Genetic specificity study using next-generation sequencing (NGS) of peritoneal metastatic colorectal cancer compared to primary colorectal cancer.

BACKGROUND: In patients with colorectal cancer, peritoneal metastases are the second most frequent metastatic lesion after liver metastases. Peritoneal metastases have a very poor prognosis, with a median survival time of 5-7 months. Currently, there is a lack of research on the genetic differences between primary colorectal cancer and peritoneal metastases. Therefore, we aimed to identify their genetic characteristics through a cancer panel test using next-generation sequencing. OBJECTIVE: We aim to investigate the specificity of genetic variants in primary colorectal cancer and peritoneal metastases. METHODS: We recruited patients with stage I, II, and III primary colorectal cancer and peritoneal metastases for genetic analysis using NGS. Samples were collected from patients who underwent surgery at Dankook University Hospital and consented to genetic testing. NGS was performed using a cancer panel. RESULTS: Among 36 patients with primary cancer, TP53 gene mutation was identified the most in 25 patients (69%), followed by APC gene mutation in 19 patients (53%), and KRAS gene mutation in 17 patients (47%). In the peritoneal metastasis patient group, unlike the primary cancer patient group, KRAS gene mutations were the most common 6 patients (55%), followed by TP53 gene mutations in 4 patients (36%) and PIK3CA gene mutations in 2 patients (18%). CONCLUSION: The small number of surgical cases of peritoneal metastases was a limitation of our sample size. Nevertheless, we identified differences in the alterations of specific genes between primary and peritoneal metastases. Acquiring additional cases and collecting more data will provide deeper insights into these cancers.

Human Endogenous Retrovirus-K (HML-2)-Related Genetic Variation: Human Genome Diversity and Disease.

Human endogenous retroviruses (HERVs) comprise a significant portion of the human genome, making up roughly 8%, a notable comparison to the 2-3% represented by coding sequences. Numerous studies have underscored the critical role and importance of HERVs, highlighting their diverse and extensive influence on the evolution of the human genome and establishing their complex correlation with various diseases. Among HERVs, the HERV-K (HML-2) subfamily has recently attracted significant attention, integrating into the human genome after the divergence between humans and chimpanzees. Its insertion in the human genome has received considerable attention due to its structural and functional characteristics and the time of insertion. Originating from ancient exogenous retroviruses, these elements succeeded in infecting germ cells, enabling vertical transmission and existing as proviruses within the genome. Remarkably, these sequences have retained the capacity to form complete viral sequences, exhibiting activity in transcription and translation. The HERV-K (HML-2) subfamily is the subject of active debate about its potential positive or negative effects on human genome evolution and various pathologies. This review summarizes the variation, regulation, and diseases in human genome evolution arising from the influence of HERV-K (HML-2).

Ankle dorsiflexion assistance of patients with foot drop using a powered ankle-foot orthosis to improve the gait asymmetry.

BACKGROUND: Foot drop is a neuromuscular disorder that causes abnormal gait patterns. This study developed a pneumatically powered ankle-foot orthosis (AFO) to improve the gait patterns of patients with foot drop. We hypothesized that providing unilateral ankle dorsiflexion assistance during the swing phase would improve the kinematics and spatiotemporal gait parameters of such patients. Accordingly, this study aims to examine the efficacy of the proposed assistance system using a strategy for joint kinematics and spatiotemporal gait parameters (stride length, swing velocity, and stance phase ratio). The analysis results are expected to provide knowledge for better design and control of AFOs in patients with foot drop. METHOD: Ten foot drop patients with hemiparesis (54.8 y ± 14.1 y) were fitted with a custom AFO with an adjustable calf brace and portable air compressor for ankle dorsiflexion assistance in the gait cycle during the swing phase. All subjects walked under two different conditions without extensive practice: (1) barefoot and (2) wearing a powered AFO. Under each condition, the patients walked back and forth on a 9-m track with ten laps of level ground under the supervision of licensed physical therapists. The lower-limb joint and trunk kinematics were acquired using 12 motion-capture cameras. RESULTS: We found that kinematic asymmetry decreased in the three lower-limb joints after ankle dorsiflexion assistance during the swing phase. The average ankle-joint angle increased after using the AFO during the entire gait cycle. Similarly, the knee-joint angle showed a slight increase while using the AFO, leading to a significantly decreased standard deviation within patients. Conversely, the hip-joint angle showed no significant improvements with assistance. While several patients exhibited noticeably lower levels of asymmetry, no significant changes were observed in the average asymmetry of the swing velocity difference between the affected and unaffected sides while using the AFO. CONCLUSION: We experimentally validated that ankle dorsiflexion assistance during the swing phase temporarily improves gait asymmetry in foot-drop patients. The experimental results also prove the efficacy of the developed AFO for gait assistance in foot-drop patients.

