This application establishes an automated transcriptome library preparation workflow for RNA extracted from insect tissues using the MGI MGISP-960 platform. It combines mRNA enrichment and RNA library preparation reagents with automation consumables to generate libraries for MGI sequencing. The example below uses four locust brain RNA samples and reports the preparation conditions, library concentrations, and fragment profiles.

Prepare RNA from Small Insect Tissue Samples

MolPure™ Micro Total RNA Kit (Cat. No. 19212ES) is an option for extracting total RNA from small samples. It combines a micro-scale DNA removal/RNA-binding spin column with a specialized buffer system for very small amounts of animal cells, microdissected tissue, and easy-to-lyse animal tissue.

RNA extraction takes approximately 8 minutes. The kit processes 1–500,000 cells or less than 5 mg of tissue. Its DNA removal/RNA-binding column and reagent formulation remove residual genomic DNA without DNase treatment. The recovered RNA can be used directly for reverse transcription, quantitative PCR, or high-throughput sequencing library preparation.

What RNA Sequencing Reveals about Insect Biology

Next-generation sequencing (NGS) gives researchers a genome-wide view of gene expression, from broad patterns to specific mechanisms. While the genome provides the genetic blueprint, the transcriptome shows how that information is used as biological activity changes. RNA sequencing (RNA-seq) captures gene expression under defined conditions, helping explain insect growth, physiology, metabolism, and behavior.

Reagent List for the Experiment

Category

Cat.No.

Product name

RNA Library Preparation

12629ES

Hieff NGS™ mRNA Isolation Master Kit V2

12340ES

Hieff NGS™ EvoMax RNA Library Prep Kit (Strand-specific)

Magnetic Beads

12601ES

Hieff NGS™ DNA selection Beads (Superior Ampure XP alternative)

Quantification

12642ES

1× dsDNA HS Assay Kit dsDNA qubit

Adapters

13350ES

Hieff NGS™ Unique Dual Barcode Primer Kit for MGI

User-Supplied Materials

—

Absolute Ethanol

The Automated Workflow

Figure 1. Automated transcriptome library preparation from cell samples on the MGISP-960.

Figure 1. Automated transcriptome library preparation from cell samples on the MGISP-960.

Pre-Experiment Preparation

1. Equilibrate magnetic beads to room temperature before use.

2. Prepare 80% ethanol.

3. Prepare 2 herbal medicine samples and 1 rapeseed sample, each containing 400 ng of DNA. The extracted DNA shows visible pigment contamination.

Experimental Conditions and Results

Sample number

1

2

3

4

Sample type

Brain tissue

Organism

Locust

Nucleic acid concentration

923.4

1882.9

1813.1

1471.1

RNA input

120 ng

RNA enrichment

mRNA capture with magnetic beads

Fragmentation conditions

94°C for 7 min

Cleanup after ligation

0.55×

Size selection

0.60× / 0.15×

PCR amplification

14 cycles

Cleanup after amplification

0.8×

Elution volume

28 µL

Library concentration (ng/µL)

15.6

24.8

36.6

35.6

Review Library Concentrations and Fragment Profiles

Figure. Fragment profiles of the RNA sequencing libraries.

Figure. Fragment profiles of the RNA sequencing libraries.

Yeasen Product Portfolio for Transcriptomic sequencing

Category

Product

Cat. No.

RNA extraction

Hieff™ Magnetic Tissue/cell Total RNA Kit

18605ES

RNA extraction

Hieff™ Flash Cell/Tissue Total RNA Kit

19221ES

RNA extraction

Hieff™ 5-min Flash Cell RNA Kit

19231ES

mRNA enrichment

Hieff NGS™ mRNA Isolation Master Kit V2

12629ES

mRNA enrichment

Hieff NGS™ MaxUp Human rRNA Depletion Kit

12257ES

Transcriptome library Prep

Hieff NGS™ EvoMax RNA Library Prep Kit (dUTP)

12340ES

Hieff NGS™ EvoMax RNA Library Prep Kit (dNTP)

12341ES

MGI adapters

Hieff NGS™ Unique Dual Barcode Primer Kit for MGI

13350-13353ES

Illumina adapters

Hieff NGS™ Stubby UDI Primer Kit for Illumina

12327-12330ES

Purification beads

Hieff NGS™ DNA Selection Beads

12601ES

Quantification

1x dsDNA HS Assay Kit

12642ES

Consumables

250 uL filter tips /
96-well plates /
deep-well plates

83130ES/
83560ES/ 
83672ES

 

Investigação