Services

Plant Isoform-sequencing with Reference Genome

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Introductions

Plant DNA Recombination

The study of full-length transcriptomes is an important way to understand the functions of biological organisms. The third-generation sequencing technology of Pacbio uses single-molecule real-time sequencing technology (SMRT). Due to its ultra-long read length (average 15kb), complete full-length transcripts can be directly obtained without splicing. Therefore, the entire sequence of a high-quality single RNA molecule can be effectively obtained, and the transcripts of isoforms, homologous genes, superfamily genes or allelic expressions that cannot be identified by second-generation sequencing can be accurately identified. At present, full-length transcriptome sequencing technology has been widely used in plant research, including Arabidopsis, corn, wheat, sorghum, salvia, petunia, etc. Isoform-sequencing with reference genome can be used to perfect species genome annotations, study changes in variable splicing events caused by different experimental treatments, study post-transcriptional regulatory mechanisms (such as variable polyadenylation), and more accurate quantitative analysis.

Services

Based on PacBio's third-generation sequencing platform, Lifeasible uses single molecule real-time sequencing technology (SMRT) to provide professional isoform sequencing and analysis services with reference genome to help your research.

Workflow of isoform sequencing and analysis services with reference genome

Plant DNA Recombination

Analysis content

Standard information analysis Raw sequencing data statistics and quality control
  • Reads category
  • Reads clustering and corre ction
  • Full-length isoform data statistics
  • Align with reference genome
  • Novel transcript prediction
  • Novel transcript function annotation
  • 1) Nr annotation
    2) GO function annotation
    3) KEGG metabolic pathway annotation
    4) Swissprot protein annotation

  • Gene structure analvsis
  • 1) INCRNA analysis
    2) Variable shear analysis
    3) Variable polyadenylation analysis
    4) Fusion gene analysis
    5) SNP/indel analysis
    6) Open reading frame analysis
    7) Transeription factor analysis
    8) Optimization of gene structure

Customized information analysis
  • Use the second-generation data to correct the third-generation data (illumina data is required)
  • Gene quantification and differential expression analysis (Illumina data required)
  • Multi-omics association analysis (such as methylation, proteomics, miRNA)

Sample requirements

  • Sample size
  • Total RNA>12μg (2 libraries)

  • Sample purity
  • OD260/280≥1.8, OD260/230≥1.0. And there is a normal peak at 260nm.

  • RNA integrity
  • RIN≥8.0, 28S/18S≥1.3, the baseline of the spectrum is not raised, and the 5S peak is normal.

  • Please put the sample in a 1.5ml tube, indicate the sample name, concentration and preparation time on the tube, and use parafilm to seal the tube mouth.
  • Before shipping, fix all sample tubes in 50ml centrifuge tubes with caps and ship them on dry ice.

Project cycle

  • The entire project generally takes about 2 months.

Feedback to customers

  • Raw data
  • A project report includes experimental process, instrument parameters, analysis methods and results, etc.

Service process

Service Process of Isoform-sequencing with Reference Genome - Lifeasible

Advantages

  • Lifeasible specializes in providing services related to agricultural molecular breeding, with standardized systems and scientific management.
  • PacBio's third-generation sequencer has the advantages of ultra-long read length, high throughput, no GC preference, and direct detection of base modifications, etc.
  • Full-length transcriptome sequencing can detect multiple alternative splicing forms and discover more splicing sites and alternative splicing events. And it can discover new functional genes and supplement genome annotations.
  • The one-stop service we provide makes your upstream and downstream experiments seamlessly connected, facilitates your experiments more conveniently and quickly.
  • Personalized customized package service.

※ It should be noted that our service is only used for research.

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