Comparison of digital PCR platforms using the molecular marker.

Assays of clinical diagnosis and species identification using molecular markers are performed according to a quantitative method in consideration of sensitivity, cost, speed, convenience, and specificity. However, typical polymerase chain reaction (PCR) assay is difficult to quantify and have various limitations. In addition, to perform quantitative analysis with the quantitative real-time PCR (qRT-PCR) equipment, a standard curve or normalization using reference genes is essential. Within the last a decade, previous studies have reported that the digital PCR (dPCR) assay, a third-generation PCR, can be applied in various fields by overcoming the shortcomings of typical PCR and qRT-PCR assays. We selected Stilla Naica System (Stilla Technologies), Droplet Digital PCR Technology (Bio-Rad), and Lab on an Array Digital Real-Time PCR analyzer system (OPTOLANE) for comparative analysis among the various droplet digital PCR platforms currently in use commercially. Our previous study discovered a molecular marker that can distinguish Hanwoo species (Korean native cattle) using Hanwoo-specific genomic structural variation. Here, we report the pros and cons of the operation of each dPCR platform from various perspectives using this species identification marker. In conclusion, we hope that this study will help researchers to select suitable dPCR platforms according to their purpose and resources.

Comparison of genetic variation between primary colorectal cancer and metastatic peritoneal cancer.

BACKGROUND: Among cancer metastases by primary colorectal cancer (CRC), peritoneal metastasis is the second most common metastatic lesion after liver metastasis. In treating metastatic CRC, it is very important to differentiate targeted-therapy and chemotherapy according to the characteristics of each lesion because the genetic variation of the primary and metastatic lesions are different. However, there are few studies of genetic characteristics on peritoneal metastasis caused by primary CRC, so molecular-level studies are continuously required. OBJECTIVE: We propose an appropriate peritoneal metastasis treatment policy by identifying the genetic characteristics between primary CRC and synchronous peritoneal metastatic lesions. METHODS: Primary CRC and synchronous peritoneal metastasis samples were analyzed in pairs from six patients using Comprehensive Cancer Panel (409 cancer-related genes, Thermo Fisher Scientific, USA) and next-generation sequencing (NGS). RESULTS: The mutations were commonly found on the KMT2C and THBS1 genes in both primary CRC and peritoneal metastasis. The PDE4DIP gene was mutated in all cases except for on a sample of peritoneal metastasis. As a result of analysis using the mutation database, we confirmed that the gene mutations of primary CRC and the peritoneal metastasis derived from it showed the same tendency, although we did not accompany the gene expression level or epigenetic study. CONCLUSION: It is thought that the treatment policy through molecular genetic testing of primary CRC can also be applied to peritoneal metastasis treatment. Our study is expected to be the basis for further peritoneal metastasis research.

Supervised chemical graph mining improves drug-induced liver injury prediction.

Drug-induced liver injury (DILI) is the main cause of drug failure in clinical trials. The characterization of toxic compounds in terms of chemical structure is important because compounds can be metabolized to toxic substances in the liver. Traditional machine learning approaches have had limited success in predicting DILI, and emerging deep graph neural network (GNN) models are yet powerful enough to predict DILI. In this study, we developed a completely different approach, supervised subgraph mining (SSM), a strategy to mine explicit subgraph features by iteratively updating individual graph transitions to maximize DILI fidelity. Our method outperformed previous methods including state-of-the-art GNN tools in classifying DILI on two different datasets: DILIst and TDC-benchmark. We also combined the subgraph features by using SMARTS-based frequent structural pattern matching and associated them with drugs' ATC code.

