ABSTRACT: Comparative analysis of transcriptome profiles of G. arboreum L. cv. and its fuzzy-lintless mutant (ANOI 1960) during fibre development stages.
Project description:Comparative analysis of transcriptome profiles of G. arboreum L. cv. and its fuzzy-lintless mutant (ANOI 1960) at 0 and 10 dpa. Cotton is one of the most commercially important fibre crops in the world and used as a source for natural textile fibre and cottonseed oil. The fuzzy-lintless ovules of cotton mutants are ideal source for identifying genes involved in fibre development by comparing with fibre bearing ovules of wild-type. To decipher molecular mechanisms involved in fibre cell development, transcriptome analysis has been carried out by comparing G. arboreum cv. (wild-type) with its fuzzy-lintless mutant (ANOI 1960). Fuzzed-lintless mutant line was generated by back cross breeding between FL and Fl (recurrent parent) lines (personal communication by Dr. I. S. Katageri). Basically Fibre less type was a RIL, first recovered from cross between G.arboreum (linted) and G. anomalum (lint less). This RIL was used as donor parent and crossed with normal arboreum (as recurrent parent) to develop G. arboreum FL and G. arboreum Fl isogenic lines. This G. arboreum Fl line is named as ANOI 1960. Cotton bolls were collected at fibre initiation (0 dpa/days post anthesis) and elongation (10 dpa) and gene expression profiles were analyzed in wild-type and ANOI 1960 mutant using Affymetrix cotton GeneChip Genome array.
Project description:Comparative analysis of transcriptome profiles of G. arboreum L. cv. and its fuzzy-lintless mutant (ANOI 1960) at 0 and 10 dpa. Cotton is one of the most commercially important fibre crops in the world and used as a source for natural textile fibre and cottonseed oil. The fuzzy-lintless ovules of cotton mutants are ideal source for identifying genes involved in fibre development by comparing with fibre bearing ovules of wild-type. To decipher molecular mechanisms involved in fibre cell development, transcriptome analysis has been carried out by comparing G. arboreum cv. (wild-type) with its fuzzy-lintless mutant (ANOI 1960). Fuzzed-lintless mutant line was generated by back cross breeding between FL and Fl (recurrent parent) lines (personal communication by Dr. I. S. Katageri). Basically Fibre less type was a RIL, first recovered from cross between G.arboreum (linted) and G. anomalum (lint less). This RIL was used as donor parent and crossed with normal arboreum (as recurrent parent) to develop G. arboreum FL and G. arboreum Fl isogenic lines. This G. arboreum Fl line is named as ANOI 1960. Cotton bolls were collected at fibre initiation (0 dpa/days post anthesis) and elongation (10 dpa) and gene expression profiles were analyzed in wild-type and ANOI 1960 mutant using Affymetrix cotton GeneChip Genome array. Cotton plants were grown under field condition. Flowers were tagged and cotton bolls were collected during fibre development stages. Total RNA was isolated from fibre bearing ovules of wild-type (WT) and fuzzy-lintless ovules of mutant (ANOI 1960) collected at various (0 and 10 dpa) fibre development stages using SpectrumTM Plant Total RNA kit (Sigma, USA) according to the manufacturerM-bM-^@M-^Ys protocol. Affymetrix cotton GeneChip Genome array (Affymetrix, USA) having 23,977 probe sets representing 21,854 cotton transcripts was used for transcriptome analysis. Three biological replicates were maintained to test the reproducibility and quality of the chip hybridization. cDNA labeling, array hybridization, staining and washing procedures were carried out as described in the Affymetrix protocols. CEL files having estimated probe intensity values were analyzed with GeneSpring GX-11.5 software (Agilent Technologies, USA) to get differentially expressed transcripts. The Robust Multiarray Average (RMA) algorithm was used for the back ground correction, quantile normalization and median polished probe set summarization to generate single expression value for each probe set. Normalized expression values were log2-transformed and differential expression analysis was performed using unpaired t-test. The p-values were corrected by applying the false discovery rate (FDR) correction (Benjamini and Hochberg, 2000).
Project description:The aim of the present study is to list the genes involved in cotton (G. arboreum) leaf epicuticular wax production and deposition. For this purpose differentially expressed genes (especially, down-regulated in wax deficient mutant plant) in wild and epicuticular wax mutant (Gawm3) plants were founded through cDNA microarray, developed from the wild plant leaves.
Project description:Small RNAs (21-24 nt) are pivotal regulators of gene expression that guide both transcriptional and post-transcriptional silencing mechanisms in diverse eukaryotes, including most if not all plants. MicroRNAs (miRNAs) and short interfering RNAs (siRNAs) are the two major types, both of which have a demonstrated and important role in plant development, stress responses and pathogen resistance. In this work, we used a deep sequencing approach (Sequencing-By-Synthesis, or SBS) to develop sequence resources of small RNAs from different Gossypium arboreum tissues (leaves, flowers and fibers). The high depth of the resulting datasets enabled us to examine in detail critical small RNA features as size distribution, tissue-specific regulation and sequence conservation between different organs in this species. We also developed database resources and a dedicated website (http://smallrna.udel.edu/) with computational tools for allowing other users to identify new miRNAs or siRNAs involved in specific regulatory pathways, verify the degree of conservation of these sequences in other plant species and map small RNAs on genes or larger regions of the maize genome under study. Small RNA libraries were derived from leaves, flowers and fibers of Gossypium arboreum. Each tissue represented a mixture of developmental stages. Total RNA was isolated using the Plant RNA Purification Reagent (Invitrogen) and submitted to Illumina (Hayward, CA, http://www.illumina.com) for small RNA library construction using approaches described in (Lu et al., 2007) with minor modifications. The small RNA libraries were sequenced with the Sequencing-By-Synthesis (SBS) technology by Illumina. PERL scripts were designed to remove the adapter sequences and determine the abundance of each distinct small RNA. We thank Thea Wilkins for providing the plant material and Kan Nobuta and Gayathri Mahalingam for assistance with the computational methods..