Somatic Cell Nuclear Transfer Embryos Research Articles

Developmental Competence of Somatic Cell Nuclear Transfer Embryos and Interspecies ICSI Zygotes From Bovine Small Antral Follicles.

Assisted reproductive technologies (ART) play a crucial role in conserving threatened wildlife species such as Bos gaurus. ART requires a large number of mature oocytes, and small antral follicles (SAFs) in the ovary are often used to obtain abundant sources of bovine oocytes. However, oocytes from SAFs often experience difficulty completing maturation and obtaining high quality and quantity of blastocyst formation compared to fully grown oocytes. This study aimed to increase the number of high-quality mature oocytes and improve their potential for ART applications in cloned and interspecies intracytoplasmic sperm injection (ICSI) embryos by utilising L-ascorbic acid (LAA) in pre invitro maturation (pre-IVM) culture. First, oocytes isolated from SAFs were cultured with the duration of pre-IVM 0, 6, 8, 10 h and different concentrations of LAA to determine good conditions for oocyte maturation. Then, mature oocytes were assessed for their developmental competence through parthenogenesis, cloned and interspecies ICSI embryos. The results showed that 8-h pre-IVM with 50 μg/mL LAA improved the maturation rate and developmental competence of parthenogenetic and clone embryos, especially, improving the high blastocyst quality by increasing cell number and expression of histone acetylation at lysine 9 (H3K9ac). In addition, the culture process improved the nuclear reprogramming of somatic cells after nuclear transfer into mature oocytes, resulting in an increased hatching rate of cloned embryos. It also enhanced the activation and the pronuclear formation rate of Gaurus-Taurus zygotes. Overall, the established pre-IVM culture method enhanced the meiotic and developmental competence of embryos. This procedure opened hope for the preservation of endangered species and other applications.

Oleanolic acid promotes porcine oocyte maturation by activating the Nrf2/HO-1 signalling pathway

This study investigated the potential role and underlying mechanisms of oleanolic acid (OA), a pentacyclic triterpene with antioxidant and anti-inflammatory properties, in porcine oocytes during in vitro maturation (IVM). The results showed that supplementation with 5 μM OA during IVM resulted in a greater percentage of mature oocytes, parthenogenetically activated embryos and somatic cell nuclear-transferred embryos. This was evidenced by significant increases in the rate of first polar body expulsion, the expansion of cumulus granulosa cells and the total cell number in blastocysts. Further analysis revealed that OA promoted fatty acid accumulation and upregulated the mRNA expression of genes involved in fatty acid β-oxidation. OA significantly increased the intracellular mitochondrial membrane potential and ATP levels and effectively inhibited BAX/BCL2 and Cleaved Caspase3 protein expression. Notably, OA increased the protein levels of intracellular Nrf2 and HO-1, and the GSH levels and the activities of the antioxidant enzymes SOD and catalase (CAT), while reducing ROS levels. Mechanistically, OA activated the Nrf2/HO-1 signalling pathway, which is crucial for regulating the expression of antioxidant-related targets in IVM porcine oocytes. Our findings indicated that OA improved antioxidant capacity by activating the Nrf2/HO-1 signalling pathway, thereby promoting porcine oocyte maturation.

Optimization of donor cell production for somatic cell nuclear transfer in the critically endangered Vietnamese Ỉ pig

