Molecular Biology Reports

TP53 is a tumour suppressor gene. Its inactivation plays a significant role in the molecular pathology of cancers. TP53 germline mutations increase the risk of developing multiple primary cancers. However, the role of alterations in TP53 germline DNA in head and neck cancers (HNCs) is not well-established. HNCs comprise one of the most frequent cancers in South Asia. The present discovery study reports the investigation of germline variations in the TP53 gene in a cohort of 30 HNC patients from Karachi, Pakistan. Blood samples were collected and genomic DNA was extracted from white blood cells. TP53 has 11 exons, where exon 1 is not transcribed. After quality control of DNA, amplification of seven selected exons along with their splice sites, two intronic regions (introns 2-3 and 3-4), and 3UTR were carried out. Sanger sequencing was carried out in order to identify germline variations. Comparison with wild type sequence revealed rs1642785 G>C (intron 2-3) variation in 63.2%, PIN3 duplication (rs17878362) in intron 3-4 in 94.7%, and rs1042522 G>C in exon 4 (p.R72P) in 66.6% of the cases. In 3UTR, 13.4% of the analyzed cases carried either one of two variants, i.e., 17:7669567_8delCA or 17:7669560C>G. The latter variations are reported for the first time in literature. In conclusion, we report three highly frequent germline variations and two newly discovered variations in 3UTR of TP53 germline DNA in HNC patients from Pakistan. These results shall contribute to delineating the genetic component of HNCs with potential translational implications.

In Brazil, it is estimated that at least 10% of the native flora, which corresponds to about 4 or 5 thousand plant species, are used for food purposes. Among these species, Pereskia aculeata Mill, known as ora-pro-nobis and belonging to the Cactaceae family, stands out. It is considered a Unconventional Food Plant (UFP) and a vegetable of traditional use, playing an important role in the diet and economy of rural and urban families. Due to their high nutrient content, they have both economic and social significance, being a target of interest for scientific research. Despite the potential for human use of this species, there is a lack of knowledge about its diversity and genomic structure, resulting in few genetic and molecular studies that can contribute to the conservation and improvement of this species in general. It is essential to acquire information about its genomic structure, as well as to carry out studies related to the comparison, selection and optimization of protocols for obtaining quality DNA and in sufficient quantity, in order to guarantee the success of molecular and evolutionary analyses. The aim of this research was to evaluate qualitatively and quantitatively methods of isolation of genomic DNA from Pereskia aculeata Mill. Significant variations were observed in the DNA extracted and in the purity observed between the different protocols tested, with the isolation by the protocol described by Sambrook et al. (1989) being the most efficient in terms of concentration and purity, followed by the modified method proposed in this work. This work is a pioneer in comparing methods for isolating DNA from Pereskia aculeata Mill. in terms of quality and quantity. In addition, it was found that there is still a scarcity of molecular studies on this species, both worldwide and in Brazil, which indicates the existence of a promising gap for the development and realization of scientific research in this field.

The use of molecular techniques to deal with plant molecular breeding requires the extraction of genomic DNA in good quantity and quality, which can be influenced by method of extraction and source of DNA (plant species, plant part or tissues). However, this research focuses on plant tissue source and tried to describe the quality and quantity of DNA isolated from different tissues of barley crop. CTAB protocol was used for the isolation of DNA and both the quality and quantity of this DNA was validated through gel electrophoresis, Qubit quantification and PCR amplifications. The result showed that DNA could be successfully extracted from all tissues of plants and the yield of DNA obtained was variable ranging from 179ng l-1 in stem to 750ng l-1 in young leave. Band intensity of genomic and PCR amplified DNA was good for DNA isolated from young and matured leave. Faint band was observed in the PCR amplification for DNA isolated from stem but no to unreliable amplification was obtained for seeds and roots, respectively. Thus, for any molecular technique in barley crop research, the best tissue for DNA isolation using modified CTAB, is young or matured leave and alternatively stem can be used as DNA source.

