As the partial fulfillment of M.Sc. in Tropical and International Forestry, I chose the field of Forest Genetics. The idea behind the selection of the topic is to utilise the availability of superb molecular lab and technical professional.
As the genetic has profound impact in every aspect of the forestry, the subject is always fascinating for me. The silivicultural treatments and change of micro environments are important but it is not so big role in changing the productivity and adaptability of the species as it is due to genetical components.
To learn the basic of forest genetics, I worked on the very fascinating tree species of Europe. Tilia species are the native species of Europe, which had faced tremendous pressure due to anthoropological causes and changes of climatic in the Remote Past. At the moment, Tilia species and its hybrids are very popular avenue species in the Europe. I have observed Tilia planted on the urban area of Italy, the Netherlands, Germany and Spain. The situation is similar for England, Turkey, Poland, Czech and many European countries.
Tilia have many species. Among them I studied on Tilia cordata (also called Short Leaved Lime or Winter Lime) and Tilia platyphyllos (Large Leaved Lime or Summer Lime) and their putative hybrid from Hainich National Park situated in Central Germany. I applied molecular methods to identify the species differentiation. Eight isozyme gene loci and 2 chlorplast microsatellites were used. Finally, we were successful to differntiate species among the mixture of these three species without any previous knowledge on species.
This study is one of the first study on the species, so widely acclaimed by the concerned people.
The relevance of the thesis to Nepalese context is seemed only upto the delivering the methods since the species is native to Europe. Further the thesis provides full insights about the importance of forest genetics in species differentiation and such others. We have also the such problems in Nepal in many other species. Upto my knowledge (which might be wrong also), we are taking Sal forest of Terai and hill as one species however they are totally different in morphological point of view. Is it only due to environmental influences or due to the genetic constituents? Are they same species of Shorea robusta or they are different species or subspecies of Shorea, needs to be studied.
There such cases in many genus such as Ficus and others. May be my thesis would provide some insights to those who want to study these differences.
Abstract: There are around 50 genera under the Tiliaceae family. Tilia is one out of them. Genus Tilia includes 35 to 50 different species. The species are distributed in Europe, America and Asia.
Among these species, the study concentrates only in three species of Tilia such as T. cordata, T. platyphyllos and T.× europaea. These species are mainly distributed all over Europe. T. cordata is widely distributed in Europe whereas T. platyphyllos has the distribution range in South and Central Europe. The distribution of T. platyphyllos is quite limited in comparison to T. cordata. T. × europaea are distributed naturally only in those areas where both of T. cordata and T. platyphyllos grow sympatrically.
Tilia species are considered as European noble tree species. They are generally grown in association with other species but they never constitute pure stands in larger areas. The species are important from the ecological, cultural and environmental perspectives; however they are neglected from the economic utilization.
T. cordata, T. platyphyllos and T.× europaea are similar in their outer appearance; however they are different in many morphological traits. Earlier studies about these species tried to distinguish the species based on morphological traits. Many morphological traits are found different among these species. The mosaic of characteristics of leaves, inflorescences, fruits and pollen has been applied in identifying the species. But the precise identification of the species in Tilia is difficult and misleading on the basis of morphological traits alone.
The morphological traits are highly influenced with local environment. The same materials collected from different parts of the tree and in different days might have different characteristics which are very difficult to compare and to take the decision about species identification. Besides, during the winter, it is almost impossible to get these materials for the identification purposes.
To overcome these problems and correct identification of the species in Tilia, the genetic approach can be rightly applied. In these contexts, we tried to distinguish the species with genetic analysis for 294 out of 332 Tilia trees in Hainich National Park, Western Thuringia, Germany. Besides, we tried to access the genetic patterns of these Tilia trees. The samples were collected from 3 plots established by Albrecht von Haller Institute of Plant Sciences at Hainich National park.
To identify precisely the species in Tilia, the genetic markers such as isozyme and chloroplast microsatellite were applied. For the isozyme analysis, overall 8 gene loci from 7 enzyme system were studied under three different running systems. GOT and PGI enzymes from Ashton and Braden, LAP, MNR and SKDH from Histidine-Citrate and PGM and ADH from Tris-Citrate were applied. Among these three running systems, Histidine-Citrate was most efficient to analyse the species. MNR and LAP were vital to discriminate the species in Tilia since MNR and LAP used to produce non overlapping species specific alleles for T. cordata and T. platyphyllos. Additionally PGI, PGM and SKDH also used to produce species specific variants for T. platyphyllos only. MNR and LAP gene loci mainly considered while determining the species.
The analysis of cpDNA was held to get additional information regarding the intra and inter species differentiation. Polymerase Chain Reaction was held in Peltier Thermocycler using ccmp3 and ccmp10. The PCR products were scanned in ABI Genetic Analyser 3100. With the gene scanning and genotyping, five different haplotypes were formed by combining linked size fragments obtained from ccmp10 and ccmp3. The ccmp10 primer produced 3 alleles and ccmp3 produced 4 alleles in the expected size ranges.
The haplotypes 1 and 2 were observed only with Summer Linden and hybrid Linden trees. These haplotypes confirmed the genetic contribution from SL tree. On the other side, haplotypes 3, 4 and 5 were recorded mainly from Winter Linden and Hybrids. Both directional pollen flows was occurred during hybridization. Both T. cordata and T. platyphyllos were contributing pollens to cross fertilisation to produce hybrids. The fragment size 128 bp and 129 bp could be taken as an allele at least contributed from the WL parent. In contrast to this logic, 1 tree from haplotype 3 and 2 from haplotype 4 were identified as SL trees. These might be due to some externalities or due to the occurrence of back crossing for longer period.
Precise identification of species could be made on total 294 trees out of 332 samples. Among the identified species, 255 were Winter Linden trees, 27 were Hybrid and 12 were SL trees. Out of 27 hybrid Linden trees, 16 were identified as the F2 generation hybrid Linden trees.
The genetic identification of the Linden trees did not match perfectly with morphological identification. In morphological identification, there was the larger number of SL and hybrids than genetically identified. The morphological traits are continuous, overlapping and highly influenced by locality factors where as genetic traits are discriminating, non-overlapping and not effected by environment. Morphological traits are continuous traits, therefore getting the cut point is always difficult while identifying species, but genetic traits are discrete hence undoubtedly identified the species on the basis of availability or absence of certain genetic trait or genotype.
Hybrid Lindens are the most genetically diversified (He=0.473) species among three analysed species of Tilia. It is taken as normal since it acquires genetic contribution from both Tilia spp. The gene diversity is higher in T. platyphyllos (He=0.411) than T. cordata (He=0.28). There is minimal or negative Fixation index (F) in Tilia species at Hainich, which describes it as the predominantly out crossing species.
The SL trees are very rare in the region. Therefore, the genetic conservation of this species should be initiated immediately.
(Your suggestions, comments and feedbacks are highly appreciated)
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