We aimed to establish efficient donor cells to produce piglets by somatic cell nuclear transfer (SCNT) of the endangered Vietnamese Ỉ pig. In Experiment 1, we assessed the effects of cell passages on the in vitro development of SCNT embryos. Cells with five and six passages showed significantly cleaved and blastocyst formation rates (86.72 and 86.64; 35.68 and 35.51, respectively, P < 0.05). The highest average total cell number per blastocyst was observed in groups of cells with five and six passages (50.45 and 50.18, respectively). Experiment 2 was performed to assess the sex of donor cells on the subsequent development of SCNT embryos. There was no significant difference in the cleaved and blastocyst formation rates, and the average total cell between female and male groups (86.51 % vs 86.94 % and 35.31 % vs 35.08 %, 50.29 % vs 50.67 %, respectively, P > 0.05). Experiment 3 was performed to assess the effect of cell lines on the development of SCNT embryos. Our results showed no significant difference in the success rate of fibroblast nuclear transfer into recipient oocytes, the cleaved and blastocyst formation rates, and the average total cell number per blastocyst among the cell lines 6004, 9154, 9155, 9156 and 9157 (P > 0.05). Experiment 4 was performed to assess the ability of SCNT embryos to induce pregnancy and to develop term. SCNT embryos were produced from Ỉ fibroblast cells established based on the results of Experiments 1, 2 and 3. Transfer of blastocyst stage embryos into 19 recipients (100–120 embryos in each) resulted in 14 pregnancies, in which 8 pregnant females terminated on Day 22–42 and 6 others produced 20 cloned piglets from donor cells of a female pig but 5 piglets died before birth and 15 healthy cloned piglets. However, 3 out of 15 healthy piglets died of unknown causes within 24h of birth and 3 out of 15 healthy piglets died at 3–5 days of age due to diarrhoea, 9 out of 15 healthy piglets are now 3 months of age. Finally, we established a protocol for the donor cell production which enabled the production of the endangered Ỉ pig embryos by SCNT and maximized blastocyst production rate by more than 35 % and pregnant rate after the transfer of cloned Ỉ pig embryos to recipients at 73.68 % for the first time in Vietnam.

Genome-Scale Analyses Reveal Roadblocks to Monkey Cloning.

Cloning by somatic cell nuclear transfer (SCNT) remained challenging for Rhesus monkeys, mostly due to its low efficiency and neonatal death. Genome-scale analyses revealed that monkey SCNT embryos displayed widespread DNA methylation and transcriptional alterations, thus including loss of genomic imprinting that correlated with placental dysfunction. The transfer of inner cell masses (ICM) from cloned blastocysts into ICM-depleted fertilized embryos rescued placental insufficiency and gave rise to a cloned Rhesus monkey that reached adulthood without noticeable abnormalities.

A retrospective analysis of sheep generated by somatic cell nuclear transfer

Somatic cell nuclear transfer (SCNT) is one of the primary methods for production of genetically engineered sheep, which allows for gene editing or transgene introduction in somatic cells. The use of SCNT eliminates the risk of genetic mosaicism in embryos and animals that is commonly observed after zygote micromanipulations. This retrospective analysis of SCNT in sheep performed at Utah State University, spanning from 2016 to 2021, examined parameters that may impact pregnancy and full-term development, including donor oocytes (donor age), donor cell lines, SCNT parameters (time of oocyte activation following SCNT, number of transferred embryos, in vitro maturation and culture conditions), and recipients (surgical number and ovulatory status), as well as factors that may correlate with large offspring syndrome or abnormal offspring syndrome (LOS/AOS) in the fetuses and lambs. Our findings indicated that compared to prepubertal oocytes, the SCNT embryos produced from adult sheep oocytes had comparable in vitro maturation rates, pregnancy and full-term development rates, as well as SCNT efficiency. In addition, earlier activation time of SCNT embryos (e.g. 24–26 h post maturation) was correlated to the early pregnancy loss rate, full-term rate, and SCNT efficiency. Compared to our standard serum-containing medium, commercial serum-free culture medium showed a positive correlation with the full-term development of sheep SCNT embryos. Transferring 15–30 embryos per recipient resulted in consistently good pregnancy rates. Surgical numbers and ovulatory status (having at least one follicle between 6 and 12 mm in size or a corpus hemorrhagicum (CH)) of recipients did not affect pregnancy and full-term development rates. In summary, this retrospective analysis identified parameters for improving pregnancy and full-term development of SCNT embryos in sheep.

Comparative transcriptome profile of embryos at different developmental stages derived from somatic cell nuclear transfer (SCNT) and in-vitro fertilization (IVF) in riverine buffalo (Bubalus bubalis).