BackgroundMoloney murine leukemia virus (MLV) replication is suppressed in mouse embryonic stem cells (ESCs) by the Trim28-SETDB1 complex. The chromatin remodeler Smarcad1 interacts with Trim28 and was suggested to allow the deposition of the histone variant H3.3. However, the role of Trim28, H3.3, and Smarcad1 in MLV repression in ESCs still needs to be fully understood. ResultsIn this study, we used MLV to explore the role of Smarcad1 in retroviral silencing in ESCs. We show that Smarcad1 is immediately recruited to the MLV provirus. Based on the repression dynamics of a GFP-reporter MLV, our findings suggest that Smarcad1 plays a critical role in the establishment and maintenance of MLV repression, as well as other Trim28-targeted genomic loci. Furthermore, Smarcad1 is important for stabilizing and strengthening Trim28 binding to the provirus over time, and its presence around the provirus is needed for proper deposition of H3.3 on the provirus. Surprisingly, the combined depletion of Smarcad1 and Trim28 results in enhanced MLV derepression, suggesting that these two proteins may also function independently to maintain repressive chromatin states. ConclusionsOverall, the results of this study provide evidence for the crucial role of Smarcad1 in the silencing of retroviral elements in embryonic stem cells. Further research is needed to fully understand how Smarcad1 and Trim28 cooperate and their implications for gene expression and genomic stability. Take homeO_LIDepletion of Smarcad1 impairs retroviral repression. C_LIO_LISmarcad1 is necessary for proper recruitment of Trim28 and H3.3 deposition. C_LIO_LIDepleting Smarcad1 and Trim28 results in enhanced derepression of the MLV provirus. C_LI

5S rDNA is essential component of all cell types. A survey was conducted to study the 5S rDNA site number, position, and origin of signal pattern diversity in 64 plants using fluorescence in situ hybridization. The species used for this experiment were chosen due to the discovery of karyotype rearrangement, and for species checked in which 5S rDNA has not yet been explored. The chromosome number was 14-160, while the chromosome length was 0.63-6.88 m, with 37 plants (58%) had small chromosome (<3 m). The chromosome numbers of three species and 5S rDNA loci of 19 species have been reported for the first time. 5S rDNA was varied and abundant in signal site number (2-18), position (e.g., interstitial, distal, and proximal position, occasionally, outside chromosome), and even as intense (e.g., strong, weak, and slight). Our results exposed modifiability in the number and location of 5S rDNA in 33 plants and demonstrated an extensive stable number and location of 5S rDNA in 31 plants. The potential origin of signal pattern diversity was probably caused by chromosome rearrangement (e.g., deletion, duplication, inversion, translocation), polyploidization, self-incompatibility, and chromosome satellites. These data characterized the variability of 5S rDNA within the karyotypes of the 64 plants that exhibited massive chromosomal rearrangements and provided anchor points for genetic physical maps. These data prove the utility of the 5S rDNA oligonucleotide as a chromosome marker in identifying plant chromosomes. This method provides a basis for developing similar applications for cytogenetic analysis in other species. Article SummaryA survey was exploited to study 5S rDNA signal pattern diversity in 64 plants by FISH. The chromosome number of three species and 5S rDNA loci of 19 species have been reported by the first time. 5S rDNA was really rather various and abundant in signal site number (2-18), position (interstitial, distal, proximal position, occasionally, outside chromosome), even as the intense (strong, week, slight). These data prove the utility of 5S rDNA oligonucleotide as chromosome marker in identifying plant chromosomes.

IntroductionTraditional disease modeling approaches have primarily utilized animal models and immortalized cell lines to investigate disease mechanisms and develop therapeutic strategies. However, previous research indicates that only about 5% of therapeutic interventions tested in animal models eventually receive regulatory approval for human use, highlighting limitations. The discovery of human-induced pluripotent stem cells (iPSCs) by Yamanakas team in 2007 has revolutionized the field with remarkable possibilities for modeling human diseases, drug testing, and regenerative medicine. Differentiated cells derived from iPSCs in two-dimensional (2D) monolayers offer a relatively straightforward system to study disease pathogenesis and underlying molecular mechanisms. ObjectiveThe present study aimed to generate and characterize an induced pluripotent stem cell (iPSC) line from peripheral blood mono-nuclear cells (PBMCs) of a healthy individual intending to serve as an age and gender-matched control for future disease modeling and regenerative medicine research. Materials and MethodsPBMCs were isolated from a healthy 31-year-old male volunteer. Somatic reprogramming was performed using episomal vectors expressing OCT3/4, SOX2, KLF4, and L-MYC. The resulting colonies were cultured and characterized for pluripotency markers by immunocytochemistry demonstrating the ability to differentiate into three germ layers. Karyotype analysis was performed to check the chromosomal abnormalities. InferencesThe resulting iPSC line exhibited pluripotency markers with the differentiation ability into three germ layers. Karyotyping analysis confirmed a normal chromosomal profile in both the donor and the reprogrammed iPSC line. This iPSC line could be a valuable resource as a healthy control for disease modeling and will contribute to advancing stem cell research with potential for regenerative medicine applications.

