Development of gene microarray in screening differently expressed genes in keloid and normal-control skin

Background Keloid is an intricate lesion that is probably regulated by many genes. In this study,the authors used the technique of complementary DNA (cDNA) microarray to analyse abnormal gene expression in keloids and normal control skins. Methods The polymerase chain reaction (PCR) products of 8400 genes were spotted in an array on chemical-material-coated-glass plates. The DNAs were fixed on the glass plates. The total RNAs were isolated from freshly excised human keloid and normal control skins,and the mRNAs were then purified. The mRNA from both keloid and normal control skins were reversely transcribed to cDNAs,with the incorporation of fluorescent dUTP,for preparing the hybridisation probes. The mixed probes were then hybridised to the cDNA microarray. After thorough washing,the cDNA microarray was scanned for differing fluorescent signals from two types of tissues. Gene expression of tissue growth factor-β_1 (TGF-β_1) and of c-myc was detected with both RT-PCR and Northern blot hybridisation to confirm the effectiveness of cDNA microarray. Results Among the 8400 human genes,402 were detected with different expression levels between keloid and normal control skins. Two hundred and fifty genes,including TGF-β_1 and c-myc,were up-regulated and 152 genes were down-regulated. Higher expressions of TGF-β_1 and c-myc in keloid were also revealed using RT-PCR and Northern blot methods. Conclusion cDNA microarray analysis provides a powerful tool for investigating differential gene expression in keloid and normal control skins. Keloid is a complicated lesion with many genes involved.

Deep Learning Enabled Microarray Gene Expression Classification for Data Science Applications

In bioinformatics applications,examination of microarray data has received significant interest to diagnose diseases.Microarray gene expression data can be defined by a massive searching space that poses a primary challenge in the appropriate selection of genes.Microarray data classification incorporates multiple disciplines such as bioinformatics,machine learning(ML),data science,and pattern classification.This paper designs an optimal deep neural network based microarray gene expression classification(ODNN-MGEC)model for bioinformatics applications.The proposed ODNN-MGEC technique performs data normalization process to normalize the data into a uniform scale.Besides,improved fruit fly optimization(IFFO)based feature selection technique is used to reduce the high dimensionality in the biomedical data.Moreover,deep neural network(DNN)model is applied for the classification of microarray gene expression data and the hyperparameter tuning of the DNN model is carried out using the Symbiotic Organisms Search(SOS)algorithm.The utilization of IFFO and SOS algorithms pave the way for accomplishing maximum gene expression classification outcomes.For examining the improved outcomes of the ODNN-MGEC technique,a wide ranging experimental analysis is made against benchmark datasets.The extensive comparison study with recent approaches demonstrates the enhanced outcomes of the ODNN-MGEC technique in terms of different measures.

Development of a 16S r RNA gene-based microarray for the detection of marine bacterioplankton community

A better understanding of bacterioplankton community shifts following change in marine environments is critical to predict the marine ecosystem function. In order to get a snapshot of the microbial taxonomy profiling of a wide range marine area, a quick, convenient and low cost method would be favorable. In this study, we developed a 16S rRNA gene-based microarray using ARB software, which contained 447 probes targeting 160 families of marine bacteria. The specificity, sensitivity and quantitative capability of this microarray were assessed by single cloned16S rRNA genes. The reliability of this microarray was tested by eight environmental samples. The results showed that the microarray was specific, only 1.16% false results were detected in five single-clone hybridization tests. The microarray could detect DNA samples as few as 1 ng/μL and the signal intensity could reflect the relative abundance of the bacteria in the range of 1 ng/μL to 100 ng/μL of DNA concentration. Hybridization with environmental samples showed that it can discriminate bacterioplankton communities by sites and time. High throughput sequencing results from the eight samples confirmed the hybridization results. It indicated that this developed microarray could be used as a convenient tool to monitor the bacterioplankton community in marine environment.

