MicroRNAs (miRNAs) are a family of ~22 nucleotide small RNA molecules that regulate gene expression by fully or partially binding to their complementary sequences. Recently, a large number of miRNAs and their expression patterns have been identified in various species. However, to date no miRNAs have been reported to modulate muscle development in beef cattle.
Total RNAs from the Chinese Qinchuan bovine longissimus thoracis at fetal and adult stages were used to construct small RNA libraries for Solexa SBS technology sequencing. A total of 15,454,182 clean reads were obtained from the fetal bovine library and 13,558,164 clean reads from the adult bovine library. In total, 521 miRNAs including 104 novel miRNA candidates were identified. Furthermore, the nucleotide bias, base edit and family of the known miRNAs were also analyzed. Based on stem-loop qPCR, 25 high-read miRNAs were detected, and the results showed that bta-miRNA-206, miRNA-1, miRNA-133, miRNAn12, and miRNAn17 were highly expressed in muscle-related tissue or organs, suggesting that these miRNAs may play a role in the development of bovine muscle tissues.
This study confirmed the authenticity of 417 known miRNAs, discovered 104 novel miRNAs in bos taurus, and identified five muscle-specific miRNAs. The identification of novel miRNAs significantly expanded the repertoire of bovine miRNAs and could contribute to further studies on the muscle development of cattle.
Bovine; Deep sequencing technology; microRNA; Muscle; Proliferation; Differentiation
An important controversy in the relationship between beef tenderness and muscle characteristics including biochemical traits exists among meat researchers. The aim of this study is to explain variability in meat tenderness using muscle characteristics and biochemical traits available in the Integrated and Functional Biology of Beef (BIF-Beef) database. The BIF-Beef data warehouse contains characteristic measurements from animal, muscle, carcass, and meat quality derived from numerous experiments. We created three classes for tenderness (high, medium, and low) based on trained taste panel tenderness scores of all meat samples consumed (4,366 observations from 40 different experiments). For each tenderness class, the corresponding means for the mechanical characteristics, muscle fibre type, collagen content, and biochemical traits which may influence tenderness of the muscles were calculated.
Our results indicated that lower shear force values were associated with more tender meat. In addition, muscles in the highest tenderness cluster had the lowest total and insoluble collagen contents, the highest mitochondrial enzyme activity (isocitrate dehydrogenase), the highest proportion of slow oxidative muscle fibres, the lowest proportion of fast-glycolytic muscle fibres, and the lowest average muscle fibre cross-sectional area. Results were confirmed by correlation analyses, and differences between muscle types in terms of biochemical characteristics and tenderness score were evidenced by Principal Component Analysis (PCA). When the cluster analysis was repeated using only muscle samples from m. Longissimus thoracis (LT), the results were similar; only contrasting previous results by maintaining a relatively constant fibre-type composition between all three tenderness classes.
Our results show that increased meat tenderness is related to lower shear forces, lower insoluble collagen and total collagen content, lower cross-sectional area of fibres, and an overall fibre type composition displaying more oxidative fibres than glycolytic fibres.
Tenderness; Beef; Meta-analysis; Muscle biochemistry
Marbling (intramuscular fat) is a valuable trait that impacts on meat quality and an important factor determining price of beef in the Korean beef market. Animals that are destined for this high marbling market are fed a high concentrate ration for approximately 30 months in the Korean finishing farms. However, this feeding strategy leads to inefficiencies and excessive fat production. This study aimed to identify candidate genes and pathways associated with intramuscular fat deposition on highly divergent marbling phenotypes in adult Hanwoo cattle.
Bovine genome array analysis was conducted to detect differentially expressed genes (DEGs) in m. longissimus with divergent marbling phenotype (marbling score 2 to 7). Three data-processing methods (MAS5.0, GCRMA and RMA) were used to test for differential expression (DE). Statistical analysis identified 21 significant transcripts from at least two data-processing methods (P < 0.01). All 21 differentially expressed genes were validated by real-time PCR. Results showed a high concordance in the gene expression fold change between the microarrays and the real time PCR data. Gene Ontology (GO) and pathway analysis demonstrated that some genes (ADAMTS4, CYP51A and SQLE) over expressed in high marbled animals are involved in a protein catabolic process and a cholesterol biosynthesis process. In addition, pathway analysis also revealed that ADAMTS4 is activated by three regulators (IL-17A, TNFα and TGFβ1). QRT-PCR was used to investigate gene expression of these regulators in muscle with divergent intramuscular fat contents. The results demonstrate that ADAMTS4 and TGFβ1 are associated with increasing marbling fat. An ADAMTS4/TGFβ1 pathway seems to be associated with the phenotypic differences between high and low marbled groups.