Cold-shock DEAD-box protein A (CsdA) is an ATP dependant cold shock DEAD-box RNA helicase. It is a major cold shock protein needed for the cold adaptation in Escherichia coli. Although the CsdA has been studied at the protein level, further studies are necessary to understand its mechanisms of gene regulations. In this regard, we have constructed a promoter less vector with the ORF of a GFP reporter and found that the promoter of the csdA gene resides far upstream (more than 800 bases) of its coding region. Furthermore, our in vivo deletion experiment has confirmed the existence of this extraordinarily long 5UTR. Our results show that it represses its own expression. In addition, the short peptide encoding (26 aa) yrbN gene resides within this 5UTR as an operon with 8 overlapping nucleotides with the csdA coding region. Besides, we observed that the csdA gene expression may also occur along with immediate upstream (180 nucleotides) nlpI gene both at 37{degrees}C and 15{degrees}C and from the pnp gene (1173 nucleotides upstream) only during cold. In conclusion, csdA gene has operon feature like prokaryotes, in contrast, it also contains an extraordinarily long 5UTR, found in eukaryotes.

A gene that encodes for Heat Shock Factor (Hsf) was characterized and designated as TaHsf2 (Gen Bank ID: KP063542) from wheat. The genomic sequence was found to be 1551bp long with open reading frame of length1123bp. The amino acid sequence so concluded showed to have HSF domain having high degree of similarity (homology) with other HSF coding gene of related species. We have reported in-silico three-dimensional model of TaHsf2 and evaluation of the predicted model, analysed with various tools. The sudden climatic variations over the years especially elevation in temperature has adversely affected the growth and productivity of winter wheat (Triticum aestivum L.). A very less number of HSFs is being reported and characterized in wheat and the data so available for wheat calls for investigation and characterization of more HSPs coding HSFs. In our present study we have cloned one of the novel HSFs and have characterized it through in silico tools.

DNA methylation patterns are closely related to the chromatin structure, and its remodeling is considered an important mechanism in the control of gene transcription during cell differentiation. In rodent, several studies have related the possibility that multipotent mesenchymal stromal cells (MSCs) undergo cardiomyogenesis. However, it has not been completely elucidated if human adult stem cell exhibits true differentiation potential for a cardiac lineage. In this study, the action of the DNA methylation inhibitor 5-azacytidine (5-aza) was examined in human adipose tissue pericytes (hATPCs: 3G5+) regarding their possible capacity to induce myocytes in vitro. Real-Time PCR revealed that cells treated with 5-aza presented time-dependent decrease in the mRNA expression of -cardiac actin (-CA). At 24 h, this diminution was statistically significant; however, there was not a correlation with the highest level of DNA demethylation at the same period using Methylation-Sensitive High Resolution Melting-PCR (MS-HRM-PCR). An evident increase in the -CA protein expression was observed by Western blotting in hATPCs treated with 5-aza at 24 h. The mRNA expression of -SMA (-smooth actin) also showed a time-dependent decrease after the treatment, however, it was not significant. The ultrastructural analysis showed similar structures such as like-cell junctions, caveolae, and actin myofilaments, which aligned in parallel. These phenotypic alterations were found only after the treatment; however, the hTAPCs after 5-aza treatment were not able to form thick myofilaments and consequently sarcomeres. These results indicated that a terminal cardiac differentiation of hTAPCs was not achieved and that the cardiomyogenesis failure could be related to the non-muscle origin of the adipose tissue.

Here, we report a whole transcriptome analysis of LlaNAC gene (from Lepidium latifolium) containing transgenic tobacco line (NC10) and wild type (WT), to attain deeper knowledge into the downstream genes activated by the over-expressing transgene. Transcriptome sequencing of NC10 and WT samples generated huge data using Illumina platform. The maximum number of unigenes GO annotated were of Biological process (8988, 3209) followed by molecular function (5155, 2577) and cellular components (3826, 1583) for WT and NC10 samples respectively. KEGG Pathway analysis revealed the unigenes were enriched in different functional pathway categories. The unigenes whose products involved in carbohydrate metabolism, glycan metabolism, and secondary metabolites synthesis were more for NC10 library in comparison to WT. Greater variety of transcription factors were involved in transgenic than wild-type plants. Genes like, Copia-like retrotransposable element, Peroxidase 64-like, Peptidyl-prolyl cis-trans isomerise, Cytochrome P450, Lipoyl synthase, CBL-interacting serine/threonine-protein kinase 5-like etc. were found differentially expressed in both the samples. Promoter analysis of these differentially expressed genes have elements for defence and stress response, abscisic acid response, shoot specific expression and light response, etc. In summary this study reports the involvement of the overexpressed genes in the dual action of cold tolerance and biomass accumulation, as sugars participate in both of these activities of the cell.

