Data Mining for Simple Sequence Repeats in Oil Palm Expressed Sequence Tags

Expressed Sequence Tags or ESTs are small pieces of DNA sequence that are generated by sequencing either one or both ends of an expressed gene. ESTs provide researchers with a quick and inexpensive route for discovering new genes, for obtaining data on gene expression and regulation, and for constructing genome maps. Oil palm EST sequences as available in public domain are downloaded. They were grouped and made contigs using CAP3 and Phrap. Microsatellite repeats are located using 5 softwares (MISA, TRA, TROLL, SSRIT, SSR primer). Among the 5 methods MISA is found to be the best. It can elucidate the compound repeat also. Frequency and total number (202) of SSR were detected. Mononucleotide repeat is more abundant especially ‘A/T’ repeats in Oil palm. Flanking primers were designed using primer3, SSR primers. The results of the study are given as an online database ‘MEMCO’ to help Oil palm researchers

Interspecific differences in single nucleotide polymorphisms (SNPs) and indels in expressed sequence tag libraries of oil palm _Elaeis guineensis_ and _E. oleifera_

Oil palm is the second largest source of edible oil, which meets one-fifth of global demands of oils and fats. Expressed sequence tag (EST) sequencing programs have provided a wealth of information, identifying novel genes from a broad range of organisms and providing an indication of gene expression level in particular tissues. It also provides the richest source of biologically useful SNPs due to the relatively high redundancy of gene sequence, the diversity of genotypes represented within databases. EST based SNPs are potential molecular markers and aid in genetic improvement. A total of 21062 and 2053 polymorphic (SNP and Indel) sites in _E. guineensis_ species and in _E. oleifera_, 4955 SNPs and 1172 Indels were detected. SNP(17.5/kbp) and Indel(4.1/kbp) frequency was higher in _E. oleifera_ than _E. guineensis_ species (16.8/kbp, 1.6/kbp). _E. oleifera_ showed higher transition to transversion ratio (1.40) than in _E. guineensis_ (1.02). The ratio of Ts vs Tv showed, the genetic divergence is occurring in this crops in different fashion and _E. guineensis_ had diverged more than _E. oleifera_. We provide the results of the study as online database ("http://riju.byethost31.com/oilpalm/":http://riju.byethost31.com/oilpalm/) for use by oil palm breeders

In Silico RAPD Priming Sites in Expressed Sequences and iSCAR Markers for Oil Palm

RAPD is a simple dominant marker system widely used in biology. Effectiveness of RAPD can be improved by selecting and redesigning primers whose priming sites occur in target sequence(s) of gene or organism at optimum distance. We developed software that uses sequences of random decamer primers and nucleotide sequence(s) as two input files. It locates the priming sites in input sequences and generates output files listing frequency and distance between priming sites. When the priming sites of a single primer occur more than once in a sequence with a distance of 200 to 2000 bp, the software also designs pairs of iSCAR primers. An input of 387 RAPD primers and 42,432 expressed sequences of oil palm are used as test. Wet-lab PCR results from a publication that used the same set of primers were compared with software output on priming sites. In the test sequences of oil palm covering 1.4% of genome, we found that at least 60% the primers chosen using software are sure of giving PCR amplification. We designed 641 iSCAR primers suitable for amplification of oil palm DNA. The software successfully predicted 92% (67 out of 73) of published polymorphic RAPD primers in oil palm

Mining for single nucleotide polymorphisms and insertions / deletions in expressed sequence tag libraries of oil palm

The oil palm is a tropical oil bearing tree. Recently EST-derived SNPs and SSRs are a free by-product of the currently expanding EST (Expressed Sequence Tag) data bases. The development of high-throughput methods for the detection of SNPs (Single Nucleotide Polymorphism) and small indels (insertion / deletion) has led to a revolution in their use as molecular markers. Available (5452) Oil palm EST sequences were mined from dbEST of NCBI. CAP3 program was used to assemble EST sequences into contigs. Candidate SNPs and Indel polymorphisms were detected using the perl script auto_snip version 1.0 which has used 576 ESTs for detecting SNPs and Indel sites. We found 1180 SNP sites and 137 indel polymorphisms with frequency 1.36 SNPs / 100 bp. Among the six tissues from which the EST libraries had been generated, mesocarp had high frequency of 2.91 SNPs and indels per 100 bp whereas the zygotic embryos had lowest frequency of 0.15 per 100 bp. We also used the Shannon index to analyze the proportion of ten possible types of SNP/indels. ESTs from tissues of normal apex showed highest values of Shannon index (0.60) whereas abnormal apex had least value (0.02). The present report deals the use of Shannon index for comparing SNP/ indel frequencies mined from ESTlibraries and also confirm that the frequency of SNP occurrence in oil palm to use them as markers for genetic studies

Microsatellites in palm (Arecaceae) sequences

Microsatellites are the most promising co-dominant markers, widely distributed throughout the genome. Identification of these repeating genomic subsets is a tedious and iterative process making computational approaches highly useful for solving this biological problem. Here 38,083 microsatellites were localized in palm sequences. A total of 2, 97,023 sequences retrieved from public domains were used for this study. The sequences were unstained using the tool Seqclean and consequently clustered using CAP3. SSRs are located in the sequences using the microsatellite search tool, MISA. Repeats were detected in 33,309 sequences and more than one SSR had appeared in 3,943 sequences. In the present study, dinucleotide repeats (49%) were found to be more abundant followed by mononucleotide (30%) and trinucleotide (19%). Also among the dinucleotides, AG/GA/TC/CT motifs (55.8%) are predominantly repeating within the palm sequences. Thus in future this study will lead to the development of specific algorithm for mining SSRs exclusively for palms

Interspecific Analysis for SNPs in EST Libraries of Oil Palm- Elaeis Guineensis and E. Oleifera

Oil palm is the second largest source of edible oil, which contributes to approximately 20% of the world's production of oils and fats. Expressed sequence tag (EST) sequencing programs have provided a wealth of information, identifying novel genes from a broad range of organisms and providing an indication of gene expression level in particular tissues. It also provides the richest source of biologically useful SNPs due to the relatively high redundancy of gene sequence, the diversity of genotypes represented within databases, and the fact that each SNP would be associated with an expressed gene. There are cultivated two species of oil palm, Elaeis guineensis Jacq. from tropical western Africa and E. oleifera from Americas. Interspecific SNP detection carried but by choosing Lambda Zap II libraries of both species. Auto_snp perl script was used to cluster the ESTs and detect SNPs. A total of 163 EST sequences belong to Lambda zap libraries analysed here, interestingly we have found 60 SNP site (35 Transition transversion 25 ) in this species specific sequences. Labda Zap tissue also shows the similar pattern of SNPs, such as transition type mutation is higher than trasnversion type in Oil palm (Riju et al, 2007)