Transcriptome analysis of PCOS arrested 2-cell embryos

Figures & data

Table 1. Summary of the sequence assembly after Illumina sequencing.

Sample name	Raw reads	Clean reads	Clean bases	Error rate (%)	Q20 (%)	Q30 (%)	GC content (%)
Normal 2-cell 1	44346717	34559454	3.46G	0.04	97.07	91.28	43.58
Normal 2-cell 2	43138152	33947088	3.40G	0.04	97.13	91.38	43.70
Normal 2-cell 3	47009259	36844353	3.69G	0.04	96.98	90.88	43.39
Non-PCOS arrest 1	33403676	22702894	2.83G	0.03	96.46	92.70	49.45
Non-PCOS arrest 2	37950082	23810672	2.97G	0.03	95.97	91.74	46.20
Non-PCOS arrest 3	69520534	26313142	3.28G	0.05	96.02	91.98	48.06
PCOS arrest 1	50948968	26618254	3.32G	0.04	95.88	91.60	48.36
PCOS arrest 2	63678672	23465086	2.92G	0.04	96.16	92.26	47.94
PCOS arrest 3	85369950	72645034	9.08G	0.03	95.80	91.42	46.33

Notes: Q20, percentage of bases with a Phred value of at least 20; Q30, percentage of bases with a Phred value of at least 30.

Figure 1. Count of differentially expressed genes between non-PCOS arrested and non-arrested 2-cell embryos (a), PCOS arrested and non-arrested 2-cell embryos (b) and between non-PCOS and PCOS arrested embryos (c). The x-axis shows the fold-change in gene expression, and the y-axis shows significant statistical differences. Red, up-regulated genes; green, down-regulated genes.

Figure 2. Cluster analysis of DEGs. A, hierarchical cluster analysis of DEGs of PCOS, non-PCOS arrested and non-arrested 2-cell embryos; B, Venn diagram showing the overlaps between groups. Red, high gene expression levels; blue, low gene expression levels.

Table 2. Functional annotation of DEGs in the down-regulated genes between PCOS arrested and 2-cell embryo.

GO accession	Description	Term types	Adjusted P values
GO:0005622	intracellular	Cellular component	5.86E-07
GO:0005623	cell	Cellular component	2.56E-06
GO:0044464	cell part	Cellular component	2.56E-06
GO:0009058	biosynthetic process	Biological process	0.0047284
GO:0003924	GTPase activity	Molecular function	0.0063557
GO:0008152	metabolic process	Biological process	0.0065239
GO:0044249	cellular biosynthetic process	Biological process	0.0065239
GO:1901576	organic substance biosynthetic process	Biological process	0.0065239
GO:0005737	cytoplasm	Cellular component	0.010424
GO:0044237	cellular metabolic process	Biological process	0.015558
GO:0044271	cellular nitrogen compound biosynthetic process	Biological process	0.034668
GO:0071704	organic substance metabolic process	Biological process	0.035282
GO:0018130	heterocycle biosynthetic process	Biological process	0.036328
GO:0043227	membrane-bounded organelle	Cellular component	0.036328
GO:0003824	catalytic activity	Molecular function	0.04082
GO:0043231	intracellular membrane-bounded organelle	Cellular component	0.04082
GO:0005525	GTP binding	Molecular function	0.04082
GO:0032561	guanyl ribonucleotide binding	molecular_function	0.041541
GO:0097159	organic cyclic compound binding	Molecular function	0.045173
GO:1901362	organic cyclic compound biosynthetic process	Biological process	0.045967
GO:0007018	microtubule-based movement	Biological process	0.045967
GO:0017111	nucleoside-triphosphatase activity	Molecular function	0.045967
GO:0019438	aromatic compound biosynthetic process	Biological process	0.045967
GO:0019001	guanyl nucleotide binding	Molecular function	0.045967
GO:0046148	pigment biosynthetic process	Biological process	0.04854
GO:0042440	pigment metabolic process	Biological process	0.04854
GO:1901363	heterocyclic compound binding	Molecular function	0.04854
GO:0010467	gene expression	Biological process	0.04854
GO:0016818	hydrolase activity, acting on acid anhydrides, in phosphorus- containing anhydrides	Molecular function	0.04854

Table 3. Functional annotation of DEGs in the up-regulated genes between PCOS and non-PCOS arrested embryo.

