Project description:Phalaenopsis aprodite subsp. formosana is one of the most important species for Phalaenopsis breeding. A mutant line with variegated leaf is found in this species. The green leaves bear unstable yellow sectors. In order to investigate the molecular mechanism of the variegated mutant line, we sequenced the transcriptome of variegated mutant by Illumina's Solexa sequencing technology. The sequence analysis results showed 22,598 unigenes by de novo assembly method, and the average unigene length was 1,286 bp. The bioinformatics tools were used to screen the differential expression between green and yellow sectors of leaves. There were 389 differentially expressed unigenes were identified. In addition, Gene ontology (GO) and KEGG pathway analyses revealed diverse biological functions and processes from differentially expressed genes. In transcriptome analysis, seven differential expression gene between the green and yellow sectors of leaves can be identified as CHLM, CRD1, POR, CLH, SGR, psbA and Lhcb6 by RNA deep sequencing. The expression of candidate genes was confirmed using semi-quantitative reverse transcription (RT) PCR and real-time RT PCR. The result showed that the significantly differential expression of CLH and SGR between green and yellow sectors was confirmed. It is suggested that the overexpressed SGR gene promotes the function of chlorophyllase, leading to the rapid degradation of chlorophyll in yellow sector. It causes the chlorophyll to not accumulate in the yellow sector, as a result, the variegated leaves are shown.
Project description:The incomplete genome annotation of non-model organisms hampers molecular and proteomic studies. Proteomics informed by transcriptomics (PIT) is suited to non-model organisms because peptides are identified using transcriptomic, not genomic, data. Aedes aegypti is the mosquito vector for the (re-)emerging dengue, chikungunya, yellow fever and Zika viruses. An Ae. aegypti genome sequence is available, however experimental evidence for >90% of the Ae. aegypti proteome or the activity of transposable elements (TEs) that constitute 50% of the Ae. aegypti genome is lacking. We used PIT to characterise the proteome of the Aedes aegypti derived cell line Aag2. Hotspots of incomplete genome annotation were identified which are not explained by poor sequence and assembly quality. We developed criteria for the characterisation of proteomically active TEs and demonstrate that protein expression does not correlate with a TE’s genomic abundance. Finally, we identify Phasi Charoen-like virus as an unrecognised contaminant of Aag2 cells. We therefore present the first proteomic characterisation of mobile genetic elements, and provide proof-of-principle that PIT can evaluate a genome’s annotation to guide annotation efforts.
Project description:An Ac/Ds transposon tagged mutant population was screened for changes in visible fruit phenotypes. One line showed orange, yellow sectors in the fruit and was named Orange ripening (Orr), its transposase free offspring showed Mendelian segregation yielding red, yellow and orange fruit bearing plants in a ratio of 1:2:1. Crossing the an orange fruit plant line to the wild-type yielded only plants bearing yellow fruit. A cross between the yellow fruit bearing progeny yielded 26 plants having red and 17 plants having yellow fruit, suggesting an over-dominant allele. Using inverse PCR analysis showed an insertion in the putative subunit M of the tomato Ndh complex. Subsequently, an Orr Ds transposon excision line was recovered which only showed red pigmented fruit. Here, we describe microarray profiling of tomato fruits from wild-type, heterozygous and homozygous Orr insertion plants and from fruits harvested from the Orr excision line. Keywords: mutant wild type