This dataset was published by Haseeb et al., 2021
and raw data can be found in the BioProject PRJNA646069.
Tissue source: LCM articular cartilage.
Age: 12 weeks old.
Sex: Both.
Layout: PAIRED.
Reads length: 100 bp.
Experimental procedure: Knee joints were harvested from control and mutant mice and were snap frozen in OCT compound. Frozen sections were cut and then processed for laser-caputure microdissection. Articular and growth plate cartilage area were removed using laser-capture mirodissection, flash frozen on dry ice, and RNA was harvested using Qiagen Kit RNeasy Plus Micro Kit. SMARTer Stranded Total RNA-seq v2 Pico Input Mammalian kit (Takara Bio) was used for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols. Sequencing was performed with Illumina NovaSeq 6000 and normalized to TPM.
Epiphysis
This dataset was published by Taylor et al., 2022
and raw data can be found in the BioProject PRJNA793842.
Tissue source: Immature epiphyseal chondrocytes isolated by collagenase digestion overnight.
Age: 5 days old.
Sex: Males.
Layout: PAIRED.
Reads length: 100 bp.
Experimental procedure: Primary epiphyseal chondrocytes were harvested from five-day-old male wildtype (n=3) mice. Chondrocytes were digested for one hour in collagenase II in DMEM culture medium. Cells were isolated, washed in ice-cold PBS three times, then resuspended in Trizol for RNA extraction. Total RNA was extracted using phenol-chloroform.
TruSeq RNA method was used for analysis of polyadenylated mRNAs that were selected using oligo dT magnetic beads. TruSeq Kits were used for indexing to permit multiplex sample loading on the flow cells of an Illumina HiSeq 2000 sequencer and normalized to TPM.
Proliferative and Hypertrophic
This dataset was published by Lui et al., 2018
and raw data can be found in the BioProject PRJNA473132.
Tissue source: Proliferative and Hypertrophic zones from Growth Plate isolated with LCM.
Age: 1 week old.
Sex: Both.
Layout: PAIRED.
Reads length: 50 bp.
Experimental procedure: Freshly frozen bone/cartilage were sectioned and microdissected with laser capture microdissection (LCM) and RNA was extracted with Rneasy Micro Kit
RNA-Seq libraries were then constructed using Nextera XT DNA Library Preparation Kit (Illumina, San Diego, CA)
Sequencing was performed with Illumina HiSeq 3000 and normalized to TPM.
Datasets come from two works:
1) This dataset was published by Vrahnas et al. 2019
and raw data can be found in the BioProject PRJNA434572
Tissue source: Femora.
Age: 12 weeks old.
Sex: Females.
Layout: PAIRED.
Reads length: 100 bp.
Experimental procedure: RNA-seq profiles were obtained from femoral bone from 3 ephrin B2 deficient (Dmp1Cre.EfnB2f/f) mice and 4 control mice (Dmp1Cre).
Femora were collected, ends removed and marrow flushed out prior to snap freezing in liquid nitrogen. The cortical bone was homogenized in QIAzol lysis reagent using a Polytron PTA 20S homogenised. RNA was extracted using a Rneasy Lipid Tissue Mini kit (Qiagen) according to manufacturer's instructions.
mRNA reverse transcription cDNA libraries were prepared using the TruSeq RNA Sample preparation kit (Illumina) following the manufacturer’s instructions.
Sequencing was performed with Illumina HiSeq 2500 and normalized to TPM.
2) This dataset was published by Davis et al. 2019
and raw data can be found in the BioProject PRJNA550395
Tissue source: Tibiae.
Age: 16 weeks old.
Sex: Males.
Layout: SINGLE.
Reads length: 75 bp.
Experimental procedure: Differential gene expression in Ctrl vs NT3-Cre+ non-loaded and loaded limbs. 16 total samples. n=4 each genotype (Ctrl or NT3-Cre+) as biological replicates. Paired non-loaded and loaded samples for each mouse.
Right and left tibiae were stripped of all muscle tissue and flushed by centrifugation to remove bone marrow. The flushed tibiae were then cut 2 mm from the articular surface and again at the distal TFJ to isolate the area of peak strain. Tissue samples were flash-frozen in liquid nitrogen and stored at -80C°. Frozen tibiae were pulverized in liquid nitrogen using a Mikro Dismembrator (Mikro-Dismembrator S; B. Braun Biotech International, Melsungen, Germany), resuspended in TRIzol (15596026; Invitrogen, USA), and stored at -80C°. Total RNA was isolated using a Direct-zol kit per manufacturer instructions (R2072; Zymo Research, USA). RNA concentration was determined using a Nanodrop (ND-2000; ThermoFisher, USA). RIN values were used to determine RNA quality (Bioanalyzer 2100; Agilent Technologies, USA).
RNA sequencing was performed on the Total RNAble platform (Cofactor Genomics, http://cofactorgenomics.com, St. Louis, MO, USA). Libraries were prepared and sequenced as single-end 75 base pair reads on an Illumina NextSeq500 following the manufacturer’s protocols.
This dataset was published by Cho et al., 2021
and raw data can be found in the BioProject PRJNA750927.
Tissue source: Achilles tendon.
Age: 12 weeks old + 3 or 6 weeks after treatment.
Sex: Males.
Layout: PAIRED.