Backgroundgeneral structure of human sperm has not been profiled yet. Human sperm DNA characterization should progress the medical diagnostic and therapeutic methods rather than developing biological sciences. The aim of the present study was to provide biological insights into the common structure of human sperm. The value of this investigation is establishing an initial basic map of sperm head structure that leads to further advanced standardization of normality in this creature. For this purpose, analytical and microscopic methods were applied. MethodsHigh-performance Liquid Chromatography (HPLC) and flow cytometry were hired to quantify the DNA compositions. As well fluorescent, confocal and advanced light microscopy was applied to identify the stained sperm DNA by chromomycinA3 (CMA3) and 5-methylcytosine antibody (5-mc) ResultsHPLC demonstrated the mean values of nucleotide bases percentage in the structure of the sperm DNA regardless of the fraction that sperm was collected from gradient wash, sequenced from 27.6%, 8.92%, 27.05% and 35.36%. Also, quantitative flow cytometry of global 5-methylcytosine showed not a regular fluctuation in individuals with normal sperm while, there is a permanent increase in 50% fraction collected from percoll gradients. CMA3-positivity levels as well, were negatively correlated with sperm quality harvest by percoll gradients (p<0.0001), and positively correlated (P<0.05) with global methylation as determined by flow cytometry. Interestingly, in this text microscopy of immunocytochemistry of sperm cells stained by CMA3, demonstrated a different view from cells heads. Conclusionsobviously these explorations suggest some new possibilities in assessment of rough chemical level of nucleotides and cytochemistry of sperm head structure. The chromatin brightness presented with CMA3 by microscopy shows a direct relation with more extensive DNA methylation in sperms collected from low gradients of percoll wash. While, fluctuated 5-methylcytosine levels show personal presentation and even exclusive to individual sperm expression. This study induces further research on new assumptions in nuclear equilibrium in the axiom of DNA ladder in related to 5-mcytosine level in human sperm.

This study supplements earlier received experimental data using modern databases. Previously tumor-specific activity of several human native and chimeric promoters was demonstrated. Here we compared tumor-specific promoters with promoters of housekeeping genes by the presence of recognition profiles for transcription factors in DNA sequences of the promoters. A number of transcription factor recognition profiles have been identified, the presence of which in promoters may indicate the tumor specificity of the promoters. Transcription factors which may directly regulate promoters of genes involved in cell proliferation and carcinogenesis were revealed by pathway analysis. The results of the study may help in studying the peculiarities of gene transcription in tumors and in the search for or the creation of tumor-specific promoters for cancer gene therapy.

Information of the Passiflora genome is still very limited. Understand the evolutionary relationship between different species of Passiflora, and develop a large number of SSR markers to provide a basis for the genetic improvement of Passiflora. Applying restriction site associated DNA sequencing (RAD-Seq) technology, we studied the phylogeny, simple sequence repeat (SSR) and marker transferability of 10 accessions of 6 species of Passiflora. Taking the partial assembly sequence of accessions P4 as the reference genome, we constructed the phylogenetic tree using the detected 46,451 high-quality single nucleotide polymorphisms (SNPs), showing that P6, P7, P8 and P9 were a single one while P5 and P10 were clustered together, and P1, P2, P3 and P4 were closer in genetic relationship. Using P8 as the reference genome, a total of 12,452 high-quality SNPs were used to construct phylogenetic tree. P3, P4, P7, P8, P9 and P10 were all single branch while P1 and P2 were clustered together, and P5 and P6 were clustered into one branch. A principal component analysis (PCA) revealed a similar population structure, which four cultivated passion fruits forming a tight cluster. A total of 2,614 SSRs were identified in the genome of 10 Passiflora accessions. The core motifs were AT, GA, AAG etc., 2-6 bases, 4-16 repeats, and 2,515 pairs of SSR primer were successfully developed. Tthe SSR transferability in cultivated passion fruits is the best. These results will contribute to the study of genomics and molecular genetics in passion fruit.