Analysis of differential genes of ischemic stroke-induced heart tissue in mice based on GEO database

Objective:The differential genes of left ventricle in middle cerebral artery occlusion model(MCAO)mice and Sham mice(Sham)mice at 24h and 72h after ischemia were compared respectively,and the differential genes and their regulated functional pathways were analyzed at different time points after ischemic stroke,so as to analyze the mechanism of inducing cardiac dysfunction after ischemic stroke and provide evidence for its treatment.Methods:Gene-chip data from the left ventricle of MCAO mice and Sham mice were downloaded from the GEO database at the National Center for Biotechnology Information(NCBI).The differentially expressed genes were obtained by R language software programming.The GO functional enrichment and KEGG pathway enrichment analysis of the obtained differential genes were performed using DAVID 6.8 online analysis tool,and the Omicshare online analysis tool was used to visualize the enrichment analysis results.Results:At 24h after ischemia,187 differentially expressed genes were obtained,including 56 GO enrichment pathways and 5 KEGG enrichment pathways with significant significance.After 72h after ischemia,51 differentially expressed genes were obtained,14 GO enrichment pathways and 3 KEGG enrichment pathways with significant significance.The two time points involved Aplnr and Itgb6 gene targets and PI3K-Akt signaling pathway.Conclusion:①By analyzing the gene expression profile data,the differentially expressed genes and related pathways of cardiac dysfunction induced by ischemic stroke were obtained.②PI3K-AKT signaling pathway is closely related to the regulation of cardiac function,and regulation of PI3K-AKT signaling pathway may be an important direction for the treatment of cardiac dysfunction after ischemic stroke.

Spatiotemporal microRNA profile in peripheral nerve regeneration:miR-138 targets vimentin and inhibits Schwann cell migration and proliferation

While the peripheral nervous system has regenerative ability,restoration of sufficient function remains a challenge.Vimentin has been shown to be localized in axonal growth fronts and associated with nerve regeneration,including myelination,neuroplasticity,kinase signaling in nerve axoplasm,and cell migration;however,the mechanisms regulating its expression within Schwann cell（SC） remain unexplored.The aim of this study was to profile the spatial and temporal expression profile of micro RNA（mi RNA） in a regenerating rat sciatic nerve after transection,and explore the potential role of mi R-138-5 p targeting vimentin in SC proliferation and migration.A rat sciatic nerve transection model,utilizing a polyethylene nerve guide,was used to investigate mi RNA expression at 7,14,30,60,and 90 days during nerve regeneration.Relative levels of mi RNA expression were determined using microarray analysis and subsequently validated with quantitative real-time polymerase chain reaction.In vitro assays were conducted with cultured Schwann cells transfected with mi RNA mimics and assessed for migratory and proliferative potential.The top seven dysregulated mi RNAs reported in this study have been implicated in cell migration elsewhere,and GO and KEGG analyses predicted activities essential to wound healing.Transfection of one of these,mi RNA-138-5 p,into SCs reduced cell migration and proliferation.mi R-138-5 p has been shown to directly target vimentin in cancer cells,and the luciferase assay performed here in rat Schwann cells confirmed it.These results detail a role of mi R-138-5 p in rat peripheral nerve regeneration and expand on reports of it as an important regulator in the peripheral nervous system.

Gene Expression Profiling of Human c-Kit Mutant D816V

The tyrosine kinase receptor III, c-Kit/stem cell factor receptor and its ligand, human stem cell factor (huSCF) are the predominant regulator of mitogenesis in the hematopoietic stem and progenitor cells. However, gain-of-function mutations alter c-Kit auto-regulatory mechanisms to aberrant c-Kit signaling, leading to the onset or progression of cancerous transformations. The most common mutation of c-Kit is the substitution of aspartic acid residue in position 816 to valine (D816V), which is majorly responsible for its ligand-independent constitutive activation, and is implicated in hematopoietic malignancies. Currently, molecular targeted therapy is increasingly becoming a hot spot due to its specificity and low toxicity. As the molecular mechanisms responsible for D816V-c-Kit mediated tumorogenicity are largely unknown, in this study, we aimed to investigate the D816V-c-Kit signaling mediated downstream molecular targets. Specifically, we created c-Kit active mutant form D816V and performed inducible gene expression of mutant D816V-c-Kit in monomyelocytic cell line U937. Mutant D816V-c-Kit expressing cells revealed significantly enhanced cellular mitogenic activity compared to wild-type c-Kit expressing cells independent of huSCF. To examine the molecular targets regulating tumorogenic proliferation, we evaluated the consequences of mutant D816V-c-Kit expression on downstream gene expression profile by high throughput microarray technology. The levels of some of the relevant genes (PIK3CB, eIF4B, PRKCDBP, MOAP1) were validated by quantitative polymerase chain reaction. SLA, STAT5B, MAP3K2 and MAPK14 emerged as important downstream molecular targets of mutant D816V-c-Kit. Further, by dissecting the signaling pathways, we also demonstrated that the D816V-c-Kit mediated hematopoietic cell proliferation is dependent on molecular target p38 MAP kinase.