Reads length: 100 bp.
Experimental procedure: The Achilles tendon was separated from soleus muscle and were homogenized using a tissue homogenizer, and total RNA was extracted using TRI reagent and the manufacturer's protocol.
The sequencing library is prepared by random fragmentation of the DNA or cDNA sample, followed by 5’ and 3’ adapter ligation. Alternatively, “tagmentation” combines the fragmentation and ligation reactions into a single step that greatly increases the efficiency of the library preparation process. Adapter-ligated fragments are then PCR amplified and gel purified.
Sequencing was performed with Illumina HiSeq 2000 and normalized to TPM.
This dataset was published by Mousa et al., 2023
and raw data can be found in the BioProject PRJNA818005.
Tissue source: Gastrocnemius and soleus muscles.
Age: 12 weeks old.
Sex: Males.
Layout: PAIRED.
Reads length: 150 bp.
Experimental procedure: Muscles were harvested and immediately snap frozen in liquid nitrogen. Total RNA was isolated using RNA STAT-60 as per manufacterer's instructions. RNA was DNase I treated as per manufacturer’s instructions (RNase-Free DNase Set, Qiagen) then cleaned up and eluted with RNAse and DNAse free molecular grade water (RNeasy MinElute Cleanup Kit, Qiagen), followed by quantification.
KAPA RiboErase. Sequencing was performed with Illumina NovaSeq 6000 and normalized to TPM.
Datasets come from two works:
1) This dataset was published by Vail et al., 2020
and raw data can be found in the BioProject PRJNA528088.
Tissue source: Articular cartilage from the femoral condyle and/or tibial plateau. (Articular Engineering, LLC).
Age: 20/20/35 years old.
Sex: Both.
Layout: PAIRED.
Reads length: 150 bp.
Sample processing: Tissue was snapped frozen in liquid nitrogen and mechanical crushed followed by RNA extraction with trizol.
All samples underwent trizol phase separation of nucleic acids followed by QIAGEN column RNA purification (catalog number 74194)
Strand specific cDNA (250-300bp) libraries constructed using the NEBNext Ultra Kit for low input (5ng-1000ng) following poly-T oligo-attached magnetic bead purification. Quality assessed by insert size (Agilent 2100).
Sequencing was performed with Illumina HiSeq 4000 and normalized to TPM.
2) This dataset was published by Fisch et al., 2018
and raw data can be found in the BioProject PRJNA454873.
Tissue source: Knee articular cartilage from different donors.
Age: 18-61 years old, mean 38.
Sex: Both.
Layout: SINGLE.
Reads length: 75 bp.
Sample processing: Cartilage was stored at -20ºC in Allprotect Tissue Reagent immediately after harvest until RNA extraction. For RNA isolation, tissue was pulverized and homogenized in Qiazol Lysis Reagent. To remove proteins and cellular debris, a initial phenol-chloroform extraction was performed.
mRNA libraries were prepared using the Encore Complete RNA-Seq DR Multiplex System 1-8 and 9-16 (NuGen) with 16 unique indexed adapters. Sequencing was performed with Illumina HiSeq 2000 and normalized to TPM.
Epiphysis
This dataset was published by Richard et al., 2023
and raw data can be found in the BioProject PRJNA801843.
Tissue source: Epiphyseal chondrocytes.
Age: Fetal, late first-trimester and early second-trimester terminations.
Sex: Not specified.
Layout: PAIRED.
Reads length: 150 bp.
Sample processing: Microdissected fetal cartilage was minced and subsequently incubated with collagenase. Liberated cells were pelleted by centrifugation and lysed in guanidine thiocyanate buffer. Total RNA was purified using silica column-based kits.
Libraries were constructed using the TruSeq RNA Library Prep Kit v2 (Illumina, San Diego). Sequencing was performed with Illumina NextSeq 500 and normalized to TPM.
Growth plate
Datasets come from two works:
1) This dataset was published by Richard et al., 2023
and raw data can be found in the BioProject PRJNA801843.
Tissue source: Growth plate chondrocytes.
Age: Fetal, late first-trimester and early second-trimester terminations.
Sex: Not specified.
Layout: PAIRED.
Reads length: 150 bp.
Sample processing: Microdissected fetal cartilage was minced and subsequently incubated with collagenase. Liberated cells were pelleted by centrifugation and lysed in guanidine thiocyanate buffer. Total RNA was purified using silica column-based kits.
Libraries were constructed using the TruSeq RNA Library Prep Kit v2 (Illumina, San Diego). Sequencing was performed with Illumina NextSeq 500 and normalized to TPM.
2) This dataset was published by Li et al., 2017
and raw data can be found in the BioProject PRJNA420979.
Tissue source: Growth plate chondrocytes.
Age: Fetal (14 to 18 weeks old).
Sex: Not specified.
Layout: PAIRED.
Reads length: 75 bp.
Sample processing: RNA was isolated and purified from five independent 14-18 week distal human femur growth plate cartilage samples. The fetal cartilage was digested in collagenase, the chondrocytes were collected by centrifugation and total RNA was isolated with TRIZol (Thermo Fisher Scientific). Genomic DNA contamination was removed by DNase I (Thermo Fisher Scientific) digestion and the RNA was further purified with the RNeasy Mini Kit (Qiagen).
