Insect Genomics in the 21st Century: Prospects and Progress

Markkandan Kesavan 한국응용곤충학회 2018 한국응용곤충학회 학술대회논문집 Vol.2018 No.04

Insects are the dominant animals in the world, with more than one million described species. Insects, not only produce direct damage to plants but also acts as a vector for various pathogens. In recent years, next-generation sequencing (NGS) techniques have provided fascinating opportunities to understand the basic biology, biochemistry, and molecular biology of these intimate and intriguing relationships. The decrease in sequencing costs and extensive sequencing services from NGS providers has brought many scientists to be involved in genome sequencing of insects and their associated entomopathogens. By using high-throughput genomic technologies, scientists can elucidate the virulence, host adaptation and gene function of the particular entomopathogen including virus, fungi, bacteria and nematode.

Overexpression of phytochrome A and its hyperactive mutant improves shade tolerance and turf quality in creeping bentgrass and zoysiagrass

Ganesan, Markkandan,Han, Yun-Jeong,Bae, Tae-Woong,Hwang, Ok-Jin,Chandrasekkhar, Thummala,Shin, Ah-Young,Goh, Chang-Hyo,Nishiguchi, Satoshi,Song, In-Ja,Lee, Hyo-Yeon,Kim, Jeong-Il,Song, Pill-Soon Springer-Verlag 2012 Planta Vol.236 No.4

<P>Phytochrome A (phyA) in higher plants is known to function as a far-red/shade light-sensing photoreceptor in suppressing shade avoidance responses (SARs) to shade stress. In this paper, the Avena PHYA gene was introduced into creeping bentgrass (Agrostis stolonifera L.) and zoysiagrass (Zoysia japonica Steud.) to improve turf quality by suppressing the SARs. In addition to wild-type PHYA, a hyperactive mutant gene (S599A-PHYA), in which a phosphorylation site involved in light-signal attenuation was removed, was also transformed into the turfgrasses. Phenotypic traits of the transgenic plants were compared to assess the suppression of SARs under a simulated shade condition and outdoor field conditions after three growth seasons. Under the shade condition, the S599A-PhyA transgenic creeping bentgrass plants showed shade avoidance-suppressing phenotypes with a 45?% shorter leaf lengths, 24?% shorter internode lengths, and twofold increases in chlorophyll concentrations when compared with control plants. Transgenic zoysiagrass plants overexpressing S599A-PHYA also showed shade-tolerant phenotypes under the shade condition with reductions in leaf length (15?%), internode length (30?%), leaf length/width ratio (19?%) and leaf area (22?%), as well as increases in chlorophyll contents (19?%) and runner lengths (30?%) compared to control plants. The phenotypes of transgenic zoysiagrass were also investigated in dense field habitats, and the transgenic turfgrass exhibited shade-tolerant phenotypes similar to those observed under laboratory shade conditions. Therefore, the present study suggests that the hyperactive phyA is effective for the development of shade-tolerant plants, and that the shade tolerance nature is sustained under field conditions.</P>

Proteomic Investigation of Seed Storage Proteins Among Korean Soybean Cultivars

Kesavan Markkandan,Bong Soo Park,Soon Ho Choi1,Hak Soo Seo 한국작물학회 2010 한국작물학회 학술발표대회 논문집 Vol.2010 No.04

and distribution of seed storage proteins are responsible for the quality of soybean and seed development. Among storage proteins, lipoxygenase isoforms (Mw. ~97 kDa) play a major role in the distinct bean flavor during storage. In this study, we compared three soybean elite cultivars viz., JIMPUM, JINPUM2 and TANMI2 (lipoxygenase null mutants, originated from Japan) along with WILLIAMS 82 (control plant, USA) to determine the seed storage proteins by proteomic approach. Phenotype of the mature seeds showed the variation in seed coat, color and appearance. Total seed proteins of the above cultivars were subjected to two dimensional gel electrophoresis (2-DE). The resulted protein profiling showed the intensity of the different quantitative spots varied among the four cultivars. We are now investigating by using other proteomic tool and the resulted difference in proteins may helpful in quality improvement or the functional roles in the seed development.