DNA样品目的基因相对拷贝数的定量斑点杂交测定方法

传统斑点杂交由于手工上样存在差异,难以进行目的基因定量分析。该文采用手动芯片点样仪,并结合高灵敏度的North2 south生物素标记和化学发光检测试剂盒进行定量斑点杂交,对水稻OsSec27p基因进行测试,结果表明样品中目的基因的拷贝数与杂交信号呈线性关系,根据该标准曲线可以测定未知样品目的基因的相对拷贝数。该文提供了一种经济、简便、切实可行的测定DNA样品中目的基因相对拷贝数的定量方法。

Altered expression profile of long non-coding RNAs during heart aging in mice

Objective:Long noncoding RNAs(lncRNAs)play an important role in regulating the occurrence and development of cardiovascular diseases.However,the role of lncRNAs in heart aging remains poorly understood.The objective of this study was to identify differentially expressed lncRNAs in the heart of aging mice and elucidate the relevant regulatory pathways of cardiac aging.Materials and methods:Echocardiography was used to detect the cardiac function of 18-months(aged)and 3-months(young)old C57BL/6 mice.Microarray analysis was performed to unravel the expression profiles of lncRNAs and mRNAs,and qRT-PCR to verify the highly dysregulated lncRNAs.Results:Our results demonstrated that the heart function in aged mice was impaired relative to young ones.Microarray results showed that 155 lncRNAs were upregulated and 37 were downregulated,and 170 mRNAs were significantly upregulated and 44 were remarkably downregulated in aging hearts.Gene ontology analysis indicated that differentially expressed genes are mainly related to immune function,cell proliferation,copper ion response,and cellular cation homeostasis.KEGG pathway analysis showed that the differentially expressed mRNAs are related to cytokine-cytokine receptor interaction,inflammatory mediator regulation of TRP channels,and the NF-kappa B signaling pathway.Conclusion:These results imply that the differentially expressed lncRNAs may regulate the development of heart aging.This study provides a new perspective on the potential effects and mechanisms of lncRNAs in heart aging.

Association of TCR-signaling pathway with the development of lacrimal gland benign lymphoepithelial lesions

·AIM: To identify the association of the T cell receptor(TCR) signaling with the development of benign lymphoepithelial lesions(BLEL) of the lacrimal gland.· METHODS: We collected affected lacrimal gland tissues from 9 patients who underwent dacryoadenectomy in the Capital Medical University Beijing Tongren Hospital Eye Center between August2010 and March 2013 and were confirmed to have lacrimal gland BLEL by histopathological analysis. Tumor tissues from 9 patients with orbital cavernous hemangioma were also collected and used as control.Whole genome gene expression microarray was used to compare gene expression profiles of affected lacrimal gland tissues from patients with lacrimal gland BLEL to those from of orbital cavernous hemangiomas.Differential expression of TCR pathway genes between these tissues was confirmed by polymerase chain reaction(PCR) and immunohistochemistry.·RESULTS: Microarray analysis showed that in lacrimal glands with BLEL, 32 signaling pathways were enriched in the upregulated genes, while 25 signaling pathways were enriched in the downregulated genes. In-depth analysis of the microarray data showed that the expression of 27 genes of the TCR signaling pathway increased significantly. To verify the differential expression of three of these genes, CD3, CD4, and interleukin(IL)-10, reverse transcription-PCR(RT-PCR)and immunohistochemistry assays were performed. RT-PCR analysis showed that CD3 and CD4 were expressed in the lacrimal glands with BLEL, but IL-10 was not expressed. Immunohistochemistry confirmed that CD3 and CD4 proteins were also present, but IL-10 protein was not. CD3, CD4, or IL-10 expression was not found in the orbital cavernous hemangiomas with either RT-PCR or immunohistochemistry.· CONCLUSION: TCR signaling pathway might be involved in the pathogenesis of lacrimal gland BLEL.