Somatic cell nuclear transfer (SCNT) is a very important reproductive technology with many diverse applications, such as fast multiplication of elite animals, the production of transgenic animals and embryonic stem (ES) cells. However, low cloning efficiency, a low live birth rate and the abnormally high incidence of abnormalities in the offspring born are attributed to incomplete or aberrant nuclear reprogramming. In SCNT embryos, the aberrant expression pattern of the genes throughout embryonic development is responsible for the incomplete nuclear reprogramming. The present study was carried out to identify the differential gene expression (DEGs) profile and molecular pathways of the SCNT and IVF embryos at different developmental stages (2 cell, 8 cell and blastocyst stages). In the present study, 1164 (2 cell), 1004 (8 cell) and 530 (blastocyst stage) DEGs were identified in the SCNT embryos as compared to IVF embryos. In addition, several genes such as ZEB1, GDF1, HSF5, PDE3B, VIM, TNNC, HSD3B1, TAGLN, ITGA4 and AGMAT were affecting the development of SCNT embryos as compared to IVF embryos. Further, Gene Ontology (GO) and molecular pathways analysis suggested, SCNT embryos exhibit variations compared to their IVF counterparts and affected the development of embryos throughout the different developmental stages. Apart from this, q-PCR analysis of the GDF1, TMEM114, and IGSF22 genes were utilized to validate the RNA-seq data. These findings contribute valuable insights about the different genes and molecular pathways underlying SCNT embryo development and offer crucial information for improving SCNT efficiency.

Distinct properties of putative trophoblast stem cells established from somatic cell nuclear-transferred pig blastocysts

BackgroundGenetically modified pigs are considered ideal models for studying human diseases and potential sources for xenotransplantation research. However, the somatic cell nuclear transfer (SCNT) technique utilized to generate these cloned pig models has low efficiency, and fetal development is limited due to placental abnormalities.ResultsIn this study, we unprecedentedly established putative porcine trophoblast stem cells (TSCs) using SCNT and in vitro-fertilized (IVF) blastocysts through the activation of Wing-less/Integrated (Wnt) and epidermal growth factor (EGF) pathways, inhibition of transforming growth factor-β (TGFβ) and Rho-associated protein kinase (ROCK) pathways, and supplementation with ascorbic acid. We also compared the transcripts of putative TSCs originating from SCNT and IVF embryos and their differentiated lineages. A total of 19 porcine TSCs exhibiting typical characteristics were established from SCNT and IVF blastocysts (TSCsNT and TSCsIVF). Compared with the TSCsIVF, TSCsNT showed distinct expression patterns suggesting unique TSCsNT characteristics, including decreased mRNA expression of genes related to apposition, steroid hormone biosynthesis, angiopoiesis, and RNA stability.ConclusionThis study provides valuable information and a powerful model for studying the abnormal development and dysfunction of trophoblasts and placentas in cloned pigs.

Reduction of H3K9 methylation by G9a inhibitors improves the development of mouse SCNT embryos

Removal of somatic histone H3 lysine 9 trimethylation (H3K9me3) from the embryonic genome can improve the efficiency of mammalian cloning using somatic cell nuclear transfer (SCNT). However, this strategy involves the injection of histone demethylase mRNA into embryos, which is limiting because of its invasive and labor-consuming nature. Here, we report that treatment with an inhibitor of G9a (G9ai), the major histone methyltransferase that introduces H3K9me1/2 in mammals, greatly improved the development of mouse SCNT embryos. Intriguingly, G9ai caused an immediate reduction of H3K9me1/2, a secondary loss of H3K9me3 in SCNT embryos, and increased the birth rate of cloned pups about 5-fold (up to 3.9%). G9ai combined with the histone deacetylase inhibitor trichostatin A further improved this rate to 14.5%. Mechanistically, G9ai and TSA synergistically enhanced H3K9me3 demethylation and boosted zygotic genome activation. Thus, we established an easy, highly effective SCNT protocol that would enhance future cloning research and applications.

Treatment of Donor Cells with Oxidative Phosphorylation Inhibitor CPI Enhances Porcine Cloned Embryo Development.