Marbling differences are possibly a function of complex signaling pathway interactions between muscle and fat. These results suggest that ADAMTS4, which is involved in connective tissue degradation, could play a role in an important biological pathway for building up marbling in cattle. Moreover, ADAMTS4 and TGFβ1could potentially be used as an early biological marker for marbling fat content in the early stages of growth.
Beef is one of the leading sources of protein, B vitamins, iron, and zinc in human food. Beef palatability is based on three general criteria: tenderness, juiciness, and flavor, of which tenderness is thought to be the most important factor. In this study, we found that beef tenderness, measured by the Warner-Bratzler shear force (WBSF), was dramatically increased by acute stress. Microarray analysis and qPCR identified a variety of genes that were differentially expressed. Pathway analysis showed that these genes were involved in immune response and regulation of metabolism process as activators or repressors. Further analysis identified that these changes may be related with CpG methylation of several genes. Therefore, the results from this study provide an enhanced understanding of the mechanisms that genetic and epigenetic regulations control meat quality and beef tenderness.
An important variability of contractile and metabolic properties between muscles has been highlighted. In the literature, the majority of studies on beef sensorial quality concerns M. longissimus thoracis. M. rectus abdominis (RA) is easy to sample without huge carcass depreciation and may appear as an alternative to M. longissimus thoracis for fast and routine physicochemical analysis. It was considered interesting to assess the muscle fibres of M. rectus abdominis in comparison with M. longissimus thoracis (LT) and M. triceps brachii (TB) on the basis of metabolic and contractile properties, area and myosin heavy chain isoforms (MyHC) proportions. Immuno-histochemical, histochemical, histological and enzymological techniques were used. This research concerned two populations of Charolais cattle: RA was compared to TB in a population of 19 steers while RA was compared to LT in a population of 153 heifers.
RA muscle had higher mean fibre areas (3350 μm2 vs 2142 to 2639 μm2) than the two other muscles. In RA muscle, the slow-oxidative fibres were the largest (3957 μm2) and the fast-glycolytic the smallest (2868 μm2). The reverse was observed in TB muscle (1725 and 2436 μm2 respectively). In RA muscle, the distinction between fast-oxidative-glycolytic and fast-glycolytic fibres appeared difficult or impossible to establish, unlike in the other muscles. Consequently the classification based on ATPase and SDH activities seemed inappropriate, since the FOG fibres presented rather low SDH activity in this muscle in comparison to the other muscles of the carcass. RA muscle had a higher proportion of I fibres than TB and LT muscles, balanced by a lower proportion either of IIX fibres (in comparison to TB muscle) or of IIA fibres (in comparison to LT muscle). However, both oxidative and glycolytic enzyme activities were lower in RA than in TB muscle, although the LDH/ICDH ratio was higher in RA muscle (522 vs 340). Oxidative enzyme activities were higher in RA than in LT muscle, whereas glycolytic enzyme activity was lower. In RA muscle, contractile and metabolic properties appeared to be less well-correlated than in the two other muscles.
RA muscle has some particularities in comparison to the LT and TB muscles, especially concerning the unusual large cross-section surface of SO fibres and the very low oxidative activity of intermediate IIA fibres.
Serotype O157:H7 Escherichia coli strains from several different bovine and meat (beef) sources were studied to determine the diversity of their virulence properties and to compare their plasmid characteristics. Eighteen strains from cattle feces, 2 from water buffalo feces, 3 from beef samples, and 2 from feces of human hemolytic uremic syndrome cases were examined. All of these strains hybridized with the CVD419 DNA probe which identifies serotype O157:H7 and many other serotypes of verocytotoxin-producing E. coli. Of 15 bovine strains that hybridized with two verocytotoxin DNA probes, 8 hybridized with both verocytotoxin 1 (VT1) and VT2 probes, 5 hybridized with only the VT2 probe, and 2 hybridized with only the VT1 probe. This distribution was similar to that reported for O157:H7 E. coli isolated from humans. All three beef isolates hybridized with both VT1 and VT2 probes. All strains that hybridized with the VT probes were positive in the verocytotoxin assay, and all probe-negative strains were negative in the assay. All the strains possessed large plasmids with molecular sizes ranging from 53 to 64 MDa. Fifteen of the 20 cattle and water buffalo strains had one or more additional small plasmids. Restriction patterns resulting from HindIII, SmaI, and BamHI digestions of the large plasmids were used to compare all possible pairs of five different single plasmid-bearing strains from different countries (Egypt, England, and the United States). The restriction patterns of these strains were distinct, and the mean coefficients of similarity for these comparisons ranged from 71 to 91%, indicating a moderate degree of genetic diversity. This diversity and the presence of multiple plasmids in many bovine and human O157:H7 strains reinforce the usefulness of plasmid analysis in future studies. Only four of the 20 bovine strains and 1 of the 3 beef strains possessed the capability for adherence to HEp-2 and Intestine 407 cells in the presence of mannose, indicating that in vitro expression of localized adherence is not a universal property of O157:H7 strains of bovine origin.
