Project description:The giant panda (Ailuropoda melanoleuca) stands as a flagship and umbrella species, symbolizing global biodiversity. While traditional assisted reproductive technology faces constraints in safeguarding the genetic diversity of giant pandas and bolstering the population size of giant pandas, induced pluripotent stem cells (iPSCs) known for their capacity to differentiate into diverse cells types, including germ cells, present a transformative potential for conservation of endangered animals. In our study, we isolated primary fibroblast cells from an individual giant panda and successfully generated giant panda induced pluripotent stem cells (GPiPSCs) through a non-integrating episomal vectors reprogramming method. Characterization of these GPiPSCs revealed their state of primed pluripotency and demonstrated their potential for differentiation. Furthermore, we innovatively formulated a species-specific chemically defined FACL medium and unraveled the intricate signaling pathway networks responsible for maintaining the pluripotency and fostering cell proliferation of GPiPSCs. This study provides key insights into rare species iPSCs, offering materials for panda characteristics research and laying the groundwork for in vitro giant panda gamete generation, potentially aiding endangered species conservation.
Project description:Next generation sequencing reveal profiles in H3K27me3 and expression levels between WT and panda mutant, anti-Flag ChIP reveal the binding sites of PANDA protein
Project description:Next generation sequencing reveal profiles in H3K27me3 and expression levels between WT and panda mutant, anti-Flag ChIP reveal the binding sites of PANDA protein
Project description:Purpose:To present the miRNA expression profiles in giant panda milk exosomes across five lactation stages (0, 3, 7, 15 and 30 days after birth), aiming to provide new information for investigations into the physiological functions of the giant panda milk Methods: Three females were sampled in all, and each individual were sampled over multiple lactations, including 0, 3, 7, 15 and 30 days after delivery. Breast milk samples (5-10 ml) were collected from each stages. Total RNA isolated from individuals in five lactation stages (0, 7, 15 and 30 days after delivery) were pooled in equal quantities for each stage Results: Here, we illustrated the species and expression characteristics of exosome-loaded miRNAs existing in giant panda breast milk during distinct lactation periods, and highlighted the enrichment of immune- and development-related endogenous miRNAs in colostral and mature giant panda milk, which are stable even in certain hash conditions, like low pH and high concentration of RNAase, by the protection of extracellular vesicles.These findings indicate that breast milk may allow dietary intake of maternal miRNAs by infants for the regulation of postnatal development. We also demonstrated that the exogenous plant miRNA from the primary food source of giant panda (bamboo) were detected in the exosomes of giant panda breast milk, which were predicted to be of regulatory role in basic cell metabolism and neuron development. This result suggested that the dietary plant miRNAs were able to be absorbed by host cell and then secreted to body fluids as potential cross-kingdom regulators. Conclusions: Exosomal miRNAs in the giant panda breast milk may be the crucial maternal regulators for the development of intrinsic ‘slink’ newborn cubs.
Project description:In this study, differentially expressed (DE)piRNAs of fresh and frozen-thawed sperm with different freeze tolerance compacity from giant panda and boar were evaluated. The results showed 1160 (22 down-regulated and 1138 up-regulated) and 384 (110 up-regulated and 274 down-regulated) differentially expressed (DE) piRNAs were identified in giant panda and boar sperm, respectively. Gene ontology (GO) enrichment analysis revealed that the target DE mRNAs of DE piRNAs were mainly enriched in biological regulation, cellular process and metabolic process in giant panda and boar sperm. Moreover, Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated that the target DE mRNAs of DE piRNAs were only distributed on DNA replication and cAMP signaling pathway in giant panda, but cAMP, cGMP and MAPK signaling pathway in boar sperm which were considered as part of olfactory transduction pathway.Conclusion:Olfactory transduction related pathways maybe contributed to different freeze tolerance compacity between giant panda and boar sperm, which will benefit to further understand the molecular mechanism of sperm cryoinjury and freezability.
Project description:While various methods exist for examining and visualizing the structure of RNA molecules, dimethyl sulfate-mutational profiling and sequencing (DMS-MaPseq) stands out for its simplicity and versatility. This technique has proven effective for studying RNA structures both in vitro and in complex biological settings. We've updated the protocol for using DMS-MaPseq, and it can also be employed to identify the binding of antisense oligonucleotides (ASOs) to RNA. By applying this updated protocol, we successfully characterized the structural ensemble of the HIV1 Rev Response Element (RRE), along with its two alternative structures. The findings align with previously published research. Additionally, we resolved the structure of the long non-coding RNA PANDA, which was previously unknown. Moreover, we used PANDA as a basis for designing ASOs and confirmed their binding through a substantial decrease in DMS-reactivities at the anticipated ASO binding locations.
Project description:Gene differential expression studies can serve to explore and understand the laws and 16 characteristics of animal life activities, and the difference in gene expression between different 17 animal tissues have been well demonstrated and studied. However, for the world-famous rare 18 and protected species giant panda (Ailuropoda melanoleuca), only the transcriptome of the blood 19 and spleen has been reported separately. Here, in order to explore the transcriptome differences 20 between the different tissues of the giant panda, transcriptome profiles of the heart, liver, spleen, 21 lung, and kidney from five captive giant pandas were constructed with Illumina HiSeq 2500 22 platform. The comparative analysis of the inter-tissue gene expression patterns was carried out 23 based on the generated RNA sequencing datasets. Analyses of Gene Ontology (GO) enrichment, 24 Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment, and protein-protein interaction 25 (PPI) network were performed according to the identified differentially expressed genes (DEGs). 26 We generated 194.52 GB clean base data from twenty-five sequencing libraries and identified 27 18,701 genes, including 3492 novel genes. With corrected p-value < 0.05 and |log2FoldChange| > 28 2, we finally obtained 921, 553, 574, 457, and 638 tissue-specific DEGs in the heart, liver, spleen, 29 lung, and kidney, respectively. In addition, we identified TTN, CAV3, LDB3, TRDN, and 30 ACTN2 in the heart; FGA, AHSG, and SERPINC1 in the liver; CD19, CD79B, and IL21R in the 31 spleen; NKX2-4 and SFTPB in the lung; GC and HRG in the kidney as hub genes in the PPI 32 network. The results of the analyses showed a similar gene expression pattern between the spleen 33 and lung. This study provided for the first time the heart, liver, lung, and kidney’s transcriptome 34 resources of the giant panda, and it provided a valuable resource for further genetic research or 35 other potential research.