Somatic cell nuclear transfer (SCNT) technology holds great promise for livestock industry, life science and human biomedicine. However, the development and application of this technology is limited by the low developmental potential of SCNT embryos. The developmental competence of cloned embryos is influenced by the energy metabolic status of donor cells. The purpose of this study was to investigate the effects of CPI, an oxidative phosphorylation inhibitor, on the energy metabolism pathways of pig fibroblasts and the development of subsequent SCNT embryos. The results showed that treatment of porcine fibroblasts with CPI changed the cellular energy metabolic pathways from oxidative phosphorylation to glycolysis and enhanced the developmental ability of subsequent SCNT embryos. The present study establishes a simple, new way to improve pig cloning efficiency, helping to promote the development and application of pig SCNT technology.

Monitoring swine virus transmission in embryos derived from commercial abattoir oocytes.

Pigs are pivotal in agriculture and biomedical research and hold promise for xenotransplantation. Specific-pathogen-free (SPF) herds are essential for commercial swine production and xenotransplantation research facilities. Commercial herds aim to safeguard animal health, welfare, and productivity, and research facilities require SPF status to protect immunocompromised patients. Somatic cell nuclear transfer (SCNT) embryos are the norm for producing cloned and genetically edited animals. Oocytes for embryo reconstruction are most conveniently sourced from commercial abattoirs with unclear disease statuses. However, research on viral clearance from donor oocytes during embryo reconstruction remains limited. SCNT has previously been shown to reduce the transmission of Porcine reproductive and respiratory syndrome virus, Bovine viral diarrhea virus, Porcine Circovirus type 2, and Porcine parvovirus. Still, it is lacking for other pathogens, including endogenous viruses. This project contains two preliminary studies investigating the polymerase chain reaction (PCR) assay detection of common swine viruses through the phases of producing parthenogenic and SCNT embryos. Exogenous pathogens detected in oocyte donor tissue or the oocyte maturation media were not detected in the produced embryos. Porcine endogenous retrovirus type C (PERVC) was not removed by parthenogenic embryo activation and was detected in 1 of the 2 tested SCNT embryos reconstructed using a PERVC-negative cell line. SCNT and parthenogenic embryo construction similarly reduced exogenous virus detection. SCNT embryo construction helped reduce endogenous virus detection. This project demonstrates the importance of screening embryos for endogenous viruses and shows the usefulness of parthenogenic embryos in future exogenous virus clearance studies.

Analysis of production efficiency of cloned transgenic Yucatan miniature pigs according to recipient breeds with embryo transfer conditions

The purpose of this study was to compare the efficiency of the production of cloned transgenic Yucatan miniature pigs (YMPs) using two recipient breeds, i.e., YMPs and domestic pigs (DPs), under various embryo transfer conditions. We initially assessed the in vitro developmental competence of embryos obtained via somatic cell nuclear transfer (SCNT) from three different transgenic donor cells. No difference was observed among the three groups regarding developmental competence. Furthermore, the cloning efficiency remained consistent among the three groups after the transfer of the SCNT embryos to each surrogate mother. Subsequently, to compare the efficiency of the production of cloned transgenic YMPs between the two recipient breeds using varying parameters, including ovulation status (preovulation and postovulation), duration of in vitro culture (IVC) (incubated within 24 h and 24–48 h), and the number of transferred SCNT embryos (less than and more than 300), we assessed the pregnancy rates, delivery rates, mean offspring counts, and cloning efficiency. Regarding the ovulation status, YMPs exhibited higher pregnancy rates, delivery rates, and cloning efficiency compared with DPs in both statuses. Moreover, the pregnancy rates, delivery rates, and cloning efficiency were affected by the ovulation status in DPs, but not in YMPs. The comparison of IVC duration between groups revealed that YMPs had higher pregnancy rates vs. DPs in both conditions. SCNT embryos cultured for 24–48 h in YMPs yielded higher delivery rates and cloning efficiency compared with those cultured for less than 24 h in DPs. Finally, the analysis based on the number of transferred SCNT embryos showed that both the pregnancy and delivery rates were higher in YMPs vs. DPs. However, the highest average number of offspring was obtained when more than 300 SCNT embryos were transferred into DPs, whereas the cloning efficiency was higher in YMPs vs. DPs. Our results suggest that YMPs are more suitable recipients than are DPs under various conditions for the production of cloned transgenic YMPs.