Cold carcass weight (CW) and longissimus muscle area (EMA) are the major quantitative traits in beef cattle. In this study, we found several polymorphisms of growth hormone-releasing hormone (GHRH) gene and examined the association of polymorphisms with carcass traits (CW and EMA) in Korean native cattle (Hanwoo).
By direct DNA sequencing in 24 unrelated Korean cattle, we identified 12 single nucleotide polymorphisms within the 9 kb full gene region, including the 1.5 kb promoter region. Among them, six polymorphic sites were selected for genotyping in our beef cattle (n = 428) and five marker haplotypes (frequency > 0.1) were identified. Statistical analysis revealed that -4241A>T showed significant associations with CW and EMA.
Our findings suggest that polymorphisms in GHRH might be one of the important genetic factors that influence carcass yield in beef cattle. Sequence variation/haplotype information identified in this study would provide valuable information for the production of a commercial line of beef cattle.
In this manuscript, we describe the generation of a gene library for the expression of HSP110/HSPH, HSP70/HSPA and HSP40/DNAJ members. First, the heat shock protein (HSP) genes were collected from the gene databases and the gene families were analyzed for expression patterns, heat inducibility, subcellular localization, and protein homology using several bioinformatics approaches. These results can be used as a working draft model until data are confirmed by experimental approaches. In addition, we describe the generation of a HSPA/DNAJ overexpression library and tested the effect of different fusion tags on HSPA and DNAJ members using different techniques for measuring chaperone activity. These results show that we have cloned a high-quality heat shock protein expression library containing most members from the HSPH, HSPA, DNAJA and DNAJB families which will be useful for the chaperone community to unravel the function of the highly diverse family of human molecular chaperones.
HspH; HspA; DnaJ; Hsp110; Hsp70; Hsp40; Human chaperones; Bio-informatics
The purpose of this study was to evaluate the effects of eight single nucleotide polymorphisms (SNP), previously associated with meat and milk quality traits in cattle, in a population of 443 commercial Aberdeen Angus-cross beef cattle. The eight SNP, which were located within five genes: μ-calpain (CAPN1), calpastatin (CAST), leptin (LEP), growth hormone receptor (GHR) and acylCoA:diacylglycerol acyltransferase 1 (DGAT1), are included in various commercial tests for tenderness, fatness, carcass composition and milk yield/quality.
A total of 27 traits were examined, 19 relating to carcass quality, such as carcass weight and fatness, one mechanical measure of tenderness, and the remaining seven were sensory traits, such as flavour and tenderness, assessed by a taste panel.
An SNP in the CAPN1 gene, CAPN316, was significantly associated with tenderness measured by both the tenderometer and the taste panel as well as the weight of the hindquarter, where animals inheriting the CC genotype had more tender meat and heavier hindquarters. An SNP in the leptin gene, UASMS2, significantly affected overall liking, where animals with the TT genotype were assigned higher scores by the panellists. The SNP in the GHR gene was significantly associated with odour, where animals inheriting the AA genotype produced steaks with an intense odour when compared with the other genotypes. Finally, the SNP in the DGAT1 gene was associated with sirloin weight after maturation and fat depth surrounding the sirloin, with animals inheriting the AA genotype having heavier sirloins and more fat.
The results of this study confirm some previously documented associations. Furthermore, novel associations have been identified which, following validation in other populations, could be incorporated into breeding programmes to improve meat quality.
Marbling defined by the amount and distribution of intramuscular fat is an economically important trait of beef cattle in Japan. We have recently reported that single nucleotide polymorphisms (SNPs) in the endothelial differentiation, sphingolipid G-protein-coupled receptor, 1 (EDG1) gene were associated with marbling in Japanese Black beef cattle. As well as EDG1, the titin (TTN) gene, involved in myofibrillogenesis, has been previously shown to possess expression difference in musculus longissimus muscle between low-marbled and high-marbled steer groups, and to be located within genomic region of a quantitative trait locus for marbling. Thus TTN was considered as a positional functional candidate for the gene responsible for marbling. In this study, we explored SNP in TTN and analyzed association of the SNP with marbling.