PolyA-tailed mRNA was isolated from total RNA and used as the input for sequencing library construction with the TruSeq RNA Preparation Kit (Illumina). Sequencing was performed with Illumina Hiseq 2000 and normalized to TPM.
Datasets come from two works:
1) This dataset was published by Farr et al., 2015
and raw data can be found in the BioProject PRJNA295101.
Tissue source: Iliac crest bone.
Age: 22-40 years old, mean 30.
Sex: Women.
Layout: PAIRED.
Reads length: 50 bp.
Sample processing: RNA-seq profiles were obtained from small needle bone biopsies from the posterior iliac crest of all subjects using an 8G needle under local anesthesia
total RNA was isolated using the RNeasy Micro Kit (Qiagen, Valencia, CA) and treated with the Turbo DNA-freeTM Kit (Life Technologies, Grand Island, NY) to remove potential contaminating DNA that may lead to false-positive amplification.
Sequencing was performed with Illumina HiSeq 2000 and normalized to TPM.
2) This dataset was published by Martinez-Calle et al., 2023
and raw data can be found in the BioProject PRJNA798962.
Tissue source: Iliac crest bone.
Age: Mean 40 years old.
Sex: Women.
Layout: SINGLE.
Reads length: 100 bp.
Sample processing: From iliac crest bone biopsies, total RNA was isolated using TRI reagent (Waltham, MA, USA) and purified RNA using RNeasy kit (Qiagen, Germantown, MD, USA) and submitted 1 µg of total RNA for sequencing.
Libraries were prepared according to Illumina's instructions, i.e, total RNA library for each individual sample was prepared using the TruSeq Total RNA-Seq Library Preparation Kit (Illumina, San Diego, CA), and the bar-coded cDNA libraries were sequenced for 100 bp single reads using Illumina HiSeq 4000.
This dataset was published by Zheng et al., 2018
and raw data can be found in the BioProject PRJNA429184.
Tissue source: Tendon.
Age: Adult (not specified).
Sex: Both.
Layout: PAIRED.
Reads length: 150 bp.
Sample processing: Tendon tissues were removed, flash frozen on dry ice, and RNA was harvested using Trizol reagent. Illumina TruSeq Standard Total RNA Prep Kit was used for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols. Sequencing was performed with Illumina HiSeq 4000 and normalized to TPM.
This dataset was published by Depuydt et al., 2022
and raw data can be found in the BioProject PRJNA837196.
Tissue source: Vastus lateralis, Semimembranosus and Rectus Femoris muscles.
Age: 26-64 years old, mean 45,7.
Sex: Men.
Layout: PAIRED.
Reads length: 50 bp.
Sample processing: RNA was extracted from muscle biopsies using the Direct-zol RNA Miniprep Plus kit ZY-R2073 (Zymo Research, Irvine, CA, USA). Samples were treated with DNase prior to their submission to the VIB Nucleomics Core (www.nucleomics.be). RNA concentration and purity were determined spectrophotometrically using the Nanodrop ND-1000 (Nanodrop Technologies, Wilmington, DE, USA). Libraries were constructed using the SMART-Seq Stranded Kit (Takara Bio USA, San Jose, CA, USA).
Sequencing was performed with Illumina NovaSeq 6000 and normalized to TPM.
Experimental condition: Transcription profile of specific deletion of EphrinB2 in osteocytes (Dmp1CreEfnB2fl/fl).
Genes: EphrinB2 (EfnB2), this gene encodes a member of the ephrin (EPH) family, precisely from ephrin-B (EFNB) class, which are transmembrane proteins. The ephrins and EPH-related receptors comprise the largest subfamily of receptor protein-tyrosine kinases.
Phenotype: EphrinB2 conditional knock out mice showed brittle bones. However, they did not show low bone mass, but defective bone material.
Tissue: Femora.
Age: 12 week old.
Sex: Females.
Layout: Paired.
Reads lenght: Paired.
Sample processing: Femora were collected, ends removed and marrow flushed out prior to snap freezing in liquid nitrogen. The cortical bone was homogenized in QIAzol lysis reagent at 4 degrees C using a Polytron PTA 20S homogenised. RNA was extracted using a Rneasy Lipid Tissue Mini kit (Qiagen) according to manufacturer's instructions.
mRNA reverse transcription cDNA libraries were prepared using the TruSeq RNA Sample preparation kit (Illumina) following the manufacturer’s instructions.
cDNA libraries were sequenced by PE sequencing (100 x 2) with an Illumina HiSeq 2500 sequencer and normalized to TPM.
This dataset was published by Vrahnas C et al., 2019 and raw data can be found in the BioProject PRJNA434572.
Experimental condition:Transcriptional analysis of Wnt7b overexpression in bone.
Genes:Wnt7b, a member of the Wnt signalling pathway. These genes are implicated in oncogenesis and in several developmental processes. Wnt7b shows the most profoundly anabolic effects on bone.
Phenotype:Mice overexpresing Wnt7b showed better bone formation.
Tissue:Femora.
Age:8 week old.
Sex:Both.
Layout:Paired.
Reads length:120 bp.
Sample processing:Total mRNA of femur with marrow flushed was collected from 8-week-old OE and Ctrl mice respectively using the RNeasy mini kit (Qiagen, Valencia, CA) according to the manufacturer’s protocol.
