Vitis 47 (2), 97–104 (2008) ‘Sangiovese’ and ‘Garganega’ are two key varieties of the Italian grapevine assortment evolution M. CRESPAN1), A. CALÒ1), S. GIANNETTO1), A. SPARACIO2), P. STORCHI3) and A. COSTACURTA1) 1) C.R.A. Centro di Ricerca per la Viticoltura, Conegliano, Italy 2) Istituto Regionale della Vite e del Vino, Palermo, Italy 3) C.R.A. Unità di Ricerca per la Viticoltura, Pratantico, Italy Summary provinces of Verona and Vicenza. ‘Sangiovese’ is better known than ‘Garganega’ and was mentioned for the first Two synonymous cases have been found using a set time by SODERINI (1590) as ‘Sangiogheto’; actually it is the of 11 SSR markers: ‘Garganega’ and ‘Grecanico do- most commonly cultivated black grape variety in Italy and rato’; ‘Catarratto bianco comune’, ‘Catarratto bianco is the basis for the production of famous wines, such as lucido’ and ‘Catarratto bianco extra lucido’. Molecular Chianti and Brunello di Montalcino. ‘Garganega’ is prob- data at 36 SSR loci showed that ‘Sangiovese’ and ‘Gar- ably less known but perhaps even older than ‘Sangiovese’, ganega’ represent two key varieties in the Italian am- as it was cited under this name as one of the grapes in the pelographic assortment evolution, as they both have a province of Padova by Pier de’ Crescenzi in the 13th cen- first degree relationship with numerous wine varieties. tury (CALÒ and COSTACURTA 2004). It is famous for the ex- ‘Sangiovese’ showed this link with ten varieties: ‘Foglia cellent Soave and Gambellara wines and its origins are still tonda’, ‘Frappato’, ‘Gaglioppo’, ‘Mantonicone’, ‘More- unknown. llino del Casentino’, ‘Morellino del Valdarno’, ‘Nerello As a consequence, the hypothesis is plausible that they mascalese’, ‘Susumaniello’, ‘Tuccanese di Turi’ and could have generated progenies over the centuries or could ‘Vernaccia nera del Valdarno’. Seven varieties resulted be related to other cultivars growing in the same area. closely related to ‘Garganega’: ‘Trebbiano toscano’ ali- Many possible close kinships emerged from the compari- as ‘Ugni blanc’, ‘Albana’, ‘Empibotte’, ‘Malvasia bian- son of ‘Sangiovese’ and ‘Garganega’ molecular profiles ca di Candia a sapore semplice’, ‘Marzemina bianca’, with those of hundreds of genotypes collected in the data- ‘Catarratto’ and ‘Greco del Pollino’. However, being base of Centro di Ricerca per la Viticoltura. These indica- ‘Sangiovese’ parents disputed and those of ‘Garganega’ tions were further investigated by increasing the molecular still unknown, it was not possible to determine the uni- analyses up to 36 SSR loci. Many varieties showed to be vocal direction of the various crosses. Identification of involved in a parent-offspring link, some of them having the “missing” parents would allow these genealogical economic or historical importance, such as ‘Trebbiano trees to be drawn up with greater precision. toscano’ alias ‘Ugni blanc’, ‘Catarratto’, ‘Albana’, ‘Frap- pato’, ‘Gaglioppo’, ‘Nerello mascalese’ and ‘Marzemina K e y w o r d s : Catarratto, Trebbiano toscano, Gre- bianca’. These results trace out the role played by ‘Sangio- canico dorato, pedigree, SSR, synonyms. vese’ and ‘Garganega’ in the appearance of many and well known Italian varieties. Introduction Material and Methods Researches on grapevine varieties pedigree determina- tion have been increased in the last years by means of mic- P l a n t m a t e r i a l : More accessions belonging to rosatellite markers (SSR) (SEFC et al. 2001), contributing to 22 varieties were used for genotyping (Tab. 1). They came clarify the evolution of the current ampelographic assort- from the Centro di Ricerca per la Viticoltura collections ment. One of the most interesting examples is the discov- of Conegliano (Treviso), Arezzo and Turi (Bari), and also ery of the origin of dozens of French varieties from a single from private vineyards in Tuscany and Sicily. pair of parents, ‘Pinot’ and ‘Gouais’ (BOWERS et al. 1999 a, N u c l e a r S S R l o c i a n a l y s i s : Genotyping BOURSIQUOT et al. 2004). This explaines why they have less was performed with 11 SSR loci (basic set) routinely em- allelic variability than groups of varieties in other countries ployed at the Centro di Ricerca per la Viticoltura of Coneg- with strong viticultural traditions and, using specific tests, liano for cultivar identification (VVS2, THOMAS and SCOTT they can be precisely assigned to their corresponding geo- 1993; VVMD5, VVMD7, VVMD27 and VVMD28, BOW- graphical region of origin (SEFC et al. 2000). ERS et al. 1996 and 1999 b; VrZAG62 and VrZAG79, SEFC ‘Sangiovese’ and ‘Garganega’ are ancient and re- et al. 1999; ISV2, ISV3, ISV4 and VMCNG4b9, CRESPAN nowned wine varieties, the former being widely cultivated 2003). Since two groups of synonymous varieties were throughout Italy, the latter in Veneto region, mainly in the found (‘Grecanico dorato’ and ‘Garganega’; ‘Catarratto Correspondence to: Dr. M. CRESPAN, C.R.A. Centro di Ricerca per la Viticoltura, Viale 28 Aprile, 26, 31015 Conegliano (TV), Italy. E-mail: [email protected] 98 M. CRESPAN et