A SNP in the promoter region of TTN, referred to as g.231054C>T, was the only difference detected between high- and low-marbled steer groups. The SNP was associated with marbling in 3 experiments using 101 sires (P = 0.004), 848 paternal half-sib progeny steers from 5 sires heterozygous for the g.231054C>T (P = 0.046), and 820 paternal half-sib progeny steers from 3 sires homozygous for C allele at the g.231054C>T (P = 0.051), in Japanese Black beef cattle. The effect of genotypes of the SNP on subcutaneous fat thickness was not statistically significant (P > 0.05).
These findings suggest that in addition to the EDG1 SNPs, the TTN SNP polymorphism is associated with marbling and may be useful for effective marker-assisted selection to increase the levels of marbling in Japanese Black beef cattle. Further replicate studies will be needed to confirm the allelic association observed here, and to expand the results to evaluate all possible genotypic combinations of alleles.
The intramuscular fat deposition and the fatty acid profiles of beef affect meat quality. High proportions of unsaturated fatty acids are related to beef flavor and are beneficial for the nutritional value of meat. Moreover, a variety of clinical and epidemiologic studies showed that particularly long-chain omega-3 fatty acids from animal sources have a positive impact on human health and disease.
To screen for genetic factors affecting fatty acid profiles in beef, we initially performed a microsatellite-based genome scan in a F2 Charolais × German Holstein resource population and identified a quantitative trait locus (QTL) for fatty acid composition in a region on bovine chromosome 27 where previously QTL affecting marbling score had been detected in beef cattle populations. The long-chain acyl-CoA synthetase 1 (ACSL1) gene was identified as the most plausible functional and positional candidate gene in the QTL interval due to its direct impact on fatty acid metabolism and its position in the QTL interval. ACSL1 is necessary for synthesis of long-chain acyl-CoA esters, fatty acid degradation and phospholipid remodeling. We validated the genomic annotation of the bovine ACSL1 gene by in silico comparative sequence analysis and experimental verification. Re-sequencing of the complete coding, exon-flanking intronic sequences, 3' untranslated region (3'UTR) and partial promoter region of the ACSL1 gene revealed three synonymous mutations in exons 6, 7, and 20, six noncoding intronic gene variants, six polymorphisms in the promoter region, and four variants in the 3' UTR region. The association analysis identified the gene variant in intron 5 of the ACSL1 gene (c.481-233A>G) to be significantly associated with the relative content of distinct fractions and ratios of fatty acids (e.g., n-3 fatty acids, polyunsaturated, n-3 long-chain polyunsaturated fatty acids, trans vaccenic acid) in skeletal muscle. A tentative association of the ACSL1 gene variant with intramuscular fat content indicated that an indirect effect on fatty acid composition via modulation of total fat content of skeletal muscle cannot be excluded.
The initial QTL analysis suggested the ACSL1 gene as a positional and functional candidate gene for fatty acid composition in bovine skeletal muscle. The findings of subsequent association analyses indicate that ACSL1 or a separate gene in close proximity might play a functional role in mediating the lipid composition of beef.
Genetic analysis of transcriptional profiles is a promising approach for identifying and dissecting the genetics of complex traits like meat performance. Accordingly, expression levels obtained by microarray analysis were taken as phenotypes in a linkage analysis to map eQTL. Moreover, expression levels were correlated with traits related to meat quality and principle components with high loadings of these traits. By using an up-to-date annotation and localization of the respective probe-sets, the integration of eQTL mapping data and information of trait correlated expression finally served to point to candidate genes for meat quality traits.
Genome-wide transcriptional profiles of M. longissimus dorsi RNAs samples of 74 F2 animals of a pig resource population revealed 11,457 probe-sets representing genes expressed in the muscle. Linkage analysis of expression levels of these probe-sets provided 9,180 eQTL at the suggestive significance threshold of LOD > 2. We mapped 653 eQTL on the same chromosome as the corresponding gene and these were designated as 'putative cis-eQTL'. In order to link eQTL to the traits of interest, probe-sets were addressed with relative transcript abundances that showed correlation with meat quality traits at p ≤ 0.05. Out of the 653 'putative cis-eQTL', 262 transcripts were correlated with at least one meat quality trait. Furthermore, association of expression levels with composite traits with high loadings for meat quality traits generated by principle component analysis were taken into account leading to a list of 85 genes exhibiting cis-eQTL and trait dependent expression.
Holistic expression profiling was integrated with QTL analysis for meat quality traits. Correlations between transcript abundance and meat quality traits, combined with genetic positional information of eQTL allowed us to prioritise candidate genes for further study.