The inflammatory signature in monocytes of Sjögren's syndrome and systemic lupus erythematosus, revealed by the integrated Reactome and drug target analysis.

BACKGROUND: The innate immune regulation, especially by the type I IFN signature in the CD14+ monocytes, is known to be critical in the pathogenesis of autoimmune Sjögren's syndrome (SjS) and systemic lupus erythematosus (SLE). OBJECTIVE: Since patients with one condition can be overlapped with another, this study is to identify shared differentially expressed genes (DEGs) in SjS and SLE compared to healthy controls (HCs) and refine transcriptomic profiles with the integrated Reactome and gene-drug network analysis for an anti-inflammation therapy. METHODS: CD14+ monocytes were purified from whole blood of SjS and SLE patients (females, ages from 32 to 62) and subject to bulk RNA-sequencing, followed by data analyses for comparison with HC monocytes (females, ages 30 and 33). Functional categorizations, using Gene Ontology (GO) and the Reactome pathway analysis, were performed and DEGs associated with therapeutic drugs were identified from the Drug Repurposing Hub (DHUB) database. RESULTS: The GO analysis revealed that DEGs in the inflammatory response and the cellular response to cytokine were highly enriched in both conditions. A propensity toward M1 macrophage differentiation appears to be prominent in SjS while the Response to Virus was significant in SLE monocytes. Through the Reactome pathway analysis, DEGs in the IFN signaling and the cytokine signaling in immune system were most significantly enriched in both. Upregulation of NGF-induced transcription activity in SjS and the complement cascade activity in SLE were also noted. Multiple anti-inflammatory drugs, such as prostaglandin-endoperoxide synthase and angiotensin-I-converting- enzyme were associated with the DEGs in these conditions. CONCLUSIONS: Taken together, our analysis indicates distinct inflammatory transcriptomic profiles shared in SjS and SLE monocytes. Comprehensive characterizations of the data from these conditions will ultimately allow differential diagnosis of each condition and identification of therapeutic targets.

Identification of Potentially Pathogenic Variants Associated with Recurrence in Medication-Related Osteonecrosis of the Jaw (MRONJ) Patients Using Whole-Exome Sequencing.

Background: Bisphosphonates are antiresorptive and antiangiogenic drugs that prevent and treat bone loss and mineralization in women with postmenopausal osteoporosis and cancer patients. Medication-related osteonecrosis of the jaw (MRONJ) is commonly caused by tooth extraction and dental trauma. Although genetic and pathological studies about MRONJ have been conducted, the pathogenesis of MRONJ still remains unclear. Methods: We aimed to identify genetic variants associated with MRONJ, using whole-exome sequencing (WES). Ten MRONJ patients prescribed bisphosphonates were recruited for WES, and jawbone tissue and blood samples were collected from the patients. Results: The analysis of the WES data found a total of 1866 SNP and 40 InDel variants which are specific to MRONJ. The functional classification assay using Gene Ontology and pathway analysis discovered that genes bearing the MRONJ variants are significantly enriched for keratinization and calcium ion transport. Some of the variants are potential pathogenic variants (24 missense mutations and seven frameshift mutations) with MAF < 0.01. Conclusions: The variants are located in eight different genes (KRT18, MUC5AC, NBPF9, PABPC3, MST1L, ASPN, ATN1, and SLAIN1). Nine deleterious SNPs significantly associated with MRONJ were found in the KRT18 and PABPC3 genes. It suggests that KRT18 and PABPC3 could be MRONJ-related key genes.

Rapid identification of SARS-CoV-2 in the point-of-care using digital PCR-based Dr. PCR™ Di20K COVID-19 Detection Kit without viral RNA extraction.