al. T a b l e 1 Kingroup v2 program was used to calculate the likeli- hoods of the hypothesized pedigree relationships and their List of 22 analysed varieties significance versus alternative relations. The kinship relat- edness estimator was applied; parent-offspring relation as Sangiovese primary hypothesis and unrelated as null hypothesis. Ciliegiolo The haplotype probability, i.e. the expected frequence Foglia tonda of an i allele at a particular locus was calculated, given the Morellino del Casentino Hardy and Weinberg law assumptions and basing on the Morellino del Valdarno third Mendel’s law, with the following formula: Tuccanese di Turi HP = 2p - p 2 Gaglioppo di Cirò i i where p is the frequence of allele i, computed with Cervus Vernaccia nera del Valdarno i Nerello mascalese 3.0.3 software (www.fieldgenetics.com). The loci non in Mantonicone HW equilibrium, as calculated by Cervus, were discarded Susumaniello (VVMD7, VrZAG62 and ISV3). The SSR loci localized on Garganega the same linkage group (ADAM-BLONDON et al. 2004, MER- Grecanico dorato DINOGLU et al. 2005) were considered as a single locus (15 Marzemina bianca LGs were used) and the allele univocally shared with ‘San- Catarratto bianco comune giovese’ or ‘Garganega’ having the lowest frequence was Catarratto bianco lucido chosen for HP computation; when the common allele was Catarratto bianco extra lucido not univocally identifiable, that with greater frequence was Trebbiano toscano selected, consequently the corresponding estimate may be Malvasia bianca di Candia a sapore semplice biased in excess. The calculations were done with Excel Empibotte computation sheet. The total haplotype probability, there- Albana fore the probability to find a particular allele combination, Greco del Pollino obtainable from the supposed parentage relationship, was computed as the product of the HPs at all LGs considered. bianco comune’, ‘Catarratto bianco lucido’ and ‘Catarratto bianco extra lucido’), only one sample for each cultivar was used and the analysis was continued for 19 varieties Results and Discussion with 25 nSSR loci, making totally 36 nSSR loci: VVS1 and VVS29 (THOMAS and SCOTT 1993); VVMD17, VVMD21, S y n o n y m s : Genotyping results with the basic VVMD24, VVMD25, VVMD26, VVMD31, VVMD32 set of SSR markers revealed two synonymies (probability and VVMD36 (BOWERS et al. 1996 and 1999 b); VrZAG21, of identity: 7.3x10-15). Firstly, 4 accessions of ‘Garganega’ VrZAG64 and VrZAG83, SEFC et al. 1999; VMC1e12, and 4 of ‘Grecanico dorato’ showed the same molecular VMC4g6, VMC2h9, VMC3d7, VMC2g2, VMC6e10, profile, confirming what previously pointed out by VAN- VMC4h6, VMC4c6, VMC2h4 and VMC5g6.1 (Vitis Mic- TINI et al. (2003) on just 2 samples. ‘Garganega’ has been rosatellite Consortium); scu05, SCOTT et al. 2000; UCH11, known in Veneto since at least 1200 and has great morpho- LEFORT et al. 2002. logical variability (COSMO and POLSINELLI 1960, CALÒ and Multiplex PCR of two or three SSR loci were suit- COSTACURTA 2004). The first citation relating to ‘Grecanico ably arranged based on expected allele lengths. The PCR dorato’, growing in Sicily, dates back to the end of 17th reaction mixture (25 µl final volume) contained 20 ng century (PASTENA 1969) and different phenotypes were de- total DNA, 10 µl Eppendorf HotMasterMix (2.5 x) and scribed also in this case. 5 pmoles of each primer. The PCR was performed in an AB In order to detect possible synonymies, suggested by 9700 thermal cycler with the following steps: 1 min 30 s at the Sicilian name of this cultivar reminding to a hypotheti- 94 ° C; 35 cycles at 94 °C for 30 s, 55 °C for 30 s, 65 °C for cal Greek origin, its molecular profile was compared with 30 s; 65 °C for 7 min and a final step of at least 10 min at those of the Vitis microsatellite databases of Centro di 8 °C to stop the reaction. Five µl of the PCR product were Ricerca per la Viticoltura of Conegliano (Italy), University tested on 2 % agarose gel; on the basis of signal intensity, of California, Davis (USA) and University of Crete, Her- 0.75-1.5 µl of amplified DNA were used for electrophore- aklion (Greece) http://www.biology.uoc.gr/gvd/ as well sis onto a sequencing gel (5 % polyacrylamide, 1 x TBE, as with genotypes from various references in literature. 7 M urea). Amplification products of cultivars with alleles ‘Garganega’/‘Grecanico dorato’ showed to be different of known molecular size were used as references for allele from all previously described varieties. sizing. Allele bands were revealed by silver staining and The second case of synonymy regarded ‘Catarratto’, visually scored at least twice, as reported in CRESPAN and the most widely white wine variety actually growing in MILANI (2001). Sicily. Three phenotypes were selected in the course of S t a t i s t i c a l a n a l y s e s : The molecular database time: ‘Catarratto bianco comune’, ‘Catarratto bianco lu- used for data elaboration encompassed the SSR profiles of cido’ and ‘Catarratto bianco extra lucido’; the first two are 668 wine and table cultivars.