Agricultural Research Management Information System

Home
Research Summary
Search
About Us
- ARMIS
- Brochure
Contact Us
Report
User Request
Data Input
Help
- Operation Manual
  - PDF
  - Video
- Program Area & Commodity
We have reached 37600 number of research entries at this moment.

- Logout

Research Detail

Home
Research
Detail

Identification and Expression Pattern Analysis of Chemosensory Receptor Genes in the Macrocentrus Cingulum (hymenoptera: Braconidae) Antennae

TOFAEL AHMED
Bangladesh Sugarcane Research Institute, Ishurdi-6620, Pabna, Bangladesh

TIAN-TAO ZHANG
State Key Laboratory for the Biology of the Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, China

Abstract/ Executive Summary:

Macrocentrus cingulum is an important polyembryonic endoparasitic wasp that invasions larvae of the Asian corn borer, Ostrinia furnacalis (Guenée) and the European corn borer, O. nubilalis (Hübner). Parasitoids use antennae as the main sensory organ to recognize herbivore-induced plant volatiles as host searching cues. The antennal olfaction proteins, odorant receptors (ORs) and ionotropic receptors (IRs) are involved in olfactory signal transduction pathway as a sensory neuron response. In the present study, we constructed a cDNA library from the male and female antennae for identifying the olfaction-related genes in M. cingulum. For that, we sequenced 3160 unique gene sequences and annotated them with gene ontology (GO), cluster of orthologous groups of proteins (COG), and KEGG ontology (KO). Through the homology search, we identified 9 odorant receptors (ORs), 3 ionotropic receptors (IRs) and 1 odorant-binding protein (OBP) genes from the cDNA library sequences. Additionally, the expression patterns of these ORs and IRs in different tissues (antennae, heads, thoraxes, abdomens, and legs) were demonstrated by RT-PCR. The qualitative gene expression analyses showed that most of the OR genes were more highly expressed in female than male antennae; whereas IRs, unlike ORs, were more expressed in various male than females tissues. We are the first to report ORs and IRs in M. cingulum, which should help in deciphering the molecular basis of olfaction system in this wasp.

Keywords: Hymenoptera, Braconidae, Macrocentrus cingulum, cDNA library, Odorant, Ionotropic receptors, Expression pattern

Location: In Bangladesh

Start Date:

End Date:

Program Area: Conservation and Biodiversity

Commodity: Diseases and insects

Objectives:

To identify olfaction-related genes and study olfactory signal transduction mechanisms.

Methodology:

Specimens of M. cingulum were obtained from O. furnacalis larvae living on corn plants at the Langfang Experiment Station of the Institute of Plant Protection, Chinese Academy of Agricultural Sciences, China. The parasitoids emerged as mature larvae from the host larvae and pupated inside the silken cocoon. Adult parasitoid wasps were fed with 20% honey solution. A laboratory colony was maintained on host larvae of O. furnacalis that were reared on an artifi cial diet as described by Zhou et al. (1980). They were maintained in a room at 25°C with a 16L : 8D regime (Ahmed et al., 2013). Antennae, heads with maxillary palps (excluding antennae), legs, thorax, and abdomen of female and male individuals were dissected 1–3 days after eclosion and immediately frozen in liquid nitrogen, then stored at –80°C until RNA extraction. The frozen antennae (100 pairs of each sex) or other tissues were homogenized with a liquid nitrogen-cooled mortar and ground with a pestle into very fine dust. Homogenized tissues were covered with 1 mL of TriZol reagent (Invitrogen, Carlsbad, CA, USA) and total RNA were extracted following the manufacturer’s instructions. After extraction, RNA integrity was verified by 1% agarose gel electrophoresis and quantity was assessed with a Nanodrop ND-1000 spectrophotometer (NanoDrop products, Wilmington, DE, USA). For the synthesis of the first-strand cDNA we used 1 μg of total RNA. The antennal cDNA library was constructed using the In Fusion® SMARTer™ Directional cDNA Library Construction Kit (Clontech, Mountain, CA, USA), according to manufacturer’s instructions. Sizes of cDNA inserts were determined using PCR. A pair of sequence-specific primers was designed based on the sequence of the pDNR-LIB plasmid. The clones that contained the cDNA insert longer than 400 bp were chosen and sequenced using ABI3730 sequencer (SANGON Sequencing Service, Shanghai, China). To explore the expression of the ORs/IRs identified from the antennal cDNA library analyses and to compare the differential expression pattern between the sexes, RT-PCR was conducted with cDNAs prepared from different tissues of male and female wasps.

Source of Information: EUROPEAN JOURNAL OF ENTOMOLOGY, 113: 76–83, 2016; ISSN (online): 1802-8829

Article Link (Online Journal): doi: 10.14411/eje.2016.009

Funding Source:

Budget:

Total Budget (Tk.) :

Achievement/Findings:

The antennal cDNA library was constructed successfully from female and male M. cingulum. Titer of this library was 1.6 × 106 pfu/mL; indicating a sufficient number of genes that were expressed. Genes, longer than ~0.4 kb were selected and sequenced from the library, which resulted in 3160 high-quality unigenes for further analysis. Most of the unigenes ranged from 1000 bp to 1400 bp with a mean length of 1203 bp. To annotate the M. cingulum antennal unigenes, distinct sequences were searched by BLASTx against the NCBI non-redundant protein database (Nr) with a cut-off E-value of 10–5. Of the total 3160 unigenes, 77.00% matched with known gene sequences; the other 23.00% were not similar to any sequence in the Nr database information at present. The top BLAST hit results for each unique sequence.

Knowledge Management: Journal