DNA database

DNA databases for macroinvertebrates that can be used for environmental DNA surveys are available

DNA database of macroinvertebrates for Environmental DNA

The Kanagawa Environmental Research Center has long conducted surveys of aquatic insects, which are closely related to river water quality.

To complement these capture surveys, we have been conducting environmental DNA surveys of macroinvertebrates since 2023.

On the other hand, the number of DNA sequences of aquatic insects registered in the Japanese DNA database (DDBJ) is insufficient, which hinders species identification during environmental DNA surveys, so the Center is developing and releasing a DNA database focusing on aquatic insects in the prefecture.

Procedures for DNA database maintenance

DNA database maintenance is conducted in accordance with the following procedures.

(1)Biological sample capture

Insects captured mainly in Kanagawa Prefecture are identified by specialists based on their external morphology in reference to the identification criteria of the Census of Rivers and Waters.

The target species are mainly those commonly found in rivers (common species).

(2)Photographing biological samples

After species identification is completed, morphological and other characteristics of the samples are recorded in photographs before DNA extraction is performed. This is to allow us to confirm morphological characteristics at a later date when the organisms are very small and may be damaged or dissolved during DNA extraction.

Please contact us if you wish to confirm the presence or absence of photographs of the organisms.

Please note that no photographs were taken of biological samples in the early years of database development (fiscal years 2020 to 2021), and therefore, no photographs are available. Please understand this in advance.

(3)DNA extraction from biological samples

DNA extraction from biological samples is now conducted in a manner that does not damage the samples as much as possible.

Please note that in the early years of the database development (FY2020 to FY2021), DNA was extracted from samples using destructive methods to ensure a certain level of DNA concentration in order to sequence the full-length mitochondrial sequence. Therefore, please be aware that there may be cases in which DNA-extracted samples do not remain.

If there are still biological samples of the same species captured at the same location, please check the sample information (Excel file) at the bottom of this page, where it is indicated as "Individuals captured at the same location".

(4)DNA Sequencing

DNA sequencing is performed using DNA extracts. Two methods of sequencing are employed: (1) mitochondrial full-length sequencing by shotgun sequencing and (2) amplicon sequencing by next-generation sequencers.

(5)Organize and check the accuracy of sequenced DNA sequences

For mitochondrial full-length sequence sequences, the AQ-db region of the 16SrRNA region of the mitochondrial DNA region (see the paper at the bottom of this page for details on the region) is extracted.

For sequences obtained from the next-generation sequencer, the primer regions are removed, and the sequence with the highest number of reads for each sample is designated as the sequence for that sample.

After completing the above process, the sequences are checked for errors due to sample mix-ups, contamination, etc. Specifically, the sequences are extracted from the same sequence from the DDBJ. Specifically, the 16SrRNA regions of the same species are collected from DDBJ, and a phylogenetic tree is created to check for the presence of DNA sequences from other species.

Database of 16SrRNA regions of mitochondrial DNA regions

Here is the DNA database of the 16SrRNA region of the mitochondrial DNA region of Insecta, which was maintained by the above procedure. In principle, the regions covered are those of the "AQdb-16S primer" in the following paper.

Takenaka,M.,Yano,K.,Suzuki,T.etal.DevelopmentofnovelPCRprimersetsforDNAbarcodingofaquaticinsects,andthediscoveryofsomecrypticspecies.Limnology24,121–136(2023).https://doi.org/10.1007/s10201-022-00710-5

version	DNA sequence (FASTA format)	Sample Information
ver1.0(April 2023)	DNA sequence (Number of data:551)	Sample Inoformation（Excel：229KB） (420species)
ver1.1(September 2023)	DNA sequence (Number of data:616)	Sample Information（Excel：271KB） (471species)
ver1.2(November 2023)	DNA sequence (Number of data:752)	Sample Information（Excel：318KB） (543species)
ver1.2.1(December 2023)	DNA sequence (Number of data:749)	Sample Information（Excel：318KB） (541species)
Ver1.3(spring 2024)	-	-

〇Introduction of companies, universities, and local environmental research institutes that have contributed to the database development

＜Publication of DNA database＞

Bioengineering Lab. Co., Ltd.

plant bio. Co., Ltd.

(Details of Cooperation)

We have received approval for the release of the DNA database of 454 mitochondrial full-length sequences of insects and the 16SrRNA regions of 405 species and genera, which were obtained through the joint research project "Development of a method for understanding benthic biota using environmental DNA focusing on mitochondrial DNA regions (FY 2020-2021)" between the two companies and our center.

＜Provision of biological samples＞

Tojo Laboratory, Shinshu University

(Contents of cooperation)

We received 132 pieces of DNA extracts of 101 insect species stored in the laboratory.

Shizuoka Institute of Environmental Health Science

(Details of cooperation)

Insects captured in Shizuoka Prefecture were donated.

How to read sample information (explanations have been added for parts that are difficult to understand)

SequenceID: The name is given by the following rule: Order abbreviation - Family abbreviation - Scientific name - Sample name - Analysis method abbreviation.

Biological samples: "X" indicates that no biological samples remain, "〇" indicates that biological samples are stored, and "Same location captured" indicates that no DNA-extracted biological samples remain, but individuals that were captured at the same location and are considered to be the same species are stored.

Comments on each sample: This section summarizes points that were noticed during the database development.

We are still in the process of developing a DNA database for insects, and we are looking for people who are willing to cooperate with us in database development. If you are interested in helping us, please contact us.

If you have any questions about the contents of this DNA database, please contact us. If necessary, we will consider re-sequencing or other measures.

For inquiries about this page, please contact

Kanagawa Environmental Research Center

Phone: 0463-24-3311

Extension: 314

Facsimile: 0463-24-3300