Myotonic dystrophy type 1 (DM1) is a neuromuscular disorder caused by the genomic expansions of CTG repeats, in which RNA-binding proteins, such as muscleblind-like protein, are sequestered in the nucleus, and abnormal splicing is observed in various genes. Although abnormal splicing reportedly occurs in the brains of patients with DM1, it is relation to the central nervous system symptoms is unknown. Several imaging studies have indicated substantial white matter (WM) defects in patients with DM1. Here, we performed RNA-sequencing and analysis of CTG repeat lengths in the frontal lobe of patients with DM1, separating the grey matter (GM) and WM, to investigate the splicing abnormalities in the DM1 brain, especially in the WM. The results demonstrated the number of repeats in the GM tended to be increase, with several genes showing similar levels of splicing abnormalities in the GM and WM, suggesting that the WM defects in DM1 are not only caused by aberrant splicing of GM RNA but also of WM RNA, which could be attributed to abnormal splicing of glial cell RNAs.

The polymorphism profiles of language genes (LG) display different patterns across various ancient and modern populations, leading to the speculation that cognition gene (CG) polymorphism profiles may exhibit similar trends. However, the evolutionary processes of language gene polymorphism patterns (LGPP) and cognition gene polymorphism patterns (CGPP) are likely to demonstrate distinct characteristics. In particular, it is intriguing to determine whether there is any overlap in the timing of significant changes in CGPP and LGPP over the large timescales of evolution. The potential existence of such overlap can also be assessed by examining whether the samples carrying significant changes in LGPP and CGPP are the same. This study investigated the genetic differences at 239 SNP loci in 18 language genes (LG) and 223 SNP loci in 18 cognition genes (CG) across 170 whole genomes. Principal component analysis (PCA) was used to cluster the SNP data of the aforementioned samples, and the similarity of SNP patterns between each sample was calculated from three perspectives: LG, CG, and CGLG. The basic conclusions are as follows: (1) If different positions in the PCA analysis results can essentially represent the pattern differences in SNP polymorphisms, then both language gene polymorphism patterns and cognition gene polymorphism patterns have undergone distinct stages of evolution; (2) There were significant differences in the early manifestations of language gene polymorphism patterns and cognition gene polymorphism patterns during human evolution: Language gene polymorphism patterns could not differentiate general animals, primates, and ancient human samples in the early stages of evolution, whereas cognition gene polymorphism patterns seemed to be initially divisible into two patterns, one closely resembling a group of animals and certain ancient human samples, and the other reflected in a different set of animal and primate samples. (3) It appears that samples from all five continents can be observed at every stage of evolution, suggesting that new evolving populations have always had ample time to spread across continents. (4) A quantitative comparison of the SNP profiles of 170 samples revealed that their CG and LG plus CGLG profiles indeed have 2-3 potential significant change points, and the samples carrying these significant change points has 2 common samples, namely ge1 (Georgia) and us2 (North America), implying that the most significant changes in language or cognition gene polymorphism patterns during human evolution may have occurred in some human populations in Europe/ North America.

AimsThe study was performed to evaluate the role of red rose extract (Pierre de Ronsard) on B lymphocytes. Specifically the gene expression of CD20, CD30, CD40, and CCR5 in human B cells were studied after treatment with rose extract. MethodsRed rose extract was prepared at the dilution of 0.0075% (v/v) and stored until use at -20{degrees}C. Cell treatment was performed at 37{degrees}C on B cells. The cells were plated in 6 well plates at 1.5x106 cells per well and stored at -80{degrees}C. Total RNA extraction and quality control were performed. RTq-PCR was performed according to Genecopoeias instructions. The cycle threshold method ({Delta}{Delta}Ct) was used for data analysis. ResultsThe comparative Ct method quantification (2^-{Delta}Ct) and fold change for CD20, CD 30, CD 40 and CCR5 were - 5.65E+01, 4.80E-01, N/A, 2.47E-01; and 0.954,0.377, N/A and 0.577, respectively. The amount of total RNA extracted from about 4.5x106 cells was low and did not allow us to measure the RNA profile. The A260/A230 ratios were very low due to the low amount of RNA. The analysis of gene expression by qRT-PCR showed that CD40 was not expressed in untreated cells and cells treated with rose extract with the Ct values over 32. All other genes were expressed and well-measured in both B cell samples. ConclusionThe treatment with rose extract at 0.0075% (v/v) did not modify the expression of CD20. However, the expression of CD30 and CCR5 decreased with the rose extract treatment. The range of the fold change showed that the result of CD30 expression was more accurate than for CCR5.