This study was conducted to determine the nutritional characteristics of horsemeat and bone meal in comparison with those of beef and pork presented by Dietary Reference Intakes For Koreans. Longissimus muscle and large metacarpal bone samples were collected from 20 fattened Jeju horses. Muscle samples were subjected to proximate analysis, assays for fatty acid profile and minerals, and bone samples to mineral assays. Horsemeat had similar levels of protein (21.1 vs 21.0 or 21.1%) and lower levels of fat (6.0 vs 14.1 or 16.1%) compared with beef or pork, respectively. Horsemeat had much higher levels of palmitoleic (8.2 vs 4.4 or 3.3%) and α-linolenic (1.4 vs 0.1 or 0.6%) acids than beef or pork, respectively. Linoleic acid was much higher in horsemeat (11.1%) and pork (10.1%) than in beef (1.6%). PUFA:SFA and n-6:n-3 ratios in horsemeat were 0.29 and 10.2, respectively. There were no big differences in mineral contents between horsemeat, beef and pork. For daily recommended mineral intakes of male adults (Dietary Reference Intakes For Koreans), phosphorus, sodium, potassium, iron, zinc and copper can be provided up to 24, 2.5, 6.7, 21, 26 and 40%, respectively, by 100 g raw horsemeat, but calcium and manganese levels are negligible. Horse cannon bone had much higher mineral contents especially in calcium (10,193 mg/100 g), phosphorus (5,874 mg/100 g) and copper (0.79 mg/100 g). Thiamin, riboflavin, niacin and retinol contents were 0.20, 0.21, 1.65 mg/100 g and 30 µg/100 g, respectively. But ascorbic acid and beta-carotene were not detected. Our data demonstrated that higher levels of palmitoleic and α-linolenic acid in horsemeat than in beef and pork may be beneficial for human health. Horsemeat and bone meal are a good source of some minerals and vitamins.
Horsemeat; beef; pork; bone; fatty acid; mineral
The possible origin of beef contamination and genetic diversity of Escherichia coli populations in beef cattle, on carcasses and ground beef, was examined by using random amplification of polymorphic DNA (RAPD) and PCR-restriction fragment length polymorphism (PCR-RFLP) analysis of the fliC gene. E. coli was recovered from the feces of 10 beef cattle during pasture grazing and feedlot finishing and from hides, carcasses, and ground beef after slaughter. The 1,403 E. coli isolates (855 fecal, 320 hide, 153 carcass, and 75 ground beef) were grouped into 121 genetic subtypes by using the RAPD method. Some of the genetic subtypes in cattle feces were also recovered from hides, prechilled carcasses, chilled carcasses, and ground beef. E. coli genetic subtypes were shared among cattle at all sample times, but a number of transient types were unique to individual animals. The genetic diversity of the E. coli population changed over time within individual animals grazing on pasture and in the feedlot. Isolates from one animal (59 fecal, 30 hide, 19 carcass, and 12 ground beef) were characterized by the PCR-RFLP analysis of the fliC gene and were grouped into eight genotypes. There was good agreement between the results obtained with the RAPD and PCR-RFLP techniques. In conclusion, the E. coli contaminating meat can originate from cattle feces, and the E. coli population in beef cattle was highly diverse. Also, genetic subtypes can be shared among animals or can be unique to an animal, and they are constantly changing.
The transversus thoracis muscle is a thin muscular layer on the inner surface of the anterior thoracic wall that is always in concern during harvesting of the internal thoracic artery. Because the muscle is poorly described in the surgical literature, the aim of the present study is to examine in details its variations.
The data was obtained at standard autopsies of 120 Caucasian subjects (Bulgarians) of both sexes (97 males and 23 females), ranging in age from 18 to 91 years (mean age 52.8 ± 17.8 years). The transversus thoracis morphology was thoroughly examined on the inner surface of the chest plates collected after routine incisions.
An overall examination revealed that in majority of cases the transversus thoracis slips formed a complete muscular layer (left - 75.8%, right - 83.3%) or some of the slips (left - 22.5%, right - 15%) or all of them (left - 1.7%, right - 1.7%) were quite separated. Rarely (left - 3.3%, right - 5.8%), some fibrous slips of the transversus thoracis were noted. In 55.8% of the cases there was left/right muscle symmetry; 44.2% of the muscles were asymmetrical. Most commonly, the highest muscle attachment was to the second (left - 53.3%, right - 37.5%) or third rib (left - 29.2%, right - 46.7%). The sixth rib was the most common lowest attachment (left - 94.2%, right - 89.2%). Most frequently, the muscle was composed of four (left - 31.7%, right - 44.2%) or fifth slips (left - 53.3%, right - 40.8%).