Laser capture microdissection enables cellular and molecular studies of tooth root development

Epithelial-mesenchymal interactions（EMIs） are critical for tooth development.Molecular mechanisms mediating these interactions in root formation is not well understood.Laser capture microdissection（LCM） and subsequent microarray analyses enable large scale in situ molecular and cellular studies of root formation but to date have been hindered by technical challenges of gaining intact histological sections of non-decalcified mineralized teeth or jaws with well-preserved RNA.Here,we describe a new method to overcome this obstacle that permits LCM of dental epithelia,adjacent mesenchyme,odontoblasts and cementoblasts from mouse incisors and molars during root development.Using this method,we obtained RNA samples of high quality and successfully performed microarray analyses.Robust differences in gene expression,as well as genes not previously associated with root formation,were identified.Comparison of gene expression data from microarray with real-time reverse transcriptase polymerase chain reaction（RT-PCR） supported our findings.These genes include known markers of dental epithelia,mesenchyme,cementoblasts and odontoblasts,as well as novel genes such as those in the fibulin family.In conclusion,our new approach in tissue preparation enables LCM collection of intact cells with well-preserved RNA allowing subsequent gene expression analyses using microarray and RT-PCR to define key regulators of tooth root development.

Characterization of the genes involved in nitrogen cycling in wastewater treatment plants using DNA microarray and most probable number-PCR

To improve nitrogen removal performance of wastewater treatment plants （WWTPs）, it is essential to understand the behavior of nitrogen cycling communities, which comprise various microorganisms. This study characterized the quantity and diversity of nitrogen cycling genes in various processes of municipal WWTPs by employing two molecular-based methods：most probable number-polymerase chain reaction （MPN-PCR） and DNA microarray. MPN-PCR analysis revealed that gene quantities were not statistically different among processes, suggesting that conventional actwated sludge processes （CAS） are similar to nitrogen removal processes in their ability to retain an adequate population of nitrogen cycling microorganisms. Furthermore, most processes in the WWTPs that were researched shared a pattern：the nitS and the bacterial amoA genes were more abundant than the nirK and archaeal amoA genes, respectivelv. DNA microarray analysis revealed that several kinds of nitrification and denitrification genes were detected in both CAS and anaerobic-oxic processes （AO）, whereas limited genes were detected in nitrogen removal processes. Results of this study suggest that CAS maintains a diverse community of nitrogen cycling microorganisms; moreover, the microbial communities in nitrogen removal processes may be specific.

Detection and identification of Vibrio parahaemolyticus by multiplex PCR and DNA-DNA hybridization on a microarray

In this paper, we developed a rapid and accurate method for the detection of Vibrio parahaemolyticus strains, using multiplex PCR and DNA--DNA hybridization. Multiplex PCR was used to simultaneously amplify three diagnostic genes （tlh, tdh andfla） that serve as molecular markers of V. parahaemolyticus. Biotinylated PCR products were hybridized to primers immobilized on a microarray, and detected by chemiluminesce with avidin-conjugated alkaline phosphatase. With this method, forty-five samples were tested. Eight known virulent strains （tlh＋/tdh＋/fla＋） and four known avirulent strains （tlh＋/tdh /fla＋） of the V. parahaemolyticus were successfully detected, and no non-specific hybridization and cross-hybridization reaction were found from fifteen closely-related strains （tlh-/tdh-/fla＋） of the Vibrio spp. In addition, all the other eighteen strains of non-Vibrio bacteria （tlh-/tdh /fla-） gave negative results. The DNA microarray successfully distinguished V. parahaemolyticus from other Vibrio spp. The results demonstrated that this was an efficient and robust method for identifying virulent strains of V. parahaemolyticus.

Effect of Aspirin on DMBA-induced mammary gland carcinogenesis and its anti-tumor mechanism in MCF-7 breast cancer cell

The effects of Aspirin on tumor chemoprevention and inhibition have been debated and researched in recent years and its effects on colorectal cancer are quite clear.For breast cancer,however,conclusions are inconsistent and the anti-tumor mechanism of Aspirin is not clear yet.In our study,we used DMBA-induced mammary gland carcinogenesis model to assess the chemoprevention effect of Aspirin on mammary precancerous lesions.After SD rats were treated with Aspirin,the total numbers of precancerous lesion in experimental groups were 16（40 mg/kg Aspirin） and 13（20 mg/kg Aspirin）,while the number in control group was 35.In vitro,we found that Aspirin inhibited cell proliferation in human breast cancer cell line MCF-7 by SRB assay with no apparent cytotoxity under the doses of 10,8,6,4 and 2 mM,the inhibitory rates were 86.96%,54.56%,24.83%,14.24% and 4.49%,respectively.In mechanism research,the results of gene microarray assay demonstrated that 4 mM and 2 mM Aspirin were effective in changing gene expression profile in MCF-7 cells.The expression of cell cycle regulator,cyclin A,was significantly down-regulated under the same doses,while the down-regulation of Cdk2 was only remarkable at 4 mM.Our findings reveal that Aspirin is effective in tumor inhibition during initial phase in rats.In MCF-7 cells,Aspirin reduces cell proliferation without significant cytotoxity and its possible mechanism involves altering tumor-related gene expression and regulating cell cycle process.

