the ITS1\ITS4 primers, were used in amplification reactions following the conditions described in Mello et al (1996). Nested PCR was performed by adding 2 µl of a 10-fold dilution of the ITS amplified using ITS1 and ITS4 from the first round of PCR to a fresh reaction mixture containing the P7 and M3 primers.…”

“…96540) from a fruit body collected in Ravenna (Emilia Romagna, Italy) and kindly provided by Dr Zambonelli ; the (Mischiati & Fontana, 1993). Total DNA was extracted from fruit bodies, ectomycorrhizal and non-mycorrhizal roots according to Mello et al (1996). Non-mycorrhizal roots of Populus alba L., P. nigra L., Quercus robur L., Corylus avellana L. and Tilia cordata Mill.…”

Taxonomy of the Truffles

“…Much of the molecular research aimed at identifying fungi in general (Gardes, White, Fortin, Bruns, & Taylor, 1991;Henrion et al, 1992;Buscot et al, 1996;Mello et al, 1996;Chillali, Wipf, Guillaumin, Mohammed, & Botton, 1998) and truffles in particular (Henrion et al, 1994;Paolocci, Angelini, Cristofari, Granetti, & Arcioni, 1995;Paolocci, Rubini, Granetti, & Arcioni, 1999) relies on comparison of sequences and RFLP patterns of the ITS region. Several phylogenetic studies of truffles have also used this DNA region (Roux et al, 1999;Diez et al, 2002).…”