COI Gene Sequencing

COI Gene Sequencing

Our COI sequencing services provides complete solution (DNA extraction, PCR amplification, Gel verification, Purification, Sequencing, Sequence assembling, Bioinformatics analysis (BLAST in Various Databases) and Construction of Phylogenetic tree. We also accept DNA samples, PCR products (Purified/Unpurified) for Sequencing. According to the samples charges may vary. For better result submit direct sample (insect, Tissue etc.,) stored in 90% ethonol.


Amplification of the COI gene:

The extracted DNA is used as the template for PCR to amplify the COI gene sequence. Universal primers (LCO1490 / HCO2198) or specific primers complementary to a conserved region are used so that COI region can be amplified. The PCR amplification produces multiple copies of the target DNA sequence. The resulting product (~650bp) is used as the template for the sequencing.

DNA sequencing:

Since we know the length and terminal base of each fragment, the sequences of the bases can be determined.

DNA sequencing can be done by the common Sanger sequencing method.

We provide below contents documents as a COI Result:

1. Sequence raw data (Forward and Reverse) in different file format.

  • .ab1 - Sequence with chromatogram
  • .pdf - Sequence with Chromatogram
  • .txt - Only sequence
  • .fasta - Only Sequence

2. Assembled Sequence (Contig) - ~ 650bp

3. Blast result generated from All GenBank + NCBI+EMBL+DDBJ+PDB sequences.

4. Summary report comprises:

  1. Methodology for DNA Extraction, PCR, Sequencing and Bioinformatic analysis.
  2. Sequence analysis report contains sequence read Normal Length; Q16 & Q20 read length, GC Content details.
  3. Proper contigs sequence with coverage image
  4. Final Result
  5. Phylogeny Tree.

Importance of Cytochrome c Oxidase subunit I gene (COI) study:

Cytochrome c Oxidase subunit I gene (COI), the gold standard in animal, insect DNA barcoding. Size of the Gene is 648-bp. This  tool is used in the identification of species.

Taxonomy is an integral part of any biological study. Scientific studies involving a taxonomy must have an accurate identification.  Morphology alone sometimes fails as an effective identifier of species. In cases of morphologically similar species, or where the specimens are derived from larval or juvenile life stages, the usefulness of traditional methods of comparative morphology could be limited. It is heavily dependent on specialists.

COI study is most widely used markers for population genetic and phylogeographical studies across the animal kingdom. COI is one of the three mitochondrial DNA encoded subunits of the respiratory complexIV, which is the third and final enzyme of the electron transport chain of mitochondrial oxidative phosphorylation. Its utility is further increased when showed that a  ~650bp fragment of the COI gene could be an efficient identification tool for many species. The diversity in the amino acid sequences coded by the gene was sufficient to place species reliably into higher taxonomic classifications (from phyla to orders). In addition, they observed that diversity in nucleotide sequences of the same gene region allowed for the discrimination of closely related lepidopteran and avian species. Therefore, it was proposed that a DNA barcoding system for most, if not all, animal life could be based upon sequence diversity in COI.

Identification is done by matching the barcode profile with those found in the Gene database. It is the most frequently used tools for computing sequence similarity is the Basic Local Alignment Search Tool, or BLAST, found in the National Center for Biotechnology Information (NCBI) website and other Gene bank. After submitting the query, the sequence will be fed into the algorithm on the BLAST server and locally aligned to the similar sequences in the database. This alignment will be translated into scores that signify similarity of sequences. A specimen is identified if that sequence matches one in the barcode library. Otherwise, the sequence could either be a novel barcode sequence for a given species, usually as a geographical variant or new haplotype, or it belonged to an unknown species.