Reprogramming mechanism dissection and trophoblast replacement application in monkey somatic cell nuclear transfer.

Somatic cell nuclear transfer (SCNT) successfully clones cynomolgus monkeys, but the efficiency remains low due to a limited understanding of the reprogramming mechanism. Notably, no rhesus monkey has been cloned through SCNT so far. Our study conducts a comparative analysis of multi-omics datasets, comparing embryos resulting from intracytoplasmic sperm injection (ICSI) with those from SCNT. Our findings reveal a widespread decrease in DNA methylation and the loss of imprinting in maternally imprinted genes within SCNT monkey blastocysts. This loss of imprinting persists in SCNT embryos cultured in-vitro until E17 and in full-term SCNT placentas. Additionally, histological examination of SCNT placentas shows noticeable hyperplasia and calcification. To address these defects, we develop a trophoblast replacement method, ultimately leading to the successful cloning of a healthy male rhesus monkey. These discoveries provide valuable insights into the reprogramming mechanism of monkey SCNT and introduce a promising strategy for primate cloning.

Overexpression of BRG1 improves early development of porcine somatic cell nuclear transfer embryos

The epigenetic modification levels of donor cells directly affect the developmental potential of somatic cell nuclear transfer (SCNT) embryos. BRG1, as an epigenetic modifying enzyme, has not yet been studied in donor cells and SCNT embryos. In this study, BRG1 was overexpressed in porcine fetal fibroblasts (PFFs), its effect on chromatin openness and gene transcription was examined, subsequently, the development potential of porcine SCNT embryos was investigated. The results showed that compared with the control group, the percentage of G1 phase cells was significantly increased (32.3 % ± 0.87 vs 25.7 % ± 0.81, P < 0.05) in the experimental group. The qRT-PCR results showed that the expression of H3K9me3-related genes was significantly decreased (P < 0.05), HAT1 was significantly increased (P < 0.05). Assay of Transposase Accessible Chromatin sequencing (ATAC-seq) results revealed that SMARCA4、NANOG、SOX2、MAP2K6 and HIF1A loci had more open chromatin peaks in the experimental group. The RNA-seq results showed that the upregulated genes were mainly enriched in PI3K/AKT and WNT signaling pathways, and the downregulated genes were largely focused on disease development. Interestingly, the developmental rate of porcine SCNT embryos was improved (27.33 % ± 1.40 vs 17.83 % ± 2.02, P < 0.05), the expression of zygotic gene activation-related genes in 4-cell embryos, and embryonic development-related genes in blastocysts was significantly upregulated in the experimental group (P < 0.05). These results suggest that overexpression of BRG1 in donor cells is benefit for the developmental potential of porcine SCNT embryos.

DNA methylation regulates RNA m6A modification through transcription factor SP1 during the development of porcine somatic cell nuclear transfer embryos.

Epigenetic modifications play critical roles during somatic cell nuclear transfer (SCNT) embryo development. Whether RNA N6-methyladenosine (m6A) affects the developmental competency of SCNT embryos remains unclear. Here, we showed that porcine bone marrow mesenchymal stem cells (pBMSCs) presented higher RNA m6A levels than those of porcine embryonic fibroblasts (pEFs). SCNT embryos derived from pBMSCs had higher RNA m6A levels, cleavage, and blastocyst rates than those from pEFs. Compared with pEFs, the promoter region of METTL14 presented a hypomethylation status in pBMSCs. Mechanistically, DNA methylation regulated METTL14 expression by affecting the accessibility of transcription factor SP1 binding, highlighting the role of the DNA methylation/SP1/METTL14 pathway in donor cells. Inhibiting the DNA methylation level in donor cells increased the RNA m6A level and improved the development efficiency of SCNT embryos. Overexpression of METTL14 significantly increased the RNA m6A level in donor cells and the development efficiency of SCNT embryos, whereas knockdown of METTL14 suggested the opposite result. Moreover, we revealed that RNA m6A-regulated TOP2B mRNA stability, translation level, and DNA damage during SCNT embryo development. Collectively, our results highlight the crosstalk between RNA m6A and DNA methylation, and the crucial role of RNA m6A during nuclear reprogramming in SCNT embryo development.

