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Additional file 3: Figure S1. Overview of Luchuan genome assembly. (a) The flowchart of the contig, scaffold, and chromosome assembly in this study; and (b) Hi-C contact heatmap. Genome-wide analysis of chromatin interactions in Luchuan genome. The numbering of a chromosome is based on the chromosome length from the longest to the shortest. Figure S2. Circos plot shows the genome characterization of the Luchuan and Duroc pigs. (I) Syntenic alignments between the Luchuan and Duroc genome based on one-to-one orthologous genes; (II) GC content in non-overlapping 1 Mb windows; (III) Percent coverage of TEs in non-overlapping 1 Mb windows; (IV) Gene density calculated on the basis of the number of genes in non-overlapping 1 Mb windows; and (V) The length of pseudo-chromosome, light red, and light purple ideograms represent the chromosomes of Luchuan and Duroc pigs, respectively. Each ticket above the ideograms is on a scale of 50 Mb. Figure S3. Comparative genomic and phylogenetic analyses. (a) Venn diagram showing shared orthologous groups among genomes of Luchuan, Duroc, goat, and human; and (b) Phylogenetic tree with divergence times and history of orthologous gene families. Numbers on the nodes represent divergence times, with the error range shown in parentheses. The numbers of gene families that expanded (red) or contracted (blue) in each lineage after speciation are shown on the corresponding branch. MRCA is the most recent common ancestor. Figure S4. PCA plots. (a) PCA of the first three principal components. BAMEI, Bamei; BER, Berkshire; BMX, Bamaxiang; CWB, Chinese wild boar; DUR, Duroc; EHL, Erhualian; GST, Tibetan (Gansu); HAM, Hampshire; HTDE, Hetao; JH, Jinhua; KBP, Korean black pig; KWB, Korean wild boar; LD, Landrace; LUC, Luchuan; LW, Large White; LWH, Laiwu; MEI, Meishan; MIN, Min; PTL, Pietrain; RC, Rongchang; SCT, Tibetan (Sichuan); TC, Tongcheng; TT, Tibetan (Tibet); WZS, Wuzhishan; YM, Yucatan Miniature; YNT, Tibetan (Yunnan); and (b) PCA plots of the first two principal components by classifying them into seven groups: Chinese wild boar (CWB), Korean wild boar (BWB), Luchuan pig (LUC), Duroc (DUR), Korean black pig (KBP), other eastern pigs (EST) and western pigs (WST). Figure S5. (a) LD decay determined by squared correlations (r2) of allele frequencies against the distance between polymorphic sites in eastern (EST; orange) and western (WST; green) pig groups. (b) LD decay in DUR (Duroc; red) and LUC (Luchuan; blue) pig groups. Figure S6. Enrichment pattern of genes under potential selection in GWAS regions associated with some economic traits. Red represents genes located in selective regions of eastern pigs while blue represents those of western pigs. * represents FDR < 0.05. ** represents FDR < 0.01. Figure S7. GO enrichment of differentially expressed genes between Luchuan and Duroc pigs in 10 tissues. GO biological process analysis of the differentially expressed mRNAs between Luchuan and Duroc pigs in each tissue. Up indicates the genes with expression levels that are higher in Luchuan pigs compared to Duroc pigs. Down indicates the genes with expression levels that are lower in Luchuan pigs compared to Duroc pigs. Figure S8. Analysis of DEG in skeletal muscle tissues. (a) The number of DEG in different tissues. Duroc: the expression level is higher in Duroc pigs compared to Luchuan pigs. Luchuan: the expression level is higher in Luchuan pigs compared to Duroc pigs; (b) Heatmap showing the expression of DEG related to “muscle contraction” in skeletal muscle tissues; and (c) Heatmap showing the expression of DEG related to “cell cycle” in skeletal muscle tissues. Figure S9. Frequencies of the Luchuan promoter allele across different pig breeds. DUR: Duroc pig; LW: Large White pig; LD: Landrace pig; YM: Yucatan miniature pig; YNT: Tibetan pig (Yunnan); TC: Tongcheng pig; TT: Tibetan pig (Qinghai-Tibetan); HTDE: Hetaodaer pig; SCT: Tibetan pig (Sichuan); WZS: Wuzhishan pig; BMX: Bamaxiang pig; EHL: Erhualian pig; LUC: Luchuan pig; LWH: Laiwu black pig; MIN: Min pig (Shandong). Figure S10. QC of ATAC-seq peaks. Heatmap of reads density around TSS for five tissues. Figure S11. Muscle fiber differences between Duroc and Luchuan pigs. Proportions of slow muscle fiber. Figure S12. Knockdown and overexpression efficiency of Tnnc1 in C2C12 cells and in vivo. (a) Tnnc1 expression after siRNA knockdown in C2C12 cells; (b) Tnnc1 expression after overexpression in C2C12 cells; and (c) Tnnc1 expression after in vivo siRNA knockdown in mouse skeletal muscle. Figure S13. (a) Expression of Ho-1 after overexpression of Tnnc1; (b) Expression of Ho-1 after siRNA knockdown of Tnnc1; (c) Expression of Ogg1 after overexpression of Tnnc1; and (d) Expression of Ogg1 after siRNA knockdown of Tnnc1. Figure S14. (a) Expression of slow muscle (Myh7) and fast muscle (Myh2, Myh4, Myh1) markers after siRNA knockdown of Sema3g in C2C12 cells; (b) Measuring the same markers as (a) after overexpression of Sema3g in C2C12 cells; (c) Choosing siRNAs with the best knockdown efficiency; (d) Evaluation of the overexpression efficiency; and (e) Comparing the expression level of Sema3g before and after knocking down Cebpa. * represents p-value < 0.05. ** represents p-value < 0.01.