Seed size: a priority trait in cereal crops

Kesavan, Markkandan,Song, Jong Tae,Seo, Hak Soo Blackwell Publishing Ltd 2013 Physiologia Plantarum Vol.147 No.2

<P>Crop production and productivity must be increased to provide a balanced diet for the global population. The entire genome sequences of crop species allow the elucidation of genes that regulate important traits related to the final crop seed yield, which frequently depends mainly on seed size. Seed size is a major factor that controls seed quantity and it is strongly affected by various biotic, abiotic and genetic factors. Epigenetic marks in the genome and phytohormones are also important factors affecting seed growth and development. Several genes are known to be involved in the control of seed size, but their interaction and functional characterization have yet to be resolved. In this review, we discuss the different factors that govern seed size in cereal crops and <I>Arabidopsis</I>.</P>

Profiling and identification of pregnancy-associated circulating microRNAs in dairy cattle

Kesavan Markkandan,안궁,이동진,김태일,당창권,홍송의,윤호백,임현주,홍창표 한국유전학회 2018 Genes & Genomics Vol.40 No.10

Holstein is one among the dairy cattle which provide higher milk yields than most other cattle breeds. Lack of high-accuracy, reliable methods for early detection of cattle pregnancy reduces overall productivity and constitutes a high economic burden to the dairy industry. The circulating microRNAs (miRNAs) in exosomes could provide information and serve as potential biomarkers for livestock health and disease. However, the complexity of miRNA in response to cattle early pregnancy remains unknown. Hence, we collected blood samples of three healthy dairy cows of normal and 30 days of pregnancy, in order to further characterize the miRNA transcriptome profile. A high-throughput RNA-Seq approach detected 794 known and 2154 novel circulating miRNAs in six libraries. A total of 29 miRNAs in the 30 days of pregnancy group showed significant differences compared to the normal group. Further, bta-miR-450b, bta-miR-146b, bta-miR-26b and bta-miR-27b were upregulated which shown to be involved in preeclampsia, immune response and mammary gland development. GO enrichment analysis showed these target genes were involved in the metabolic process, signal transducer activity, and membrane etc., while KEGG analysis showed that these genes were enriched in membrane trafficking, chromosome and associated proteins, exosome and G protein-coupled receptors pathways. These results provide an experimental basis to reveal the potential role of miRNAs as biomarkers in early diagnosis of pregnancy and other molecular functions.

Analysis of chigger mite genome and its application

Sathiyamoorthy Subramaniyam,Kesavan Markkandan,Myunghee Jung,Seungil Yoo,Junhyung Park,JongYul Roh,E-hyun Shin,Mi-Yeoun Park,Young Ran Ju,Ju Hyeon Kim,Deok Ho Kwon,Kyungjae A. Yoo,Si Hyeock Lee 한국응용곤충학회 2015 한국응용곤충학회 학술대회논문집 Vol.2015 No.10

Leptotrombidium pallidum is the major vector mites for Orientia tsutsugamushi, the causative agent of scrub typhus. To understand the molecular mechanism of L. pallidum, we sequenced the whole genome using Illumina sequencing technology. Totally four genomic libraries with different insert sizes ranging from 280 bp to 8 kb were used to generate 45.1 Gb of genome in the combination of paired-end and mate-pairs sequencing reads. Quality filtering and correction of paired-end reads for very small and/or bad-quality sequences yielded 26.9 Gb of high-quality sequences, which are used to estimate the genome size as 175 Mbusing kmer methods and assembled into a 193.7 Mb genomic sequence scaffolds with N50 length of 92,945 bp. Furthermore, 94% of CEGMA completeness score were obtained from genome scaffold assembly. To facilitate gene annotation, we used a combination of de novo and homology based tools to predict gene models in the chigger mite genome. A combination of evidence-based and de novo approaches predicted 15,842 high-confidence protein-coding genes with an average transcript length of 1,511 bp and 2.4 exons per gene which corresponds to about 12.4% total gene length. Bacterial endosymbiosis are very common in mite species and can range from mutualistic to pathogenic associations. Henceforth, the endosymbionts in L. pallidum were predicted using the NCBI microbial draft genomes and mitochondrial genome. Besides, this L. pallidum draft genome can be used as a significant reference for comparative genomic studies across mite species.

Current trends and future prospects in insect genomics

Junhyung Park,Kesavan Markkandan,Seong-Eui Hong,Sathiyamoorthy Subramaniyam,Seungil Yoo,Hyung Wook Kwon 한국응용곤충학회 2016 한국응용곤충학회 학술대회논문집 Vol.2016 No.04

In recent years, high-throughput next-generation sequencing (NGS) techniques have provided fascinating opportunities to understand the biology of non-model organisms, especially insect species. The decrease in sequencing costs and extensive sequencing services from NGS providers has brought many entomologists to be involved in genome sequencing. However, poor planning can lead to extremely fragmented genome assemblies which prevents high quality gene annotation and other desired analyses. Insect genomes can be problematic to assemble, due to combinations of high polymorphism, inability to breed for genome homozygosity, and small physical sizes limiting the quantity of DNA able to be isolated from a single individual. Given to the rapid development of host resistance to multiple classes of insecticides, it is indispensable to study the comprehensive genomic information of insects. Recent advances in sequencing technology and assembly strategies can able to fetch breakthroughs in deciphering the genetic information of insects. Here, we present the cost effective high throughput genome sequencing and assembly strategies for insect species in respects to taxonomy, evolutionary history, immune response, drug development, insect host-virus interactions and pest management etc.