Libraries were prepared according to Illumina's instructions of RNA sample. Paired-end method was adopted for further processes.
cDNA libraries were sequenced by PE sequencing with an Illumina HiSeq 2500 sequencer and normalized to TPM.
This dataset was published by Yu F et al., 2020 and raw data can be found in the BioProject PRJNA479739.
Experimental condition: Transcriptional analysis of the effects of Piezo1 deletion in bone.
Genes: Piezo1 encodes a mechanically-activated ion channel gene related with the sensing of mechanical forces by cells.
Phenotype: Mice lacking of Piezo1 showed shorter bones and osteoporosis-like phenotyphe with a decreased in bone thickness, mass and volume. This lost of bone was independent of the gender.
Tissue: Femora.
Age: 8 week old.
Sex: Both.
Layout: Paired.
Reads length: 150 bp.
Sample processing: Total mRNA from cortical bones of 6-week WT and Prx1CrePiezo1fl/fl mice were collected and grinded with liquid nitrogen to extract the RNA. RNA was harvested using Trizol reagent. Illumina TruSeq RNA Sample Prep Kit was used for the construction of sequencing libraries.
Libraries were prepared according to Illumina's instructions.
cDNA libraries were sequenced by PE sequencing with an Illumina HiSeq 2500 sequencer and normalized to TPM.
This dataset was published by Wang et al., 2020 and raw data can be found in the BioProject PRJNA558239.
Experimental condition: Transcriptional analysis of the effects of Tet1 and Tet2 deletion in bone.
Genes: Ten Eleven Translocation (TET), enzymes that oxidize 5-methylcytosines (5mCs) and promote locus-specific reversal of DNA methylation.
Phenotype: Mice lacking of Tet1 and Tet2 (enzymes that modify DNA producing 5hmC) showed severe skeletal defects mimicking bone phenotypes of Vitamin C-insufficient, Gulo knockout mice, a model of Vitamin C deficiency and scurvy.
Tissue: Femora.
Age: 11 week old.
Sex: Both.
Layout: Paired.
Reads length: 50 bp.
Sample processing:11 week old femoral bones from Prrx1-Cre driven Tet1fl/fl, Tet2fl/fl or Tet1fl/fl and Tet2 fl/fl mice were dissected, bone marrow was flushed, periost was removed and remaining bone tissue was thorogly washed in ice cold PBS.
RNA was extraced with commercially available extraction kits (Zymo). RNA-Seq libraries were prepared using the Illumina TruSeq v2 library preparation kit and sequenced with paired-end 50 bp reads
cDNA libraries were sequenced by PE sequencing with an Illumina HiSeq 4000 sequencer and normalized to TPM.
This dataset was published by Thaler R et al., 2022 and raw data can be found in the BioProject PRJNA577623.
Experimental condition: Transcriptional analysis of the effects of vitamin C supplementation in bone from Gulo-/- mice.
Genes: Gulo, a gene implied in the synthesize of vitamin C.
Phenotype: Mice lacking of enough vitamin C exhibited severe defects in bone structure and mechanical strength.
Tissue: Femora.
Age: 24 week old.
Sex: Both.
Layout: Paired.
Reads length: 50 bp.
Sample processing: 20 week old femoral bones from Wt and Gulo-/- mice with normal or defficiency of VitC supplementation were dissected, bone marrow was flushed, periost was removed and remaining bone tissue was thorogly washed in ice cold PBS.
RNA was extraced with commercially available extraction kits (Zymo). RNA-Seq libraries were prepared using the Illumina TruSeq v2 library preparation kit and sequenced with paired-end 50 bp reads
cDNA libraries were sequenced by PE sequencing with an Illumina HiSeq 4000 sequencer and normalized to TPM.
This dataset was published by Thaler R et al., 2022 and raw data can be found in the BioProject PRJNA577623.
Experimental condition: Transcriptional analysis of the effects of Oscar deletion in osteoarthritis, precisely in a Destabilization of the Medial Meniscus (DMM) model.
Genes: Oscar is an immunoglobulin (Ig)-like activating receptor of the leukocyte receptor complex that is specifically expressed on pre-osteoclasts. An activating receptor for collagen that co-stimulates osteoclast differentiation.
Phenotype: Mice lacking of Oscar gene showed a delay in osteoarthritis development.
Tissue: Articular cartilage.
Age: 10 week old + 2 or 4 weeks after surgery.
Sex: Males.
Layout: Paired.
Reads length: 100 bp.
Sample processing: Articular cartilage mRNA profiles of 10-weeks-old wild-type (WT) and Oscar knockout (Oscar-/-) mice were performed for the destabilization of the medial meniscus (DMM) or sham surgery and each cartilage tissue sample was harvested from two time-points (2- and 4-weeks after surgery). Sequencing libraries were prepared according to the manufacturer’s instructions (TruSeq Stranded mRNA Library Prep Kit; Illumina, San Diego, CA, USA).
cDNA libraries were sequenced by PE sequencing with an Illumina HiSeq 2500 sequencer and normalized to TPM.
This dataset was published by Park DR et al., 2020 and raw data can be found in the BioProject PRJNA615179.
Experimental condition: Transcriptional analysis of the effects of Osteocalcein (Bglap and Bglap2) deletion in bone.