Unreprogrammed H3K9me3 prevents minor zygotic genome activation and lineage commitment in SCNT embryos

Somatic cell nuclear transfer (SCNT) can be used to reprogram differentiated somatic cells to a totipotent state but has poor efficiency in supporting full-term development. H3K9me3 is considered to be an epigenetic barrier to zygotic genomic activation in 2-cell SCNT embryos. However, the mechanism underlying the failure of H3K9me3 reprogramming during SCNT embryo development remains elusive. Here, we perform genome-wide profiling of H3K9me3 in cumulus cell-derived SCNT embryos. We find redundant H3K9me3 marks are closely related to defective minor zygotic genome activation. Moreover, SCNT blastocysts show severely indistinct lineage-specific H3K9me3 deposition. We identify MAX and MCRS1 as potential H3K9me3-related transcription factors and are essential for early embryogenesis. Overexpression of Max and Mcrs1 significantly benefits SCNT embryo development. Notably, MCRS1 partially rescues lineage-specific H3K9me3 allocation, and further improves the efficiency of full-term development. Importantly, our data confirm the conservation of deficient H3K9me3 differentiation in Sertoli cell-derived SCNT embryos, which may be regulated by alternative mechanisms.

Inhibition of Wnt activity improves peri-implantation development of somatic cell nuclear transfer embryos.

Somatic cell nuclear transfer (SCNT) can reprogram differentiated somatic cells into totipotency. Although pre-implantation development of SCNT embryos has greatly improved, most SCNT blastocysts are still arrested at the peri-implantation stage, and the underlying mechanism remains elusive. Here, we develop a 3D in vitro culture system for SCNT peri-implantation embryos and discover that persistent Wnt signals block the naïve-to-primed pluripotency transition of epiblasts with aberrant H3K27me3 occupancy, which in turn leads to defects in epiblast transformation events and subsequent implantation failure. Strikingly, manipulating Wnt signals can attenuate the pluripotency transition and H3K27me3 deposition defects in epiblasts and achieve up to a 9-fold increase in cloning efficiency. Finally, single-cell RNA-seq analysis reveals that Wnt inhibition markedly enhances the lineage developmental trajectories of SCNT blastocysts during peri-implantation development. Overall, these findings reveal diminished potentials of SCNT blastocysts for lineage specification and validate a critical peri-implantation barrier for SCNT embryos.

Chromatin accessibility memory of donor cells disrupts bovine somatic cell nuclear transfer blastocysts development.

The post-transfer developmental capacity of bovine somatic cell nuclear transfer (SCNT) blastocysts is reduced, implying that abnormalities in gene expression regulation are present at blastocyst stage. Chromatin accessibility, as an indicator for transcriptional regulatory elements mediating gene transcription activity, has heretofore been largely unexplored in SCNT embryos, especially at blastocyst stage. In the present study, single-cell sequencing assay for transposase-accessible chromatin (scATAC-seq) of in vivo and SCNT blastocysts were conducted to segregate lineages and demonstrate the aberrant chromatin accessibility of transcription factors (TFs) related to inner cell mass (ICM) development in SCNT blastocysts. Pseudotime analysis of lineage segregation further reflected dysregulated chromatin accessibility dynamics of TFs in the ICM of SCNT blastocysts compared to their in vivo counterparts. ATAC- and ChIP-seq results of SCNT donor cells revealed that the aberrant chromatin accessibility in the ICM of SCNT blastocysts was due to the persistence of chromatin accessibility memory at corresponding loci in the donor cells, with strong enrichment of trimethylation of histone H3 at lysine 4 (H3K4me3) at these loci. Correction of the aberrant chromatin accessibility through demethylation of H3K4me3 by KDM5B diminished the expression of related genes (e.g., BCL11B) and significantly improved the ICM proliferation in SCNT blastocysts. This effect was confirmed by knocking down BCL11B in SCNT embryos to down-regulate p21 and alleviate the inhibition of ICM proliferation. These findings expand our understanding of the chromatin accessibility abnormalities in SCNT blastocysts and BCL11B may be a potential target to improve SCNT efficiency.