This study provides detailed basic information on the variety of the transversus thoracic muscle. It also defines the range of the clearly visible, uncovered by the muscle part of the internal thoracic artery and the completeness of the muscular layer over it. The knowledge of these peculiar muscle-arterial relations would definitely be beneficial to cardiac surgeon in performing fast and safe arterial harvesting.
Meat quality is determined by properties such as carcass color, tenderness and drip loss. These properties are closely associated with meat composition, which includes the types of muscle fiber and content of intramuscular fat (IMF). Muscle fibers are the main contributors to meat mass, while IMF not only contributes to the sensory properties but also to the plethora of physical, chemical and technological properties of meat. However, little is known about the molecular mechanisms that determine meat composition in different pig breeds. In this report we show that Jinhua pigs, a Chinese breed, contains much higher levels of IMF than do Landrace pigs, a Danish breed. We analyzed global gene expression profiles in the longissimus dorsi muscles in Jinhua and Landrace breeds at the ages of 30, 90 and 150 days. Cross-comparison analysis revealed that genes that regulate fatty acid biosynthesis (e.g., fatty acid synthase and stearoyl-CoA desaturase) are expressed at higher levels in Jinhua pigs whereas those that regulate myogenesis (e.g., myogenic factor 6 and forkhead box O1) are expressed at higher levels in Landrace pigs. Among those genes which are highly expressed in Jinhua pigs at 90 days (d90), we identified a novel gene porcine FLJ36031 (pFLJ), which functions as a positive regulator of fat deposition in cultured intramuscular adipocytes. In summary, our data showed that the up-regulation of fatty acid biosynthesis regulatory genes such as pFLJ and myogenesis inhibitory genes such as myostatin in the longissimus dorsi muscles of Jinhua pigs could explain why this local breed produces meat with high levels of IMF.
Affymetrix GeneChip arrays are widely used for transcriptomic studies in a diverse range of species. Each gene is represented on a GeneChip array by a probe-set, consisting of up to 16 probe-pairs. Signal intensities across probe-pairs within a probe-set vary in part due to different physical hybridisation characteristics of individual probes with their target labelled transcripts. We have previously developed a technique to study the transcriptomes of heterologous species based on hybridising genomic DNA (gDNA) to a GeneChip array designed for a different species, and subsequently using only those probes with good homology.
Here we have investigated the effects of hybridising homologous species gDNA to study the transcriptomes of species for which the arrays have been designed. Genomic DNA from Arabidopsis thaliana and rice (Oryza sativa) were hybridised to the Affymetrix Arabidopsis ATH1 and Rice Genome GeneChip arrays respectively. Probe selection based on gDNA hybridisation intensity increased the number of genes identified as significantly differentially expressed in two published studies of Arabidopsis development, and optimised the analysis of technical replicates obtained from pooled samples of RNA from rice.
This mixed physical and bioinformatics approach can be used to optimise estimates of gene expression when using GeneChip arrays.
Specific DNA probes were used to identify Shiga-like toxin I (SLT I)- and SLT II-producing Escherichia coli in vegetables, meats, cattle, and farm animals in Thailand. SLT-producing E. coli was isolated from 9% of market beef specimens, from 8 to 28% of fresh beef specimens at slaughterhouses, and from 11 to 84% of fecal specimens from cattle. Animals were frequently infected with several different SLT-producing E. coli types that hybridized with either the SLT I, SLT II, or both SLT probes. Of 119 SLT-producing E. coli isolates, 24% hybridized with the SLT I probe, 31% hybridized with the SLT II probe, and 44% hybridized with both SLT probes. The enterohemorrhagic E. coli plasmid probe hybridized with 64% (68 of 106) of SLT-producing E. coli isolates from food and cattle and with 8% (17 of 201) of E. coli isolates from pigs. No SLT-producing E. coli was detected in pigs. Seventy-six percent (26 of 34) of E. coli isolates that hybridized with the SLT II probe were cytotoxic to Vero but not to HeLa cells, suggesting that they produced the variant of SLT II. The high prevalence of SLT-producing E. coli in beef-producing animals suggests that exposure to animals and eating beef may pose a health risk for acquiring enterohemorrhagic E. coli infections in Thailand.
The fat components of red meat products have been of interest to researchers due to the health aspects of excess fat consumption by humans. We hypothesized that differences in protein expression have an impact on adipose tissue formation during beef cattle development and growth. Therefore, in this study we evaluated the differences in the discernable proteome of subcutaneous adipose tissues of 35 beef crossbred steers [Charolais × Red Angus (CHAR) (n = 13) and Hereford × Angus (HEAN) (n = 22)] with different back fat (BF) thicknesses. The goal was to identify specific protein markers that could be associated with adipose tissue formation in beef cows.