Genes: Bglap and Bglap2 are highly abundant bone proteins secreted by osteoblasts that regulate bone remodeling and energy metabolism.
Phenotype: Not provided.
Tissue: Tibiae.
Age: 16 week old.
Sex: Unknown.
Layout: Paired.
Reads length: 75 bp.
Sample processing: RNA was extracted from cortical tibial bone and libraries were constructed using the SMART-Seq v4 UltraLow RNA Input Kit. Library construction was completed using the ThruPLEX DNA-seq kit.
cDNA libraries were sequenced by PE sequencing with an Illumina NextSeq 550 sequencer and normalized to TPM.
This dataset is not published yet and raw data can be found in the BioProject PRJNA616424.
Experimental condition: Transcriptional analysis of Crtap ko and oim/oim Osteogenesis Imperfecta models.
Genes: Crtap is a scaffold protein that holds two other proteins, P3H1 and PPIB, in a heterotrimeric complex involved in post-translational modification of the pro-chains of type I collagen. The oim/oim model presents a null mutation in the COL1A2 gene of type I collagen.
Phenotype: Both mice models displayed Osteogenesis Imperfecta phenotypes.
Tissue: Femorae and tibiae.
Age: 12 week old.
Sex: Both.
Layout: Single.
Reads length: 75 bp.
Sample processing: RNA was extracted from cortical tibial and femoral bone and libraries were constructed using the SMART-Seq v4 UltraLow RNA Input Kit. Library construction was completed using the ThruPLEX DNA-seq kit.
cDNA libraries were sequenced by PE sequencing with an Illumina NextSeq 550 sequencer and normalized to TPM.
This dataset was published by Zimmerman SL et al., 2019 and raw data can be found in the BioProject PRJNA647367.
Experimental condition:Transcriptional analysis of femur from an Implant-Associated Osteomyelitis (IAOM) model.
Surgical procedure: Orthopedic stainless pin was surgically placed in the right femoral midshaft of mice, followed by an inoculation of Staphylococcus aureus into the medullary cavity.
Phenotype: Typical characteristics of IAOM, like periosteal reaction and intraosseous abscess, occurred by day 14 postinfection. By day 28 postinfection, necrotic abscess, sequestrum formation, and deformity of the whole femur were observed.
Tissue: Femora.
Age: 10 week old.
Sex: Males.
Layout: Paired.
Reads length: 150 bp.
Sample processing: The right femur was collected and pulverized in liquid nitrogen and total RNA was extracted by TRIzol. Sequencing libraries were generated using NEBNext Ultra RNA Library Prep Kit for Illumina (NEB, USA) following manufacturer's protocols and index codes were added to attribute sequences for each sample. PCR products were purified using AMPure XP system (Beckman Coulter, USA) and library quality was assessed on the Agilent Bioanalyzer 2100 system. The clustering of the index-coded samples was performed on a cBot Cluster Generation System using TruSeq PE Cluster Kit v3-cBot-HS (Illumia) according to the manufacturer's instructions.
After cluster generation, the library preparations were sequenced on an Illumina NovaSeq 6000 and 150 bp PE reads were generated and normalized to TPM.
This dataset was published by Lin Y et al., 2021 and raw data can be found in the BioProject PRJNA701190.
Experimental condition: Transcriptional analysis of mice suffering osteomyelitis with or without IL-27 pretreatment.
Surgical procedure: Mice were intramuscularly injected with recombinant adeno-associated virus expressing murine IL-27 (rAAV-IL-27) or recombinant adeno-associatd virus expressing GFP (rAAV-GFP, control) and then challenged with a S. aureus contaminated trans-tibial implant. Subsequently, infected tibia were collected on days 1, 3, 7 and 14 post-septic surgery for RNA sequencing.
Phenotype: At early stage of the infection, IL-27 is highly expressed in osteoblasts and macrophages but not in osteoclasts where the pathogen can persist. Pretreatment with IL-27 prevents abceses formation and reduce bone loss due to osteomyelitis.
Tissue: Tibiae.
Age: 8 week old.
Sex: Females.
Layout: Single.
Reads length: 100 bp.
Sample processing: Tibiae were pulverized at liquid nitrogen temperature and homogenized using Bullet Blender Gold (Next Advance). Collection of Total RNA from homogenized tibia was performed using TRIzol extraction (ThermoFisherScientific) and RNeasy Mini Kits (Qiagen).
The TruSeq Stranded Total RNA Library Prep Gold (Illumina) was utilized for next generation sequencing library preparation per the manufacture’s instructions. Then sequencing was performed by SE with an Illumina NovaSeq 6000 sequencer and normalised to TPM.
This dataset was published by Morita Y et al., 2019 and raw data can be found in the BioProject PRJNA714525.
Experimental condition: Transcriptional analysis of femorae after hindlimb unloading.
Procedure: Mice were subjected to hindlimb unloading for 7 days. Under isoflurane anesthesia, the mouse tail was taped to a freely rotating harness connected to a wheel that moved along the central axis of the custom-made cage. The harness was adjusted such that the mouse could not touch its hind limbs to the floor or the walls of the cage.
Phenotype: Decrease in bone mass and increase the risk of fracture.
Tissue: Femorae.
Age: 14 week old.
Sex: Females.