Additional file 1. Additional materials and methods [20, 21, 24–52, 54–57, 59–66, 118–130]

The maternal-to-embryonic transition (MET) is a complex process that occurs during early mammalian embryogenesis and is characterized by activation of the zygotic genome, initiation of embryonic transcription, and replacement of maternal mRNA with embryonic mRNA. The objective of this study was to reveal the temporal expression and localization patterns of PTTG1 during early porcine embryonic development and to establish a relationship between PTTG1 and the MET. To achieve this goal, reverse transcription-polymerase chain reaction (RT-PCR) was performed to clone porcine PTTG1. Subsequently, germinal vesicle (GV)- and metaphase II (MII)-stage oocytes, zygotes, 2-, 4-, and 8-cell-stage embryos, morulas, and blastocysts were produced in vitro and their gene expression was analyzed. The results revealed that the coding sequence of porcine PTTG1 is 609-bp in length and that it encodes a 202-aa polypeptide. Using qRT-PCR, PTTG1 mRNA expression was observed to be maintained at high levels in GV- and MII-stage oocytes. The transcript levels in oocytes were also significantly higher than those in embryos from the zygote to blastocyst stages. Immunohistochemical analyses revealed that porcine PTTG1 was primarily localized to the cytoplasm and partially localized to the nucleus. Furthermore, the PTTG1 protein levels in MII-stage oocytes and zygotes were significantly higher than those in embryos from the 2-cell to blastocyst stage. After fertilization, the level of this protein began to decrease gradually until the blastocyst stage. The results of our study suggest that porcine PTTG1 is a new candidate maternal effect gene (MEG) that may participate in the processes of oocyte maturation and zygotic genome activation during porcine embryogenesis.

Somatic cloning, also known as somatic cell nuclear transfer (SCNT), is a promising technology which has been expected to rapidly extend the population of elaborately selected breeding boars with superior production performance. Chinese Guike No. 1 pig breed is a novel swine specialized strain incorporated with the pedigree background of Duroc and Chinese Luchuan pig breeds, thus inherits an excellent production performance. The present study was conducted to establish somatic cloning procedures of adult breeding boars from the Chinese Guike No. 1 specialized strain. Ear skin fibroblasts were first isolated from a three-year-old Chinese Guike No. 1 breeding boar, and following that, used as donor cell to produce nuclear transfer embryos. Such cloned embryos showed full in vitro development and with the blastocyst formation rate of 18.4 % (37/201, three independent replicates). Finally, after transferring of 1187 nuclear transfer derived embryos to four surrogate recipients, six live piglets with normal health and development were produced. The overall cloning efficiency was 0.5 % and the clonal provenance of such SCNT derived piglets was confirmed by DNA microsatellite analysis. All of the cloned piglets were clinically healthy and had a normal weight at 1 month of age. Collectively, the first successful cloning of an adult Chinese Guike No. 1 breeding boar may lay the foundation for future improving the pig production industry.