Establishment and gene expression profiling of LKB1 stable knockdown lung cancer cell line

Background Lung cancer is the leading cause of cancer-related death in China. Mutation analysis reveals that LKB1 inactivation is present in 30% of non-small-cell lung cancer （NSCLC）, indicating its role as a tumor suppressor. However, the molecular mechanism is still not clear. Our study attempted to establish LKB1 stable knockdown NSCLC cell line, detect alterations in gene expression and identify the genes regulated by LKBI. Methods LKB1 stable knockdown H1299 cell line was established using a lentiviral short hairpin RNA. To identify the knockdown effect, LKB1 mRNA and protein expression level were evaluated with quantitative real-time PCR and Western blotting. We treated the cell lines with 2-deoxyglucose to determine if LKB1 protein function was impacted. Gene microarray analysis was performed to detect the gene expression alterations in LKB1 stable knockdown H1299 cells. Results LKB1 mRNA and protein expression were significantly suppressed in LKB1 stable knockdown H1299 cell line. 2-DG treatment had little impact on the phosphorylation of AMPK, which is the downstream target of LKB1, indicating the loss of function of LKBI. The microarray data showed that LKB1 knockdown resulted in expression alterations of 1243 kinds of genes, including those involved in cell migration, cell proliferation and cell apoptosis. Conclusions The establishment of LKB1 stable knockdown H1299 cell line provides us with a great tool to investigate various genes regulated by LKB1 through microarray. The discovery of cell proliferation and migration-related genes regulated by LKB1 is critical for unraveling molecular mechanisms of LKBI＇s role in the development and metastasis of lung cancer.

Influence of 3D Microgrooves on C2C12 Cell Proliferation,Migration,Alignment,F-actin Protein Expression and Gene Expression

In this paper, we fabricated three kinds of 3D microgrooves with different depth on biocompatible poly（lactic-co-glycolic acid） （PLGA） substrate via combination of soft-lithography and melt-casting methods, and investigated in detail their influence on C2C12 cell behaviors. It is found that cell proliferation, migration, alignment, spatial distribution, F-actin protein expression and gene expression are all remarkably distinct on these microgrooved samples and the smooth control PLGA substrate. The associated underlying mechanisms were further analyzed and discussed using real-time living cell monitoring, confocal laser scanning microscopy and gene microarray. Our preliminary results suggested that 3D microstruc- ture could affect cell behaviors in a much more extensive manner than what we had understood before.

Human brain arteriovenous malformation:an analysis of differential expressed genes

Background:Much still remains unknown about the pathogenesis of brain arteriovenous malformations (AVMs).Previous studies have revealed the abnormal expression of various angiogenesis-related genes in AVMs.To further understand this disease,we sought to identify genes differently expressed in AVMs by means of the gene microarray technique.Methods:Nine AVMs specimen and nine samples of normal vessels are collected.Total RNA isolated from these specimen is hybridized with Oligonucleotide array and gene analysis was conducted.Analyzing data with the help of significance analysis of microarrays (SAM) and a free web-based molecular annotation system 3.0 (MAS 3.0).Results:The SAM method identify 37 gene significantly up-regulated and 10 genes down-regulated in AVMs.Conclusions:Among those genes,VACN,SPARK and ARHGAP18 seem to play a facilitating role during the genesis of AVMs.Multiple pathways,as MAPK pathway,may also be involved.

The impact of space environment on gene expression in Arabidopsis thaliana seedlings

One of the important questions in space biology is the mechanisms underlying plant responses to an outer space environment,i.e.,how gene expression is altered in space.In this study,the transcriptome of Arabidopsis thaliana seedlings was analyzed as a part of Germany SIMBOX(science in microgravity box)spaceflight experiment on Shenzhou 8 spacecraft.This experiment involved the following treatments:spaceflight with microgravity(Fμg),spaceflight with 1g centrifugal force(F 1g),and ground 1g control(G 1g).Gene chips were used to screen gene expression differences in Arabidopsis thaliana seedlings among these treatments.Microarray analysis revealed that 621 genes were differentially expressed in samples Fμg vs.G 1g,249 genes in samples F 1g vs.G 1g,and 368 genes in samples Fμg vs.F 1g.Gene ontology analysis indicated that the genes were involved in metabolism of stress response,gravitropic response,and DNA damage and repair,suggesting that plants adjust these metabolic pathways to space environmental stress,microgravity,and radiation.

