| The genome is the entirety of an organism’s hereditary information. Genomics is adiscipline in genetics to study the nature of life, which carried out the research on the genomestructure, gene function, and expression regulation network and metabolism pathway of anorganism at the genomic level. Because of their large and complex genomes, the genomicstudy of plants is difficult and challenging. The advent and development of high-throughputsequencing technology (next generation sequencing technology) provided a new opportunityfor the genomic research of the non-model plants. In this study, we carried out the preliminarygenomics research on the wheat chromosome arm7DL and the transcriptome and chloroplastgenome of Ageratina adenophora using the Illumina\Solexa sequencing technology, with thepurpose to obtain huge sequence resource and reveal their genetic characteristics, and thenestablish a platform for their genomic study. The main results are followed:(1) Survey sequencing and analysis the wheat chromosome arm7DLDNA was firstly isolated from the7DL chromosome arm obtained from flow cytometricsorting of telomeric, and then was used to construct the Solexa sequencing library andsequenced on Hiseq2000platform. A total of14.54Gb data were generated, which coveredthe7DL42fold. After de novo assembled by velvet software,160,021contigs were obtainedwith the total base of223Mb, covering65%of the chromosome arm. Then, we successfullypredict2,893genes based on the assembled contigs. Using the bioinformatics method, wepredicted149miRNA precursors coding20miRNA families, five of which were randomlyselected to validat by Real-time PCR and result indicated that all of them were expressed inwheat. Furthermore,120,262transposon elements and16,325SSR were found in7DL surveysequences, accounting for84.8%. Finally,33SSR markers were developed using the foundSSR motifs and then were validated among20wheat cultivars and two nulli-tetrasomic stocks.Results found that18(70%) markers showed polymorphic and all of them are7DL-specific,which provide a new way to develop specific markers at chromosomal level.(2) De novo sequencing and analysis of the leaf transcriptome of A.adenophora byRNA-seq. In this study, we use this method to accomplish the de novo sequencing and analysis theleave transcritptome of crofton weed. RNA-Seq generated4G raw data, which was then denovo assembled into127,189unigenes with the aveage length of702bp. Of all the unigenes,53,886(42.37%) had significant similarity with proteins in the NCBI non-redundant proteindatabase. Furthermore,41,481unigenes were assigned into65GO functional classifications,and17,498unigenes could be assigned into25COG functional categories. The functions ofthese annotated unigenes were closely related to cell component organization and biogensis,signal transduction and cellular metabolism. KEGG pathway analysis showed that18,306unigene were assigned into297pathways, involved in signal transduction, materialmetabolism and second metabolism biosynthesis. According to the RPKM differentialscreening,9,495differential expression unigenes(︱log2Ratio︱≥1and FDR≤0.001) werefound, which may play an important role in rapid invasion and adaption of crofton weed.Finally, GO functional enrichment and pathway enrichment analysis found that thesedifferential genes are mainly related to secondary metabolism、biosynthesis and signaltransduction. Our present study provided the first comprehensive transcriptome informationabout the leaf of the major invasive weed, crofton weed.(3) The complete chloroplast genome sequences of A. adenophora and comparativegenomic analysis of six Astereace plastomes.We sequenced the chloroplast genome of crofton weed based on Illumina sequencing.Resulted showed that its chloroplast genome is150,689bp in length, including a smallsingle-copy (SSC) region of18,358bp and a large single-copy (LSC) region of84,815bpseparated by a pair of inverted repeats (IRs) of23,755bp. The genome contains130uniquegenes, including86protein-coding genes,36tRNAs and8RNA, of which17genes containedintrons. In addition,31tandem repeats and28dispersed repeats were also found in thisgenome, which would be useful for marker development. This is the first sequencedchloroplast genome in Eupatorieae tribe. Furthermore, we compared the gene content,sequence similarity and genome structure of this chloroplast genome with other five astereacechloroplast genomes and five DNA regions (ndhD-ccsA, psbItrnS, ndhF-ycf1, ndhI-ndhG andatpA-trnR) containing parsimony-informative characters higher than2%, which may bepotential informative markers for barcoding and phylogenetic analysis. Finally, phylogeneticanalysis based on35genes of33species demonstrated a sister relationship between A.adenophora and G. abyssinica and supported a monophyly of the Asterales.(4) Development and characterization of SSR markers in A.adenophora using thehigh-throughput sequencing technology. In this chapter, we sequenced the genome of crofton weed using the Illumina GAIIplatform with the2fold coverage. After assembly using SOAPdenovo,58,432contigs withthe length no less than200bp were obtained, which comprises49Mb of assembled data. Then,these assembled contigs were used to identify the repeats motifs. A total of3,012microsatellites were identified, including mono-, di-, tri-, tetra-, penta-and hexa-nucleotiderepeat motifs. Among them, the di-nucleotide repeat motifs were the most abundant type ofrepeat motifs. Thirty primer pairs were developed according to the identified SSR loci andthen were validated in24individuals of crofton weed. Results indicated that fifteen locidisplayed polymorphism with the number of alleles per locus ranging from two to five, withthe average of3, values for observed and expected heterozygosities ranging from0to1andfrom0.08to0.63, with the averages of0.44and0.37, respectively. The PIC of thesepolymorphic markers ranged from0.08to0.59, with the average of0.30. These novelmicrosatellites provide a valuable tool for population genetics and molecular ecological studyof this invasive weed. |
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