The plant height of rapeseed is one of the important factors that affects the production of rapeseed. If the plant height of rapeseed is too high, on the one hand, it will cause rapeseed to lodge and affect the yield, on the other hand, it will also affect the mechanized harvesting of rapeseed. In this research, the high-stalked line (YY50) and the dwarfed line (DW871) are crossed to obtain an F2 rapeseed population which was used to build pools, and then we used this to mine the main dwarfing genes. In the pools composed of tall and short stalks, we obtained 192.80Mb clean reads, which can be used for BSA (bulked segregant analysis). Preliminary positioning around the candidate section identified 23 SNP markers. Then 17 polymorphic SNP markers were obtained through polymorphism screening. Further we narrowed the candidate interval, and finally determined between 15.51-16.60Mb of ChrA10. Through identifying 231 genes from the above interval, its predicted that the production of dwarf traits may be related to lignin synthesis and limited inflorescence. It provides a basis for further mapping and cloning of the dwarfing gene DW871.

BACKGROUNDResearch on peripheral leukocyte gene expression in human health and disease is growing rapidly. However, how to process sample efficiently, simply and stably, and how to reflect human physiological state preferably remains critical issues in large cohort studies.\n\nMETHODSWe used RNA-seq to explore the differences of gene expression profiles among whole blood (WB) and three groups of leukocytes from buffy coat (BC) extraction, red blood cell (RBC) lysis and peripheral blood mononuclear cell (PBMC) isolation.\n\nRESULTSThe residual globin mRNA in leukocytes from RBC lysis (1.00% {+/-}1.23%) and PBMC isolation (0.06% {+/-} 0.03%) was much less than that in leukocytes from BC extraction (17.48% {+/-} 6.95%) and WB (24.46% {+/-} 6.43%), resulting in higher transcriptome mapping rates and larger numbers of detected genes. The expression of 616 genes associated with leukocyte function was slightly higher in leukocytes from RBC lysis than that from BC extraction and WB, but barely detected in leukocytes from PBMC isolation.\n\nCONCLUSIONSWe suggest that sample processing based on RBC lysis could allow better applications of gene expression profiling of peripheral leukocytes in large cohort studies.

Hemoglobinopathies are the most common type of inherited disease in human. in India the most frequent and clinically significant hemoglobin structural variants are HbS, HbD and HbE. The HbS mutation, in which a glutamic acid at position 6 in the {beta} chain is substituted for valine Sickle cell disease is a major health problem in some parts of India. 2 ml blood sample was collected from 350 anemia patient and PCR-RFLP method was used for hemoglobin S analysis. Out of 350 samples, in four individuals, HbS mutation was found in homozygous ({beta} 6/{beta} 6) condition. All four individuals are Sickle cell cases. In conclusion, the percentage of Sickle cell disease was observed as 1.14% in Eastern UP anemic patients.

BackgroundFFPE tissue samples are commonly used in biomedical research and are a valuable source for next-generation sequencing in oncology, however, extracting RNA from these samples can be difficult the quantity and quality achieved can impact the downstream analysis. This study compared the effectiveness of seven different commercially available RNA extraction kits specifically designed for use with FFPE samples in terms of the quantity and quality of RNA recovered. MethodsThis study used 9 samples of FFPE tissue from three different types of tissue (Tonsil, Appendix and lymph node of B-cell lymphoma) to evaluate RNA extraction methods. Three sections of 20m of each sample were combined per sample. The slices were distributed in a systematic manner to prevent any biases. Each of the 7 commercially available RNA extraction kits were used according to manufacturers instructions, with each sample being tested in triplicate resulting in a total of 189 extractions. The concentration, RNA integrity number (RIN) and DV200 of each extraction was analysed using a LabChip to determine the quantity and quality of the recovered RNA. ResultsThis study found that despite processing the FFPE samples in the same standardized way, there were disparities in the quantity and quality of RNA recovered across the different tissue types. Additionally, the study found notable differences in the quantity of RNA recovered when using different extraction kits. In terms of quality, three of the kits performed better than the other four in terms of RNA integrity number (RIN) and DV200 values. ConclusionThough many laboratories have developed their own protocols for specific tissue types, using commercially available kits is still a popular option. Although these kits use similar processes and extraction procedures, the amount and quality of RNA obtained can vary greatly between kits. In this study, among the kits tested, while the Roche kit, provided a nearly systematic better-quality recovery than other kits, the ReliaPrep FFPE Total RNA miniprep from Promega yielded the best ratio of both quantity and quality on the tested tissue samples.