Genome-wide gene expression analysis for target genes to differentiate patients with intestinal tuberculosis and Crohn’s disease and discriminative value of FOXP3 mRNA expression

Background and aims:Crohn’s disease(CD)and intestinal tuberculosis(ITB)are both chronic granulomatous conditions with similar phenotypic presentations.Hence,there is need for a biomarker to differentiate between both these two diseases.This study aimed at genome-wide gene expression analysis of colonic biopsies from confirmed cases of ITB and CD in comparison with controls.To evaluate the role of T regulatory cells,forkhead box P3(FOXP3)mRNA expression was quantified in serum as well as in colonic biopsies from patients with ITB and with the controls.Methods:Paired samples,including serum and colonic biopsies,were taken from 33 study subjects(CD,ITB and controls),and total RNA was extracted.Human whole genome gene expression microarray analysis was performed using the Illumina HumanWG-6 BeadChip Kit with six total RNA samples of the three groups in duplicates.Real-time PCR for FOXP3 mRNA expression was analyzed in serum samples and colonic biopsy samples(4-CD,5-ITB,4-controls).Results:In CD and ITB there was 1.5-fold upregulation of 92 and 382 genes and 1.5-fold downregulation of 91 and 256 genes,respectively.Peroxisome proliferators via the PPARc pathway were most significantly downregulated(P<0.005)in CD.Additionally,the IL4/5/6 signaling pathways and Toll-like receptor signaling pathway were identified as significantly differentially regulated(P<0.005)at>2-fold change.In ITB,the complement activation pathway,specifically the classical pathway,was the most significantly upregulated.FOXP3 mRNA expression was significantly elevated in colonic biopsies obtained from ITB patients as compared with CD cases(4.7062.21 vs 1.4860.31,P=0.016).Conclusions:FOXP3 mRNA expression in colonic mucosa could be a discriminatory marker between ITB and CD.Upregulation of the complement activation pathway in ITB suggests that pathogenetic mechanisms for ITB are similar to those of pulmonary tuberculosis.In CD,downregulation of PPARc was seen in colonic tissue,suggesting that restoration of PPARc-dependent anti-microbial barrier function may be a therapeutic target.

Gene Selection for Classifications Using Multiple PCA with Sparsity

A gene selection algorithm was developed using Multiple Principal Component Analysis with Sparsity （MSPCA）. The MSPCA algorithm is used to analyze normal and disease gene expression samples and to set these component Ioadings to zero if they are smaller than a threshold for sparse solutions. Next, genes with zero Ioadings across all samples （both normal and disease） are removed before extracting feature genes. Feature genes are genes that contribute differentially to variations in normal and disease samples and, thus, can be used for classification. The MSPCA is applied to three microarray datasets to select feature genes with a linear support vector machine to evaluate its performance. This method is compared with several previous gene selection results to show that this MSPCA gene selection algorithm has good classification accuracy and model stability.

Global expression profile of silkworm genes from larval to pupal stages： Toward a comprehensive understanding of sexual differences

Sexual dimorphism is a widespread phenomenon in many higher animals. The genes and gene networks that underlie sex differences are poorly understood. Using microarray data we analyzed sex-related differences in the global expression profiles of silkworm genes from larval to pupal stages. Sex-biased genes could be divided into three clusters. Cluster 1 contained 932 genes that showed a female-biased expression trend at first and a male-biased trend afterward. Cluster 2 included 283 male-biased genes. Cluster 3 was comprised of 497 female-biased genes that were expressed during the late pupal stage. Cluster 1 genes were found to be related closely to cuticle proteins, hormones, binding proteins, enzyme regulators, structural proteins, transcription regulators and so on. Several genes in clusters 2 and 3 were associated with spermatogenesis and oogenesis, respectively. The chromosomal distribution of sex-biased genes showed evidence of chromosomal enrichment. In particular a large number of the silkworms＇ male-biased genes are located on the Z chromosome. These results provide new insights into the molecular differences that dictate sexual dimorphism in the silkworm.