A long non-coding RNA essential for early embryonic development improves SCNT efficiency in goats.

Although abundant long non-coding RNAs (lncRNAs) have been found to be highly expressed in early embryos, the functions and mechanisms of most lncRNAs in regulating embryonic development remain unclear. This study was conducted to identify the key lncRNAs during embryonic genome activation (EGA) for promoting embryonic development after somatic cell nuclear transfer (SCNT) in goats. We screened and characterized lncRNAs from transcriptome data of in vitro-fertilized, 2-cell (IVF-2c) and 8-cell embryos (IVF-8c) and 8-cell SCNT embryos (SCNT-8c). We obtained 12 differentially expressed lncRNAs that were highly expressed in IVF-8c embryos compared to IVF-2c, and less expressed in SCNT-8c embryos. After target gene prediction, expression verification, and functional deletion experiments, we found that the expression level of lncRNA3720 affected the early embryonic development in goats. We cloned full length lncRNA3720 and over-expressed it in goat fetal fibroblasts (GFFs). We identified histone variants by analyzing the transcriptome data from both GFFs and embryos. Gene annotation of the gene library and literature search revealed that histone variants may have important roles in early embryo development, so we selected them as the potential target genes for lncRNA3720. Lastly, we compensated for the low expression of lncRNA3720 in SCNT embryos by microinjection, and showed that the development rate and quality of SCNT embryos was significantly improved. We speculate that lncRNA3720 is a key promoter of embryonic development in goats by interacting with histone variants.

HiC4D: forecasting spatiotemporal Hi-C data with residual ConvLSTM.

The Hi-C experiments have been extensively used for the studies of genomic structures. In the last few years, spatiotemporal Hi-C has largely contributed to the investigation of genome dynamic reorganization. However, computationally modeling and forecasting spatiotemporal Hi-C data still have not been seen in the literature. We present HiC4D for dealing with the problem of forecasting spatiotemporal Hi-C data. We designed and benchmarked a novel network and named it residual ConvLSTM (ResConvLSTM), which is a combination of residual network and convolutional long short-term memory (ConvLSTM). We evaluated our new ResConvLSTM networks and compared them with the other five methods, including a naïve network (NaiveNet) that we designed as a baseline method and four outstanding video-prediction methods from the literature: ConvLSTM, spatiotemporal LSTM (ST-LSTM), self-attention LSTM (SA-LSTM) and simple video prediction (SimVP). We used eight different spatiotemporal Hi-C datasets for the blind test, including two from mouse embryogenesis, one from somatic cell nuclear transfer (SCNT) embryos, three embryogenesis datasets from different species and two non-embryogenesis datasets. Our evaluation results indicate that our ResConvLSTM networks almost always outperform the other methods on the eight blind-test datasets in terms of accurately predicting the Hi-C contact matrices at future time-steps. Our benchmarks also indicate that all of the methods that we benchmarked can successfully recover the boundaries of topologically associating domains called on the experimental Hi-C contact matrices. Taken together, our benchmarks suggest that HiC4D is an effective tool for predicting spatiotemporal Hi-C data. HiC4D is publicly available at both http://dna.cs.miami.edu/HiC4D/ and https://github.com/zwang-bioinformatics/HiC4D/.

Lysophosphatidic acid improves development of porcine somatic cell nuclear transfer embryos

Lysophosphatidic acid improves development of porcine somatic cell nuclear transfer embryos