Layout: Paired.
Reads length: 50 bp.
Sample processing: After cutting the ends of bones and flushing marrow with PBS, the cortical bone was immediately snap-frozen in liquid nitrogen.
Subsequently, Trizol was added while the bone is maintained frozen in an eppendorf tube placed in a liquid nitrogen bath, homogenized in the same eppendorf tube with the Fastprep24 machine (MP Biomdicals), and total RNA was isolated and purified using manufacture recommendations for the PureLink RNA kit (Life Technologies).
RNA libraries were prepared for sequencing using standard Illumina protocols and samples were sequenced by PE sequencing with an Illumina HiSeq 2000 sequencer and normalized to TPM.
This dataset was published by Spatz JM et al., 2021 and raw data can be found in the BioProject PRJNA716188.
Experimental condition: Transcriptional analysis of the effects of Hbb gene in cartilage and its function in hypoxia adaption.
Gene: Hbb gene codifies one of the two types of polypeptide chains in adult hemoglobin.
Phenotype: Mice lacking of Hbb gene showed slow growth, shorter length, delayed cartilage hipertrophy with few pattern deffects.
Tissue: Knee cartilaginous tissues.
Age: P6.
Sex: Both.
Layout: Paired.
Reads length: 150 bp.
Sample processing: The cartilaginous tissues were collected and lysed in TRIzol (Invitrogen) for RNA isolation according to the manufacturer’s standard protocol.
An Agilent Bioanalyzer 2100 (Agilent Technologies) was used to check the integrity of the extracted and purified RNA. The TruSeq RNA sample preparation kit (Illumina) was used to generate the libraries.
Libraries were sequenced using an Illumina HiSEq 2500 sequencer and normalized to TPM.
This dataset was published by Zhang et al., 2023 and raw data can be found in the BioProject PRJNA757252.
Experimental condition: Transcriptional analysis of the effects of Kindlin-2 gene in cartilage.
Gene: Kindlins genes codify family of adaptor proteins that are recruited to integrin-containing adhesion sites, termed focal adhesions, Kindlin-2 is ubiquitously expressed.
Phenotype: Mice lacking of Kindlin2 gene showed early development and exacerbated signs of osteoarthritis.
Tissue: Articular cartilage.
Age: 20 week old.
Sex: Both.
Layout: Paired.
Reads length: 150 bp.
Sample processing: Total RNA was extracted from homogenized articular cartilage tissues of control and cKO mice at 5 months after tamoxifen injections using a TransZol Up Plus RNA Kit (Transgen, China). Sequencing libraries were generated using NEBNext R UltraTM RNA Library Prep Kit (Illunina, NEB, United States) and the library quality was assessed on the Agilent Bioanalyzer 2100 system.
After cluster generation, the library preparations were sequenced and 150 bp paired-end reads were generated. Libraries were sequenced using an Illumina NovaSeq 6000 sequencer and normalized to TPM.
This dataset was published by Wu et al., 2022 and raw data can be found in the BioProject PRJNA773746.
Experimental condition: Transcriptional analysis of osteoblast and osteocytes specific deletion of Hnf4α in bone.
Gene: Hepatocyte nuclear factor 4α (HNF4α) is a highly conserved transcription factor. HNF4α is constitutively localized in the nucleus and does not require binding of a ligand to homodimerize and interact with the response elements of its target genes. HNF4α can function as an activator or repressor of genes involved in cell metabolic activity, transport, glucose and lipid homeostasis, and detoxification of xenobiotic.
Phenotype: Since birth, the mice were smaller and hypomineralized. In bones, trabecular part was reduced compared with control littermates. In males, cortical bone was not different, but females suffered a reduction during their growth.
Tissue: Tibiae.
Age: 8 week old.
Sex: Both.
Layout: Single.
Reads length: 50 bp.
Sample processing: RNA from the tibiae of 6 week-old male mice with the epyphises removed and the marrow flushed was isolated RNA from tissues and from cell cultures using TRI Reagent (MRC) and purified RNA using an RNeasy kit (Qiagen).
Libraries were prepared using the TruSeq Total RNA-Seq Library Preparation Kit (Illumina) following the manufacture’s instructions. Then sequencing was performed by SE with an Illumina HiSeq 4000 sequencer and normalised to TPM.
This dataset was published by Martinez-Calle M et al., 2023 and raw data can be found in the BioProject PRJNA786626.
Experimental condition: Transcriptional analysis of the role of Byglican in bone development and regeneration after fracture.
Gene: Biglycan (Bgn) is a member of the SLRP family and is highly abundant in the ECM of a variety of tissues. During skeletal development, high levels of Bgn mRNA are detected in areas of endochondral and membranous bone formation.
Phenotype: Mice lacking of Bgn showed a delay in growth and reduction in cortical thickness and trabecular number, and an increase in trabecular spacing and bone mineral content (BMC). Analysis of serum after suffering a bone fracture showed reduction in monocyte chemoattractant protein 1 (MCP-1) and IL-6 secretion and slow recovering signs in bone.
Tissue: Femorae.
Age: 8 week old.
Sex: Males.
Layout: Paired.
Reads length: 37 bp.
