Task 1 Construction of a geographic map of genetic types

SYNTHETIC MAPS OF GENE DIVERSITY AND PROVENANCE PERFORMANCE FOR UTILIZATION AND CONSERVATION OF OAK RESOURCES IN EUROPE

Task 1 Construction of a geographic map of genetic types

Aim : The objective is to build a geographic map of CpDNA haplotypes. The map will be based on the analysis of chloroplast DNA polymorphism according to a systematic sampling of populations every 50 km throughout western Europe. But before analysing with this sampling, populations present in provenance tests will also be analysed. The map of genetic types will be compared with the map of pollen deposits to retrace postglacial recolonization routes.

Subtask 1.1. Standardization of molecular techniques for analysis Chloroplast DNA polymorphism

Participants Results Research action

P1, P3, P4, P5, P6, P8, P9, P10 R4 Themap can only be constructed if standard techniques are used for the analysis of CpDNA polymorphism. Methods have been developed and tested by partner P1 and will be shared with other participants involved in Task 1 during a 1 week technical meeting.
The molecular technique used is a PCR-RFLP assay. For the time being, 28 different haplotypes can be identified from amplification products with 4 primer pairs, digested with 3 restriction enzymes. Scoring of haplotypes and construction of data files will be standardized.

Subtask 1.2. Cp DNA analysis of populations established in provenance tests (Table 1)

Participants Results Research action

P1, P2, P3, P4, P5, P7, P9, P10, P11 R1,R5 For each provenance, buds or leaves will be collected from 5 trees per provenance. DNA will be extracted and the PCR-RFLP assay will be made on each sample to identify the haplotype(s) present in the provenance.
Results will be stored in the provenance data base (Subtask 2.1). Efforts will be made during the project to obtain and analyze populations from eastern European countries especially from the balkan region, which is considered as a major glacial refugium.

Table 1. Distribution of activities for CpDNA analysis in provenance tests

Participant hosting the test and collecting the material

Species

Prov. Test

(see 1st column annex1)

Nb of prov

Participant doing the PCR-RFLP assay.

P1 Q.petraea 1 120 P1

P2 Q.petraea
Q.robur
2

3 116

68 P3

P3

P3 Q. petraea 4 34 P3

P4 Q.petraea
Q.robur
5

6 39

11 P9

P9

P5 Q.robur 7 30 P5

P7 Q.robur 10 23 P9

P7 Q.robur 9 10 P9

P10 Q. petraea 12 12 P10

P10 Q.robur 13 15 P10

P11 Q.petraea+robur 11 24 P8

Task 1.3. Cp DNA analysis of populations sampled on a grid of 50 km and construction of the CpDNA geographic map (Table 2)

Participants Results Research action

P1, P3, P4, P5, P6, P8, P10, P11 R1,R5 Samples will be collected throughout Europe on a grid system (50 km*50km). In the nearest oak forest of natural origin, buds or leaves will be collected on 5 trees per forest (on each species if both are present). The PCR-RFLP assay will be conducted as in subtask 1.2. Haplotypes present in each forest will be identified.

Table 2: Distribution of activities for CpDNA analysis on the grid system

Country

Maximum number of sample points

Collection of samples done by:

Samples analysed by:

Austria 32 P8 P8

Belgium 12 P1 or P5 P1 or P5

France 168 P1 P1

Germany 135 P3 P3

Great Britain 70 (10*) P4 P4

Great Britain 2 P9 P9

Italy 80 Italian Subcontractor P6

Netherlands 12 P5 P5

Spain 146 P10 P10

Portugal 23 P10 P10

Sweden/Norway 20 P11 P8

Switzerland 15 P6 P6

* : 70 geographic points on the grid system, but only 10 natural stands.

Subtask 1.4. Analysis of pollen fossil deposits

Participant Results Research action

P12 R2 The current pollen database comprises data on pollen deposits collected in about 550 points distributed throughout Europe (Mostly France, Great Britain, Spain, Poland ..). Data on 300 additional points will be added through cooperative efforts with colleagues who have already collected the samples., mostly in Germany, Italy and in the Netherlands. Data relative to oak deposits will be extracted from the data base in order to construct isopollen maps for a series of ten-time slices since 15000 BP.

Participants	Results	Research action
P1, P3, P4, P5, P6, P8, P9, P10	R4	Themap can only be constructed if standard techniques are used for the analysis of CpDNA polymorphism. Methods have been developed and tested by partner P1 and will be shared with other participants involved in Task 1 during a 1 week technical meeting. The molecular technique used is a PCR-RFLP assay. For the time being, 28 different haplotypes can be identified from amplification products with 4 primer pairs, digested with 3 restriction enzymes. Scoring of haplotypes and construction of data files will be standardized.

Participant hosting the test and collecting the material	Species	Prov. Test (see 1st column annex1)	Nb of prov	Participant doing the PCR-RFLP assay.
P1	Q.petraea	1	120	P1
P2	Q.petraea Q.robur	2 3	116 68	P3 P3
P3	Q. petraea	4	34	P3
P4	Q.petraea Q.robur	5 6	39 11	P9 P9
P5	Q.robur	7	30	P5
P7	Q.robur	10	23	P9
P7	Q.robur	9	10	P9
P10	Q. petraea	12	12	P10
P10	Q.robur	13	15	P10
P11	Q.petraea+robur	11	24	P8

Country	Maximum number of sample points	Collection of samples done by:	Samples analysed by:
Austria	32	P8	P8
Belgium	12	P1 or P5	P1 or P5
France	168	P1	P1
Germany	135	P3	P3
Great Britain	70 (10*)	P4	P4
Great Britain	2	P9	P9
Italy	80	Italian Subcontractor	P6
Netherlands	12	P5	P5
Spain	146	P10	P10
Portugal	23	P10	P10
Sweden/Norway	20	P11	P8
Switzerland	15	P6	P6

Participant	Results	Research action
P12	R2	The current pollen database comprises data on pollen deposits collected in about 550 points distributed throughout Europe (Mostly France, Great Britain, Spain, Poland ..). Data on 300 additional points will be added through cooperative efforts with colleagues who have already collected the samples., mostly in Germany, Italy and in the Netherlands. Data relative to oak deposits will be extracted from the data base in order to construct isopollen maps for a series of ten-time slices since 15000 BP.