Approximately 541-580 protein spots were detected and compared in each crossbred group, and 33 and 36 protein spots showed expression differences between tissues with high and low BF thicknesses from HEAN and CHAR crossbed, respectively. The annexin 1 protein was highly expressed in both crossbred steers that had a higher BF thickness (p < 0.05) and this was further validated by a western blot analysis. In 13 tissues of CHAR animals and 22 tissues of HEAN animals, the relative expression of annexin 1 was significantly different (p < 0.05) between tissues with high and low BF thicknesses.
The increased expression of annexin 1 protein has been found to be associated with higher BF thickness in both crossbred steers. This result lays the foundation for future studies to develop the protein marker for assessing animals with different BF thickness.
High-density oligonucleotide (oligo) arrays are a powerful tool for transcript profiling. Arrays based on GeneChip® technology are amongst the most widely used, although GeneChip® arrays are currently available for only a small number of plant and animal species. Thus, we have developed a method to improve the sensitivity of high-density oligonucleotide arrays when applied to heterologous species and tested the method by analysing the transcriptome of Brassica oleracea L., a species for which no GeneChip® array is available, using a GeneChip® array designed for Arabidopsis thaliana (L.) Heynh. Genomic DNA from B. oleracea was labelled and hybridised to the ATH1-121501 GeneChip® array. Arabidopsis thaliana probe-pairs that hybridised to the B. oleracea genomic DNA on the basis of the perfect-match (PM) probe signal were then selected for subsequent B. oleracea transcriptome analysis using a .cel file parser script to generate probe mask files. The transcriptional response of B. oleracea to a mineral nutrient (phosphorus; P) stress was quantified using probe mask files generated for a wide range of gDNA hybridisation intensity thresholds. An example probe mask file generated with a gDNA hybridisation intensity threshold of 400 removed > 68 % of the available PM probes from the analysis but retained >96 % of available A. thaliana probe-sets. Ninety-nine of these genes were then identified as significantly regulated under P stress in B. oleracea, including the homologues of P stress responsive genes in A. thaliana. Increasing the gDNA hybridisation intensity thresholds up to 500 for probe-selection increased the sensitivity of the GeneChip® array to detect regulation of gene expression in B. oleracea under P stress by up to 13-fold. Our open-source software to create probe mask files is freely available and may be used to facilitate transcriptomic analyses of a wide range of plant and animal species in the absence of custom arrays.
Arabidopsis thaliana; Brassica oleracea; Brassicaceae; cross-species; microarray; oligonucleotide; phosphate; phosphorus; plant nutrition; probe masking; transcriptomics
The conjugated linoleic acid (CLA) content of beef can be increased by supplementing appropriate beef cattle diets with vegetable oil or oil seed. Yet the effect of consumption of such beef on adipose tissue characteristics is unclear, thus the study was conducted to compare adipose tissue responses of rats to diets containing beef from steers either not provided or provided the oil supplements to alter CLA composition of the fat in muscle.
Effects of feeding synthetic (industrial hydrogenation) CLA or CLA from beef on growth and adipose tissue responses of weanling, male, Wistar rats (n = 56; 14 per treatment diet) were investigated in a completely randomized design experiment. Diets were: control (CON) diet containing casein and soybean oil, synthetic CLA (SCLA) diet; where 1.69% synthetic CLA replaced soybean oil, two beef-diets; CONM and CLAM, containing freeze dried beef from steers either not fed or fed 14% sunflower seeds to increase CLA content of beef. Diets were isonitrogenous (20% protein) and isocaloric. Rat weights and ad libitum intakes were recorded every 2 wk. After 9 wk, rats were fasted for 24 h, blood sampled by heart puncture, sacrificed, tissue and organs were harvested and weights recorded. The adipose tissue responses with regard to cellularity and fatty acid compositions of retroperitoneal and inguinal adipose tissue were determined.
Body weights and gains were comparable, but organ weights as percent of body weight were greater for rats fed SCLA than CONM. Fasting blood glucose concentration was lower (p < 0.01) in rats fed SCLA than those fed CONM or CLAM. Retroperitoneal and inguinal fat weights, as percent of body weight were greater (p < 0.01) in rats fed CONM or CLAM than those fed CON or SCLA diets. Adipocyte numbers were least in retroperitoneal tissue of rats fed SCLA, while inguinal tissue cell density and total number were lower (p = 0.02) in rats fed CLAM (7.26 × 107 cells/g and 8.03 × 108 cells) than those fed CONM (28.88 × 107 cells/g and 32.05 × 108 cells, respectively).
Study suggests that dietary CLA either as synthetic or high CLA-beef may alter adipose tissue characteristics by decreasing the number of adipocytes and by decreasing the size of the tissue.