Sample processing: Femoral shafts were isolated and frozen in liquid nitrogen. The bones were centered in a tissue tube (Covaris) under liquid nitrogen and crushed using a CP02 cryoPREP Dry Pulverizer. Total mRNA was extracted and purified from the pulverized tissue using TriPure (Sigma, United States) followed by RNeasy mini kit (Qiagen) hybrid protocol.
RNA was transcribed by Superscript IV (Thermo Fisher Scientific) and full-length 2nd strand cDNA amplified by LongAmp DNA polymerase (New England BioLabs). Sequencing libraries were prepared using a Nextera XT kit (Illumina). Libraries were sequenced using an Illumina NextSeq500 or NextSeq2000 sequencer and normalized to TPM.
This dataset was published by Shainer R et al., 2023 and raw data can be found in the BioProject PRJNA791910.
Experimental condition: Transacriptional analysis of the effects of partial ablation of DTA in osteocytes.
Procedure: Use of a mouse model of conditional deletion of osteocytes by the expression of diphtheria toxin subunit α (DTA) in dentin matrix protein 1 (DMP1)-positive osteocytes.
Phenotype: Mice heterozygotes for DTA showed a delay in growth, loss of bone mass, kyphosis and deformities as they get older. Had more empty lacunae without the presence of osteocytes within cortical and trabecular bone matrix and impairment of osteocyte network. These changes were gender insensitive.
Tissue: Femorae.
Age: 8 week old.
Sex: Both.
Layout: Paired.
Reads length: 92-141 bp.
Sample processing: Total RNA of whole cortical bone with bone marrow flushed out from 4-week WT and DTAhet mice was extracted using TRIzol reagent (Thermo Fisher), quantified and purified using Bioanalyzer 2100 and RNA 6000 Nano LabChip Kit (Agilent).
Following purification, mRNA library were constructed following the vendor’s recommended protocol and sequencing was performed by PE with an Illumina NovaSeq 6000 sequencer and normalised to TPM.
This dataset was published by Ding P et al., 2022 and raw data can be found in the BioProject PRJNA835705.
Experimental condition: Transcriptional analysis of the effects of deletion of Sclerostin gene in an Osteogenesis imperfecta mouse model.
Gene: Sost gene codifies Sclerostin, a secreted molecule that binds to LRP5 on the surface of osteoblasts, whereby it interferes with WNT signaling and thus decreases bone formation.
Phenotype: OI mice lacking of Sost gene showed more bone density and mass that they got older. These mice also showed higher P1NP serum concentrations and lower TRAP5b serum levels than Jrt;Sost-het mice at the age of 8 weeks.
Tissue: Tibiae.
Age: 8 week old.
Sex: Males.
Layout: Paired.
Reads length: 100 bp.
Sample processing: Male mice were euthanized at the age of 8 weeks (n=4 mice per group). Tibia shafts were dissected, the bone marrow was flushed out and the bones were immediately immersed in RNAlater (Thermo Fisher) and stored at − 80 °C until processing. After mincing with scissors, the bones were blotted to remove excess RNAlater and manually grinded using a liquid nitrogen-cooled mini mortar and pestle set, and then transfered to TRIzol. The total RNA was extracted using the phenol–chloroform method.
RNA libraries for RNA-seq were prepared using SMARTER mRNA-Seq Library Prep Kit following manufacturer's protocols. mRNA enrichment was performed using the NEBNext Poly(A) Magnetic Isolation Module (New England Biolabs, Ipswich, MA, USA). cDNA synthesis was achieved with the NEBNext Modules (New England Biolabs). The remaining steps of the library preparation were done using the NEBNext Ultra II DNA Library Prep Kit for Illumina (New England Biolabs)and sequencing was performed by PE with an Illumina NovaSeq 6000 sequencer and normalised to TPM.
This dataset was published by Marulanda J et al., 2022 and raw data can be found in the BioProject PRJNA910516.
Condition: Transcriptome analysis of the effects of meniscus tears in ACL injury Tissue: Anterior Cruciate Ligament (ACL) Age: Between the twenties and thirties Sex: Both
ACL remnants were homogenized with Trizol reagent (Invitrogen, Carlsbad, CA) and the use of a Polytron homogenizer (Kinematica AG, Lucerne, Switzerland). Aliquot of the homogenized suspension was transferred to a microfuge tube and incubated at room temperature for 5 min to permit the complete dissociation of nucleoprotein complexes. After addition of chloroform, the sample was mixed vigorously and the contents were transferred to phase lock gel tube and centrifuged for 15 min at 4°C. The clear aqueous layer was transferred to a fresh tube and one volume of 70% ethyl alcohol was added to precipitate RNA. RNA was collected using RNeasy spin columns (Qiagen Inc., Valencia, CA) according to the instruction manual. RNA was quantified by using a NanoDrop ND-100 spectrophotometer (NanoDrop) while quality was assessed with the use of an Agilent Bioanalyzer (Agilent Technologies, Santa Clara, CA). Libraries were prepared using Sigma SeqPlex (Sigma–Aldrich, St. Louis, MO) kit according to manufacturer's protocol. cDNA libraries were sequenced by SE sequencing with an Illumina HiSeq 3000 sequencer and normalized to TPM
This dataset was published by Brophy RH et al., 2018 and raw data can be found in the BioProject PRJNA430860.
