Development of GS-RLP markers for detection of epialleles in cotton (Gossypium L. )

Development of GS-RLP Markers for Detection of Epialleles in Cotton (Gossypium L. ) Ayşe SAĞIR M.Sc. Thesis in the Department of Field Crops Supervisor: Prof. Dr. Mehmet KARACA February 2020; 63 pages As an industrial crop Cotton (Gossypium L.), is the main raw material of many types of the industry, and it is important for the world and Turkey. The rapidly increasing world population and changing and developing life standards of people increase the need for cotton. The word epigenetics, was used in the past to express the differentiation of multicellular organisms during development to form tissues and organs from uniform cells. However, nowadays it is used to describe hereditary changes occur in the expression of genes without alteration in DNA sequences. Among epigenetic regulations, the most studied and understood mechanism is the DNA methylation. Different methods has been used for determining DNA methylation. The aim of this study, loci and primer pairs for detection of epialles by the Gen Specific Restriction Length Polymorphism (GS-RLP) method developed in our labratory and similar to Methylation Sensitive Amplification Polymorphism (MSAP). In the study, DNA sequences of two cotton types, G. hirsutum L. and G. barbadense L. species, constituting more than 98% of the world cotton production was used. The sequences were downloaded from the GenBank database and primer pairs were designed using certain criteria. Palindromic recognition regions of MspI and HpaII isochisomers were present in the developed primer sequences. The differences in DNA sequence can be determined based to the methyl group found on the cytosine nucleotide of DNA in the palindromic region. In the study, 88 primer pairs covering 54 loci was developed. Half of the developed primer pairs was from G. hirsutum and the other half from G. barbadense genomic sequences. It was demonstrated in preliminary experiments that primer pairs can amplify target sequences from the genomes of both species. For this, six loci and spesific primer pairs were selected and utilized in the GS-RLP method. Primer pairs developedand used in preliminary experiments were promising for the detection of epialleles. In this context, the results suggested that these primer pairs can be used for the detection of epialleles related to fiber development. Therefore the developed primer pairs can also be used to developing markers in deterimination of the quality and quantity of fiber in cotton. G. hirsutum and G. barbadense are two cotton species constituting 98% of the world cotton production. Methilation sensistive primer pairsspesific to these species developed in this study is not only used for the development of markers but also useful for understanding the epigenetic arrangements and regulations of genes and gene elements related to fiber properties. KEY WORDS: DNA Methylation, Epigenetics, Epigenetic Arrangements, Fiber Properties, GS-RLP marker, Gossypium barbadense, Gossypium hirsutum, Primer Pair COMMITTEE: Prof. Dr. Mehmet KARACA Prof. Dr. Bayram ÇEVİK Assoc. Prof. Dr. Ayşe Gül İNCE