Gene expression microarray experiments are expensive to conduct and guidelines for acceptable quality control at intermediate steps before and after the samples are hybridised to chips are vague. We conducted an experiment hybridising RNA from human brain to 117 U133A Affymetrix GeneChips and used these data to explore the relationship between 4 pre-chip variables and 22 post-chip outcomes and quality control measures.
We found that the pre-chip variables were significantly correlated with each other but that this correlation was strongest between measures of RNA quality and cRNA yield. Post-mortem interval was negatively correlated with these variables. Four principal components, reflecting array outliers, array adjustment, hybridisation noise and RNA integrity, explain about 75% of the total post-chip measure variability. Two significant canonical correlations existed between the pre-chip and post-chip variables, derived from MAS 5.0, dChip and the Bioconductor packages affy and affyPLM. The strongest (CANCOR 0.838, p < 0.0001) correlated RNA integrity and yield with post chip quality control (QC) measures indexing 3'/5' RNA ratios, bias or scaling of the chip and scaling of the variability of the signal across the chip. Post-mortem interval was relatively unimportant. We also found that the RNA integrity number (RIN) could be moderately well predicted by post-chip measures B_ACTIN35, GAPDH35 and SF.
We have found that the post-chip variables having the strongest association with quantities measurable before hybridisation are those reflecting RNA integrity. Other aspects of quality, such as noise measures (reflecting the execution of the assay) or measures reflecting data quality (outlier status and array adjustment variables) are not well predicted by the variables we were able to determine ahead of time. There could be other variables measurable pre-hybridisation which may be better associated with expression data quality measures. Uncovering such connections could create savings on costly microarray experiments by eliminating poor samples before hybridisation.
miRNAs are a class of small, single-stranded, non-coding RNAs that perform post-transcriptional repression of target genes by binding to 3’ untranslated regions. Research has found that miRNAs involved in the regulation of many metabolic processes. Here we uncovered that the beef quality of Angus cattle sharply diversified after acute stress. By performing miRNA microarray analysis, 13 miRNAs were significantly differentially expressed in stressed group compared to control group. Using a bioinformatics method, 135 protein-coding genes were predicted as the targets of significant differentially expressed miRNAs. Gene Ontology (GO) term and Ingenuity Pathway Analysis (IPA) mined that these target genes involved in some important pathways, which may have impact on meat quality and beef tenderness.
miRNA; Bovine; Beef tenderness; Stress
Marbling defined by the amount and distribution of intramuscular fat, so-called Shimofuri, is an economically important trait of beef cattle in Japan. The c17-25 expressed sequence tag (EST) has been previously shown to possess expression difference in musculus longissimus muscle between low-marbled and high-marbled steer groups, and to be located within genomic region of a quantitative trait locus for marbling. Thus, the akirin 2 (AKIRIN2) gene containing the c17-25 EST sequence was considered as a positional functional candidate for the gene responsible for marbling. In this study, we explored single nucleotide polymorphism (SNP) in the AKIRIN2 and analyzed association of the SNP with marbling.
A SNP in the 3' untranslated region of the AKIRIN2, referred to as c.*188G>A, was the only difference detected between high- and low-marbled steer groups. The SNP was associated with marbling in 3 experiments using 100 sires (P = 0.041), 753 paternal half-sib progeny steers from 4 sires heterozygous for the c.*188G>A (P = 0.005), and 730 paternal half-sib progeny steers from 3 sires homozygous for the A allele at the c.*188G>A (P = 0.047), in Japanese Black beef cattle. The effect of genotypes of the SNP on subcutaneous fat thickness was not statistically significant (P > 0.05).
These findings suggest that the AKIRIN2 SNP polymorphism is associated with marbling and may be useful for effective marker-assisted selection to increase the levels of marbling in Japanese Black beef cattle.
Presence of Mycobacterium avium subsp. paratuberculosis (MAP) in beef has been reported as a public health concern because asymptomatically infected cattle may contain MAP in tissues that are used for human consumption. Associations between MAP carcasses contamination and animal characteristics such as age, breed, production type, and carcass classification were assessed. Cheek muscles from 501 carcasses were sampled cross-sectionally at a Danish abattoir and tested for presence of viable MAP and MAP DNA by bacterial culture and IS900 realtime PCR, respectively. Cheek muscle tissues from carcasses of two dairy cows were positive by culture whereas 4% of the animals were estimated with ≥10 CFU/gram muscle based on realtime PCR. Age was found to be associated with carcass contamination with MAP. The observed viable MAP prevalence in beef carcasses was low. However, detection of MAP and MAP DNA in muscle tissues suggested that bacteremia occurred in slaughtered cattle.