A Step Towards Cotton Genome Assembly Through Construction Of High Density Genetic Map From Interspecific Cross Of G. Hirsutum×G. Tomentosum

Cotton is the leading natural fiber crop known as "white gold". It is a crop of commerce since long ago. It is also world’s important oil-seed crop. Cotton (Gossypium spp) is extensively grown species across the globe from tropical to subtropical regions. So mapping cotton genome will provide foundation for advanced research. Cotton being a large genome needs high density genetic mapping to progress towards the whole genome assembly. A genetic map estimates the relative position of molecular markers along the chromosome which are corresponding to the genes based on recombination events. Genetic maps not only paved the way in understanding genome organization and structure but also offer the opportunity for tagging superior traits for crop improvement through marker-assisted selection (MAS). Genetic maps developed through PCR-based markers will be more efficient in marker-assisted selection as these markers are environmentally neutral, abundant across the genomes and made it possible to observe the genetic variations at DNA level.It has been proven that upland cotton germplasm has narrow genetic base hence it needs to be enriched by introducing superior genes from wild relatives through introgression. Therefore, it is imperative to have better understanding of genome organization of wild relative. There are five tetraploid cottons including G. tomentosum which has many useful genes like stresses resistance genes that need to be introgressed into the G. hirsutum for crop improvement. Up till now, researchers mostly used G. barbadense as one of the parent to generate interspecific cross to get high levels of polymorphism for the construction of high density genetic maps. This is the main reason why we used G. tomentosum in present study.Firstly, raimondii-derived SSR, form174scaffolds of G. raimondii were designed. A total of449SSR ubiquitous repeats distributed among101scaffolds,428pairs of SSRs were developed. The frequency of these SSR loci was58%with an average one locus per28.7kb. The highest motifs were tri-nucleotides and hexa-nucleotides with frequencies of20.9%and38.3%respectively. The optimum length of a locus was20bp with a range of18-22bp. They revealed16.59%polymorphic rate between CRI-12and G. tomentosum. Out of the35primer pairs,34loci were mapped which increased the gSSR map from1575to1609. This divulged2.11%increase in the gSSR.88.24%were integrated on D sub-genome, we can infer that D sub-genome may have gone through concerted evolutionary process but it needs further verification. Secondly, a high density genetic map was constructed using F2population derived from an interspecific cross of G. hirsutum (the cultivar name as CRI-12) x G. tomentosum. The map was exclusively based on survey of genome wide simple sequence repeat (SSR) form24DNA sources. The map consisted of3,093markers loci distributed across all the26chromosomes and covered4,365.3cM of cotton genome, with an average inter-marker distance of1.48cM. Marker number anchored on the chromosomes varied from65to215with an average of118.96on each chromosome. More markers were mapped on D sub-genome (51.86%) than A sub-genome (48.14%). The maximum length of chromosome was218.38cM and the minimum was122.09cM with an average length of167.90cM. A sub-genome covers more genetic distance (2,189.01cM) with an average distance of1.53cM than D sub-genome which covers a length of2,176.29cM with an average distance of1.43cM. The smallest homeologous chromosome pair was Chr.04(A04) and22(D04), and the largest was Chr.05(A05) and19(D05) as far as mapped loci are concerned. There were716distorted loci in the map accounting for23.14%and more distorted loci were distributed on D sub-genome (25.06%) than A sub-genome (21.23%). In our map49segregation hotspots (SDR) were distributed across the genome with more on D sub-genome as compared to A genome. All the skewed alleles within one SDR segregate in the same direction. Two post-polyploidization reciprocal translocations of "A2/A3and A4/A5" were suggested by7pairs of duplicate loci. The map constructed through these studies is one of the three densest genetic maps in cotton; however, this is the first dense genome wide SSR interspecific genetic map between G. hirsutum and G. tomentosum. This map will play an important role in understanding the genome structure of G. tomentosum. It will also open the doors for further in-depth genome research such as fine mapping, map-based cloning, evolutionary studies, tagging genes of interest from wild relatives, MAS and comparative mapping not only in cotton but also with other species as well.