BACKGROUND: Since COVID-19 was declared the pandemic by the WHO, it has continued to spread. There is a need for rapid, efficient, and accurate diagnostic kits and techniques to control its spread. OBJECTIVE: The diagnostic capability of the qRT-PCR-based Real-Q 2019-nCoV Detection Kit and dPCR-based Dr. PCR™ Di20K COVID-19 Detection Kit was compared and evaluated. METHODS: Diagnostic tests for COVID-19 were performed using two different COVID-19 kits and 301 individual specimens with confirmed COVID-19 positive/negative at the government-accredited medical institution. Assessment of diagnostic capability was measured through diagnostic sensitivity, specificity, Cohen's Kappa coefficient, and dilutional linearity tests. RESULTS: The COVID-19 diagnostic test results using two kits and 301 individual specimens perfectly matched the pre-diagnosis results of the medical institution. In addition, the measurement results of diagnostic sensitivity and specificity were "1", indicating high diagnostic capability. Cohen's Kappa coefficient value is "1", which means that the diagnosis concordance between the two kits is "Almost Perfect". As a result of dilutional linearity tests to evaluate their detection capability, both kits were measured with very high detection reliability. CONCLUSION: Here, we propose that the dPCR-based Dr. PCR™ Di20K COVID-19 Detection Kit has the advantages of the dPCR method reported in the previous study and is suitable for point-of-care testing (POCT) by overcoming the limitations of space, test time, cross-over contamination, and biosafety due to omitting RNA extraction process.

High-accuracy quantitative principle of a new compact digital PCR equipment: Lab On An Array.

Digital PCR (dPCR) is the third-generation PCR that enables real-time absolute quantification without reference materials. Recently, global diagnosis companies have developed new dPCR equipment. In line with the development, the Lab On An Array (LOAA) dPCR analyzer (Optolane) was launched last year. The LOAA dPCR is a semiconductor chip-based separation PCR type equipment. The LOAA dPCR includes Micro Electro Mechanical System that can be injected by partitioning the target gene into 56 to 20,000 wells. The amount of target gene per wells is digitized to 0 or 1 as the number of well gradually increases to 20,000 wells because its principle follows Poisson distribution, which allows the LOAA dPCR to perform precise absolute quantification. LOAA determined region of interest first prior to dPCR operation. To exclude invalid wells for the quantification, the LOAA dPCR has applied various filtering methods using brightness, slope, baseline, and noise filters. As the coronavirus disease 2019 has now spread around the world, needs for diagnostic equipment of point of care testing (POCT) are increasing. The LOAA dPCR is expected to be suitable for POCT diagnosis due to its compact size and high accuracy. Here, we describe the quantitative principle of the LOAA dPCR and suggest that it can be applied to various fields.

Diagnostic evaluation of qRT-PCR-based kit and dPCR-based kit for COVID-19.

BACKGROUND: Coronavirus disease of 2019 (COVID-19) is well known as a fatal disease, first discovered at Wuhan in China, ranging from mild to death, such as shortness of breath and fever. Early diagnosis of COVID-19 is a crucial point in preventing global prevalence. OBJECTIVE: We aimed to evaluate the diagnostic competency and efficiency with the Allplex™ 2019-nCoV Assay kit and the Dr. PCR 20 K COVID-19 Detection kit, designed based on the qRT-PCR and dPCR technologies, respectively. METHODS: A total of 30 negative and 20 COVID-19 positive specimens were assigned to the diagnostic test by using different COVID-19 diagnosis kits. Diagnostic accuracy was measured by statistical testing with sensitivity, specificity, and co-efficiency calculations. RESULTS: Comparing both diagnostic kits, we confirmed that the diagnostic results of 30 negative and 20 positive cases were the same pre-diagnostic results. The diagnostic statistics test results were perfectly matched with value (1). Cohen's Kappa coefficient was demonstrated that the given kits in two different ways were "almost perfect" with value (1). In evaluating the detection capability, the dilutional linearity experiments substantiate that the Dr. PCR 20 K COVID-19 Detection kit could detect SARS-CoV-2 viral load at a concentration ten times lower than that of the Allplex™ 2019-nCoV Assay kit. CONCLUSIONS: In this study, we propose that the dPCR diagnosis using LOAA dPCR could be a powerful method for COVID-19 point-of-care tests requiring immediate diagnosis in a limited time and space through the advantages of relatively low sample concentration and small equipment size compared to conventional qRT-PCR.