GO accession	Description	Term types	Adjusted P values
GO:0003676	nucleic acid binding	Molecular function	4.99E-07
GO:0043565	sequence-specific DNA binding	Molecular function	3.19E-06
GO:0005488	binding	Molecular function	3.19E-06
GO:0016070	RNA metabolic process	Biological process	1.44E-05
GO:0003677	DNA binding	Molecular function	3.58E-05
GO:0090304	nucleic acid metabolic process	Biological process	5.98E-05
GO:0006351	transcription, DNA-dependent	Biological process	0.00018512
GO:0032774	RNA biosynthetic process	Biological process	0.00019373
GO:0008270	zinc ion binding	Molecular function	0.00019373
GO:1901363	heterocyclic compound binding	Molecular function	0.00019373
GO:0097159	organic cyclic compound binding	Molecular function	0.00019373
GO:0034654	Nucleobase containing compound biosynthetic process	Biological process	0.00057696
GO:0006139	nucleobase-containing compound metabolic process	Biological process	0.00057696
GO:2000112	regulation of cellular macromolecule biosynthetic process	Biological process	0.00057696
GO:0010556	regulation of macromolecule biosynthetic process	Biological process	0.00057696
GO:0031326	regulation of cellular biosynthetic process	Biological process	0.00057696
GO:0009889	regulation of biosynthetic process	Biological process	0.00057696
GO:0006355	regulation of transcription, DNA-dependent	Biological process	0.00057696
GO:0051252	regulation of RNA metabolic process	Biological process	0.00057696
GO:2001141	regulation of RNA biosynthetic process	Biological process	0.00057696
GO:0010468	regulation of gene expression	Biological process	0.00073122
GO:0060255	regulation of macromolecule metabolic process	Biological process	0.0011985
GO:0019438	aromatic compound biosynthetic process	Biological process	0.0014888
GO:0046483	heterocycle metabolic process	Biological process	0.0015037
GO:0006725	cellular aromatic compound metabolic process	Biological process	0.0015037
GO:0018130	heterocycle biosynthetic process	Biological process	0.0019823
GO:0031323	regulation of cellular metabolic process	Biological process	0.0021782
GO:1901360	organic cyclic compound metabolic process	Biological process	0.0022871
GO:0044271	cellular nitrogen compound biosynthetic process	Biological process	0.0022949
GO:0019219	regulation of nucleobase-containing compound metabolic process	Biological process	0.0022949
GO:0051171	regulation of nitrogen compound metabolic process	Biological process	0.0022949
GO:0080090	regulation of primary metabolic process	Biological process	0.0024321
GO:0019222	regulation of metabolic process	Biological process	0.0024394
GO:1901362	organic cyclic compound biosynthetic process	Biological process	0.0024457
GO:0001071	nucleic acid binding transcription factor activity	Molecular function	0.0025382
GO:0003700	sequence-specific DNA binding transcription factor activity	Molecular function	0.0025382
GO:0034641	cellular nitrogen compound metabolic process	Biological process	0.0035068
GO:0046872	metal ion binding	Molecular function	0.0057461
GO:0043169	cation binding	Molecular function	0.0068433
GO:0046914	transition metal ion binding	Molecular function	0.0071118
GO:0010467	gene expression	Biological process	0.0093075
GO:0022613	ribonucleoprotein complex biogenesis	Biological process	0.011344
GO:0006807	nitrogen compound metabolic process	Biological process	0.014291
GO:0006367	transcription initiation from RNA polymerase II promoter	biological process	0.016731
GO:0042254	ribosome biogenesis	Biological process	0.025702
GO:0005515	protein binding	Molecular function	0.029572
GO:0006396	RNA processing	Biological process	0.04445
GO:0005634	nucleus	Cellular component	0.044722
GO:0005622	intracellular	Cellular component	0.046387

Figure 3. GO enrichment in biological process analysis of DEGs. The x-axis shows −log10 (pvalue), and the y-axis shows the five representative GO terms in biological process. A, down-regulated genes between non-PCOS arrested and non-arrested 2-cell embryos; B, up-regulated genes between non-PCOS arrested and non-arrested 2-cell embryos; C, down-regulated genes between non-PCOS and PCOS arrested embryos; D, up-regulated genes between non-PCOS and PCOS arrested embryos; E, down-regulated genes between PCOS arrested and non-arrested 2-cell embryos; F, up-regulated genes between PCOS arrested and non-arrested 2-cell embryos.

Figure 4. KEGG enrichment analyses of DEGs. The ratio of the number of DEGs to the total gene number is represented by the enrichment factor. Size of dots: gene number; colour of dots: range of P-values. A, representative terms between non-PCOS arrested and non-arrested 2-cell embryos in down-regulated genes; B, representative terms between non-PCOS arrested and non-arrested 2-cell embryos in up-regulated genes; C, representative terms between non-PCOS and PCOS arrested embryos in down-regulated genes; D, representative terms between non-PCOS and PCOS arrested embryos in up-regulated genes; E, representative terms between PCOS arrested and non-arrested 2-cell embryos in down-regulated genes; F, representative terms between PCOS arrested and non-arrested 2-cell embryos in up-regulated genes.

Figure 5. Representative cell cycle pathway between PCOS arrested and non-arrested 2-cell embryos. Red, down-regulated gene names.

Figure 6. Relative expression of 5 selected genes by Q-PCR. The relative gene expression levels were calculating using the 2^−ΔΔCt method. R.Q., relative expression; *P < 0.05 indicates that the genes were significantly down-regulated in PCOS and non-PCOS arrested embryos relative to the non-arrested 2-cell embryo.