Protein disulfide isomerase has a regulatory role in endosperm development in rice

Yeon Jeong Kim,Ga Hyun Son,Markkandan Kesavan,Sung-Il Kim,Hak Soo Seo 한국육종학회 2012 한국육종학회 심포지엄 Vol.2012 No.07

Protein disulfide isomerase (PDI) is a chaperone protein that involves in oxidative protein folding by acting as catalysts and folding assistants in the endoplasmic reticulum (ER). Genome database showed that rice contains three PDI-like genes. But, their functions and subcellullar localization are not clearly identified. Here, we show possible functions of rice PDI (OsPDI) during seed development. Seeds of OsPDI T-DNA insertion mutants which were identified by genomic DNA PCR and western blot display chalky phenotype. Electron microscope analysis revealed that endosperms of the OsPDIL1-1Δ mutant show imperfect packing of round starch granules, causing floury-white color. Abnormal form of protein body I (PB-I) containing prolamin and thick aleurone layer were also observed in the OsPDIL1-1Δ mutants. Protein content per seed was significantly low in the OsPDIL1-1Δ mutant. However, free sugar content was high in the OsPDIL1-1Δ mutant seed. Northern and western blot analyses showed that during seed development, OsPDI protein is steadily accumulated in the seed until maturation while its transcript level was highest at 10 days after flowering and rapidly decreased to basal level. In addition, OsPDI strongly interacts with cysteine protease OsCP1 and chaperone BiP protein accumulates in OsPDIL1-1Δ mutant. Besides, proteomic analysis of the OsPDIL1-1Δ mutant seed showed that OsPDI is post-translationally regulated and its loss causes accumulation of many types of seed proteins. Our results indicate that OsPDI plays a critical role in seed development through its regulatory activity for various proteins.

An efficient and tunable parameter to improve variant calling for whole genome and exome sequencing data

안용주,Kesavan Markkandan,백인표,문세영,이우석,김희수,한규동 한국유전학회 2018 Genes & Genomics Vol.40 No.1

Next generation sequencing (NGS) has traditionally been performed in various fields including agricultural to clinical and there are so many sequencing platforms available in order to obtain accurate and consistent results. However, these platforms showed amplification bias when facilitating variant calls in personal genomes. Here, we sequenced whole genomes and whole exomes from ten Korean individuals using Illumina and Ion Proton, respectively to find the vulnerability and accuracy of NGS platform in the GC rich/poor area. Overall, a total of 1013 Gb reads from Illumina and ~39.1 Gb reads from Ion Proton were analyzed using BWA-GATK variant calling pipeline. Furthermore, conjunction with the VQSR tool and detailed filtering strategies, we achieved high-quality variants. Finally, each of the ten variants from Illumina only, Ion Proton only, and intersection was selected for Sanger validation. The validation results revealed that Illumina platform showed higher accuracy than Ion Proton. The described filtering methods are advantageous for large population-based whole genome studies designed to identify common and rare variations associated with complex diseases.

De novo assembly and functional annotation of the emma field cricket (Teleogryllus emma) transcriptome

이준하,Kesavan Markkandan,김인우,김선영,서민철,김미애,김성현,황재삼 한국응용곤충학회 2019 Journal of Asia-Pacific Entomology Vol.22 No.1

In recent years, insects have been used in animal-based foods for sustainable live-stock management and in the human diet as therapeutic foods. These insects are classified as edible and are widely consumed around the world for their broad medicinal and nutritional values. Antimicrobial peptides/proteins (AMPs) confer resistance to microbial pathogens and contribute to the immune function of insects. The emma field cricket (Teleogryllus emma) is widely consumed around the world for its broad-spectrum therapeutic effects. However, the absence of genetic information on T. emma is an obstacle to understanding the diversity and activity of its AMPs. Hence, deep sequencing of the T. emma transcriptome and expression profiling are essential. We report whole-body de novo transcriptome profiling of T. emma and analysis of its potential bacterial immunization-induced AMPs. We assembled 18.2 Gb of Illumina reads, generated 102,311 transcripts (minimum length: > 500 bp, N50: 2585 bp), and identified 67,838 unigenes. Of these unigenes, 39,937 (58.87%) matched known proteins in a Basic Local Alignment Search Tool search of the National Center for Biotechnology Information database. Functional analysis demonstrated that the 39,937 unigenes clustered under 5707 Gene Ontology terms. In addition, we identified 10 cDNAs encoding new AMPs via an in-silico approach using public databases. Our transcriptome data set and AMP profile greatly improve the understanding of T. emma genetics and provide a huge number of gene sequences, including those for genes of known and unknown function, for further study.