Post-vaccination Monitoring to Assess Foot-and-Mouth Disease Immunity at Population Level in Korea.

In South Korea, domestic cattle, pigs, and goats were subjected to mandatory foot-and-mouth disease (FMD) vaccination and year-round serosurveillance since 2011. In 2020, approximately USD 95 million was spent solely for FMD vaccine purchase for 59 million livestock, and 1.25 million samples were tested to estimate the population immunity and demonstrate the absence of virus circulation. As the FMD vaccination program was revised in 2018, the post-vaccination monitoring (PVM) was designed to evaluate the effectiveness of the vaccine program of three vaccines approved for routine use. To this end, monitoring post-vaccination immunity has been conducted by collecting 35,626 serum samples at 28 days post-vaccination following regular national vaccinations, which were carried out in April and in October in 2020. The design of the serological test for PVM was specially targeted at particular livestock groups, including dairy cattle, goats, and beef cattle aged 6-12 months, which were generally estimated to have a low expected seroprevalence. The risk factors had also been identified, considering the increased likelihood of infection in a particular location, herd size, and husbandry system applied in a targeted sample collection. Serum sample collection and SP-O and NSP antibody tests were performed by local veterinary laboratories using commercially available ELISAs. The current FMD vaccination program, which was performed twice a year following the regimen of primary vaccination and boost, resulted in over 80% population immunity. The seroprevalence monitored after the vaccination in fall was higher than the one studied in spring except in pigs. It was demonstrated that the seroprevalence of risk-based targeted samples ranged from 93.8 to 100% in cattle, 63.2 to 100% in pigs, and 20.0 to 100% in goats. Of note is the area near the North Korean borders which showed a relatively low seroprevalence among the targeted regions, and no NSP sero-positive reactor was detected in this region. When subpopulation immunity at the individual level was assessed, the seroprevalence in young cattle stock was slightly lower (95.8%) than that of adults (98.4%). In conclusion, the FMD vaccination campaign has been successfully implemented in Korea, and the PVM can be a supplementary program for massive routine surveillance in terms of providing timely information needed both to estimate population immunity and to properly target "risk-based surveillance."

Differential expressions of L1-chimeric transcripts in normal and matched-cancer tissues.

L1s are a cis-regulatory elements and contain bidirectional internal promoters within the 5' untranslated region (UTR). L1s provide bidirectional promoters that generate alternative transcripts and affect differential expressions in the human genome. In particular, L1 antisense promoters (L1ASPs) could produce aberrant transcripts in cancer tissues compared to normal tissues. In this study, we identified the L1-chimeric transcripts derived from L1ASPs and analyzed relative expression of L1-chimeric transcripts between normal and matched-cancer tissues. First, we collected 425 L1-chimeric transcripts by referring to previous studies. Through the manual inspection, we identified 144 L1-chimeric transcripts derived from 44 L1 antisense promoters, suggesting that the antisense promoter acted as an alternative promoter. We analyzed relative gene expression levels of 16 L1-chimeric transcripts between matched cancer-normal tissue pair (lung, liver, gastric, kidney, thyroid, breast, ovary, uterus, and prostate) using real-time quantitative PCR (RT-qPCR) and investigated putative transcription factor binding motifs to determine activity of L1ASPs. Taken together, we propose that L1ASPs could contribute to the differential gene expression between normal and cancer tissues.

Quantitative evaluation of the molecular marker using droplet digital PCR.

Transposable elements (TEs) constitute approximately half of Bovine genome. They can be a powerful species-specific marker without regression mutations by the structure variation (SV) at the time of genomic evolution. In a previous study, we identified the Hanwoo-specific SV that was generated by a TE-association deletion event using traditional PCR method and Sanger sequencing validation. It could be used as a molecular marker to distinguish different cattle breeds (i.e., Hanwoo vs. Holstein). However, PCR is defective with various final copy quantifications from every sample. Thus, we applied to the droplet digital PCR (ddPCR) platform for accurate quantitative detection of the Hanwoo-specific SV. Although samples have low allele frequency variation within Hanwoo population, ddPCR could perform high sensitive detection with absolute quantification. We aimed to use ddPCR for more accurate quantification than PCR. We suggest that the ddPCR platform is applicable for the quantitative evaluation of molecular markers.