Condition: Transcriptome analysis from muscles obtained from healthy donors and patients suffering Limb-girdle muscular dystrophy R12 (LGMD-R12) Tissue: Three different thigh muscles that are severely (semimembranosus), moderately (vastus lateralis) or mildly (rectus femoris) affected in LGMD-R12 Age: Range from 26-64 year old Sex: Men
RNA was extracted from muscle biopsies using the Direct-zol RNA Miniprep Plus kit ZY-R2073 (Zymo Research, Irvine, CA, USA) according to the kit’s instructions. Samples were treated with DNase prior to their submission to the VIB Nucleomics Core (Leuven, Belgium, www.nucleomics.be). The Nanodrop ND-1000 (Nanodrop Technologies, Wilmington, DE, USA) was used to measure RNA concentration and purity spectrophotometrically. A Bioanalyzer 2100 (Agilent, Santa Clara, CA, USA) was applied to assess RNA integrity. Libraries were prepared using SMART-Seq Stranded Kit (Takara Bio USA, San Jose, CA, USA) and sequenced by PE sequencing using an Illumina NovaSeq 6000 sequencer and normalized to TPM.
This dataset was published by Depuydt CE et al., 2022 and raw data can be found in the BioProject PRJNA837196.
Condition: Transcriptome analysis from hip cartilage of patients with Neck Of femur Fracture (NOF) or Osteoarthritis (OA) undergoing acetabulofemoral joint replacement Tissue: Hip cartilage Age: Range from 70-95 year old (NOF) and 50-85 (OA) Sex: Women
Cartilage samples were ground into powder and homogenized using Invitrogen TRIzol Reagent after which RNA was isolated by RNeasy Kit cDNA libraries were prepared for sequencing using Illumina TrueSeq mRNA kits with the manufacturers’ protocols. The cDNA libraries were sequenced by PE sequencing using an Illumina HiSeq 3000 sequencer and normalized to TPM.
This dataset was published by Ajekigbe B et al., 2019 and raw data can be found in the BioProject PRJNA436709.
Condition: Transcriptome analysis of cartilage from healthy donors and patients suffering osteoarthritis (OA) Tissue: Knee Articular Cartilage Age: Ranging from 18 to 82 Sex: Both
Cartilage was stored at −20°C in Allprotect Tissue Reagent (Qiagen, Valencia, CA) immediately after harvest until RNA extraction. For RNA isolation, cartilage was pulverized using a 6770 Freezer/Mill Cryogenic Grinder (SPEX SamplePrep, Metuchen, NJ), and homogenized in Qiazol Lysis Reagent (Qiagen, Valencia, CA). To remove proteins and cellular debris, a initial phenol-chloroform extraction was performed. RNA purity was assessed using NanoDrop (ND-1000, Thermo Scientific, Wilmington, differentially expressed (DE) genes) and RNA integrity number (RIN) was calculated using a 2100 Bioanalyzer (Agilent, Santa Clara, CA). Libraries were prepared using the Encore Complete RNA-Seq DR Multiplex System 1–8 and 9–16 (NuGen, San Carlos, CA) with 16 unique indexed adapters (L2V6DR-BC2-L2V6DR-BC16). cDNA libraries were sequenced by SE sequencing with an Illumina HiSeq 2000 sequencer and normalized to TPM
This dataset was published by Fisch KM et al., 2018 and raw data can be found in the BioProject PRJNA454873.
Condition: Transcriptome analysis from articular cartilage from healthy and osteoarthritic cartilage Tissue: Knee articular cartilage Age: Range from 25-79 year old Sex: Both
Patients fulfilled the American College of Rheumatology classification criteria for OA of the knee and had no history of knee injury, surgery, rheumatoid arthritis, or pseudogout. Total RNA was isolated from homogenized cartilage using the RNeasy Fibrous Tissue Mini Kit (Qiagen) according to the manufacturer's instructions. The RNA concentration and purity were determined using a NanoDrop spectrophotometer (NanoDrop Technologies). The integrity of the RNA was assessed in an Agilent Bioanalyzer 2100. Samples were prepared for RNA sequencing using BRB-seq protocol following manufacturer's protocols. The cDNA libraries were sequenced by PE sequencing using an Illumina HiSeq 3000 sequencer and normalized to TPM.
This dataset was published by Liao S et al., 2022 and raw data can be found in the BioProject PRJNA857575.
Condition: Transcriptome analysis from patients suffering Chronic Kidney Disease (CKD) with Low Bone Remodeling (LR) and High Bone Remodeling (HR) Renal Osteodystrophy (ROD) Tissue: Iliac crest bone Age: Middle age Sex: Both
Iliac crest bone biopsies from 9 healthy volunteers and 20 patients with CKD, segregated by bone histomorphometry in low (9 biopsies) or high (11 biopsies) bone remodeling groups. RNA was isolated from bone using TRI reagent (Waltham, MA, USA) and purified using RNeasy kit (Qiagen, Germantown, MD, USA). Libraries were prepared according to Illumina's instructions, using the TruSeq Total RNA-Seq Library Preparation Kit (Illumina, San Diego, CA), and the bar-coded cDNA libraries were sequenced by SE sequencing using an Illumina HiSeq 4000 sequencer and normalized to TPM.
This dataset was published by Martinez-Calle M et al., 2023 and raw data can be found in the BioProject PRJNA798962.