Application of NanoString technologies in angioimmunoblastic T cell lymphoma.

BACKGROUND: Angioimmunoblastic T-cell lymphoma (AITL) is an aggressive disease. Most cancer diagnoses are determined by anatomical histology. Therefore, many samples are stored in FFPE blocks for H&E staining. However, RNAs extracted from the FFPE block have a high level of fragmentation, making it difficult to perform accurate DEG analysis using RNA sequencing. OBJECTIVE: To overcome fragmented RNA's drawback in NGS application, we applied the NanoString nCounter® technique of hybridization method that can be used for DEG analysis without PCR amplification. METHODS: We characterized the gene expression profiling of AITLs though transcriptome analysis based on the nCounter® PanCancer IO 360™ Panel and NanoString platform. To perform the analysis of differential expression gene (DEG) profiles in AITLs, we compared the NanoString data from eight AITL patients with a healthy control donor. RESULTS: Ninety-one genes were up-regulated and six genes were down-regulated in AITLs compared to control. The Gene Ontology (GO) analysis of 97-DEGs revealed that they were closely related to cytokine, MAPK cascade, leukocyte differentiation, and immune response, suggesting that this affect the immune system. In addition, KEGG analysis revealed that AITL DEGs were found to be highly involved in cytokine-cytokine receptor interaction and PI3K-Akt signaling pathway. CONCLUSION: We believe that comprehensive multiplex studies, along with NanoString analysis, may be helpful to understand the molecular mechanisms of AITL, including mutations, gene expression, and protein expression studies.

Impact of Sleep Disorder as a Risk Factor for Dementia in Men and Women.

Sleep is an essential physiological process, especially for proper brain function through the formation of new pathways and processing information and cognition. Therefore, when sleep is insufficient, this can result in pathophysiologic conditions. Sleep deficiency is a risk factor for various conditions, including dementia, diabetes, and obesity. Recent studies have shown that there are differences in the prevalence of sleep disorders between genders. Insomnia, the most common type of sleep disorder, has been reported to have a higher incidence in females than in males. However, sex/gender differences in other sleep disorder subtypes are not thoroughly understood. Currently, increasing evidence suggests that gender issues should be considered important when prescribing medicine. Therefore, an investigation of the gender-dependent differences in sleep disorders is required. In this review, we first describe sex/gender differences not only in the prevalence of sleep disorders by category but in the efficacy of sleep medications. In addition, we summarize sex/gender differences in the impact of sleep disorders on incident dementia. This may help understand gender-dependent pathogenesis of sleep disorders and develop therapeutic strategies in men and women.

Comparison of library construction kits for mRNA sequencing in the Illumina platform.

BACKGROUND: The emergence of next-generation sequencing (NGS) technologies has made a tremendous contribution to the deciphering and significance of transcriptome analysis in biological fields. Since the advent of NGS technology in 2007, Illumina, Inc. has provided one of the most widely used sequencing platforms for NGS analysis. OBJECTIVE: Although reagents and protocols provided by Illumina are adequately performed in transcriptome sequencing, recently, alternative reagents and protocols which are relatively cost effective are accessible. However, the kits derived from various manufacturers have advantages and disadvantages when researchers carry out the transcriptome library construction. METHODS: We compared them using a variety of protocols to produce Illumina-compatible libraries based on transcriptome. Three different mRNA sequencing kits were selected for this study: TruSeq® RNA Sample Preparation V2 (Illumina, Inc., USA), Universal Plus mRNA-Seq (NuGEN, Ltd., UK), and NEBNext® Ultra™ Directional RNA Library Prep Kit for Illumina® (New England BioLabs, Ltd., USA). We compared them focusing on cost, experimental time, and data output. RESULTS: The quality and quantity of sequencing data obtained through the NGS technique were strongly influenced by the type of the sequencing library kits. It suggests that for transcriptome studies, researchers should select a suitable library construction kit according to the goal and resources of experiments. CONCLUSION: The present work will help researchers to choose the right sequencing library construction kit for transcriptome analyses.

Novel Discovery of LINE-1 in a Korean Individual by a Target Enrichment Method.

Long interspersed element-1 (LINE-1 or L1) is an autonomous retrotransposon, which is capable of inserting into a new region of genome. Previous studies have reported that these elements lead to genomic variations and altered functions by affecting gene expression and genetic networks. Mounting evidence strongly indicates that genetic diseases or various cancers can occur as a result of retrotransposition events that involve L1s. Therefore, the development of methodologies to study the structural variations and interpersonal insertion polymorphisms by L1 element-associated changes in an individual genome is invaluable. In this study, we applied a systematic approach to identify human-specific L1s (i.e., L1Hs) through the bioinformatics analysis of high-throughput next-generation sequencing data. We identified 525 candidates that could be inferred to carry non-reference L1Hs in a Korean individual genome (KPGP9). Among them, we randomly selected 40 candidates and validated that approximately 92.5% of non-reference L1Hs were inserted into a KPGP9 genome. In addition, unlike conventional methods, our relatively simple and expedited approach was highly reproducible in confirming the L1 insertions. Taken together, our findings strongly support that the identification of non-reference L1Hs by our novel target enrichment method demonstrates its future application to genomic variation studies on the risk of cancer and genetic disorders.

Investigation of Hanwoo-specific structural variations using whole-genome sequencing data.

BACKGROUND: The total length of the cattle genome is approximately ~ 3 billion base pairs. About half of the bovine genome (46.5%) is composed of transposable elements (TEs). The TEs could be a major source of genomic structural variations (SVs) between cattle breeds. These SVs have led to genomic fluidity and rearrangements between interspecies. OBJECTIVE: TE-mediated insertion and deletion events could have a strong influence on the bovine genome. This study aimed to investigate TE-mediated deletion events that are common to 12 Hanwoo genome resequencing data. RESULTS: We compared 12 Hanwoo genome resequencing data with the cattle reference genome (Bos taurus_UMD_3.1.1) and six other open source data (2 Jersey, 2 Holstein, 2 Angus). By using BreakDancer program, the common SVs to the 12 Hanwoo genomes were detected. A total of 299 Hanwoo-specific SV candidates were detected. Among them, 56 Hanwoo-specific TE-mediated deletion candidate loci were validated by PCR and Sanger sequencing. Finally, we identified one locus, DEL_96, which is an authentic Hanwoo-specific deletion. The DEL_96 event occurred by nonallelic homologous end-joining between LINE (BovB) and unique sequence with 1 bp microhomology. The 370 bp deletion event appeared to be only in the Hanwoo individuals after the divergence of Hanwoo and Holstein lineages. CONCLUSION: Our study showed that one of the SVs, TE-mediated deletion, could be utilized as a molecular maker to distinguish between Hanwoo and Holstein.

A Simple Guideline to Assess the Characteristics of RNA-Seq Data.

Next-generation sequencing (NGS) techniques have been used to generate various molecular maps including genomes, epigenomes, and transcriptomes. Transcriptomes from a given cell population can be profiled via RNA-seq. However, there is no simple way to assess the characteristics of RNA-seq data systematically. In this study, we provide a simple method that can intuitively evaluate RNA-seq data using two different principal component analysis (PCA) plots. The gene expression PCA plot provides insights into the association between samples, while the transcript integrity number (TIN) score plot provides a quality map of given RNA-seq data. With this approach, we found that RNA-seq datasets deposited in public repositories often contain a few low-quality RNA-seq data that can lead to misinterpretations. The effect of sampling errors for differentially expressed gene (DEG) analysis was evaluated with ten RNA-seq data from invasive ductal carcinoma tissues and three RNA-seq data from adjacent normal tissues taken from a Korean breast cancer patient. The evaluation demonstrated that sampling errors, which select samples that do not represent a given population, can lead to different interpretations when conducting the DEG analysis. Therefore, the proposed approach can be used to avoid sampling errors prior to RNA-seq data analysis.