Cryopreservation technology has become a primary method for preserving animal genetic resources in the biomedical field. It achieves long-term storage by placing gametes or embryos in an ultra-low-temperature environment, enabling them to retain biological activity after thawing. However, the cytotoxicity issues associated with traditional cryoprotectants limit their application, making the development of non-toxic and highly effective novel cryoprotectants a current research priority. This review systematically reviews the development history and principles of cryopreservation technology, and summarizes the classification and mechanisms of action of cryoprotectants, with a particular focus on five types of novel cryoprotectants featuring bioinspired structural characteristics. Their ice-suppression performance and mechanisms of action are analyzed. These protectants possess structures similar to those of natural antifreeze substances and exhibit low toxicity, thereby demonstrating higher cryopreservation efficiency, and are thus referred to as novel bioinspired cryoprotectants. Furthermore, this review examines the potential applications of the aforementioned protectants in the cryopreservation of animal gametes (sperm, oocytes, and embryos), offering new insights into expanding the use of bioinspired cryoprotectants in the preservation of animal genetic resources.

In mammals, imprinted genes are characterised by a monoallelic expression, which is based on parental origin and is essential for both foetal and placental development. The ZFAT gene encodes a transcriptional factor, and its non-coding antisense RNA, ZFAT-AS1, overlaps with the ZFAT locus. Both ZFAT and ZFAT-AS1 are maternally imprinted in human placentas. In bovines, the imprinting status of the ZFAT and ZFAT-AS1 genes has yet to be reported. In this study, we analysed the allelic expression of three transcript variants (X1-X3) of the bovine ZFAT and ZFAT-AS1 genes in somatic tissues and placentas using a single nucleotide polymorphism-based method. The results showed that bovine ZFAT exhibited isoform-specific paternal expression. The ZFAT X2 variant exhibited monoallelic expression in the bovine placentas and biallelic expression in the six bovine somatic tissues (heart, liver, spleen, lung, kidney and brain). However, the ZFAT X1 and X3 variants were biallelically expressed in both bovine tissues and placentas. A 311 bp bovine ZFAT-AS1 complementary DNA (cDNA) sequence was obtained by aligning the human ZFAT-AS1 cDNA sequence with the bovine genome and conducting reverse transcription polymerase chain reaction amplification. Bovine ZFAT-AS1 have monoallelic expression in bovine placentas and somatic tissues. In addition, the DNA methylation of two regions was characterised, including the partial promoter, and exon 1 and intron 1 regions of ZFAT, and there were no differentially methylated regions.

In the face of increasingly serious environmental problems, such as global warming, passive daytime radiative cooling (PDRC) technology offers a green solution. The process involving the radiation of heat to outer space through transparent windows in the atmosphere and spontaneous reflection of solar radiation does not consume energy. However, when used in outdoor applications, radiative cooling materials are often disturbed by environmental factors, such as rain and dust. The disturbance caused can affect the cooling performance of the materials. Moreover, to facilitate the widespread use of radiative cooling, new requirements for the stability of the materials used have been proposed. In this study, a PDRC coating with superhydrophobic properties was prepared by combining ethylene tetrafluoroethylene and multi-scale Al2O3 particles, with complementary emission performances in an atmospheric window. The PDRC coating exhibited self-cleaning properties with a high solar reflectivity (95.04 %) and a high atmospheric window emissivity (95.43 %). The water contact angle of the PDRC coating surface was 158.81 degrees. The coating could achieve a temperature drop of 9.2 degree celsius and a cooling power of 98.6 W/m2 under direct outdoor sunlight conditions. The PDRC coatings exhibit good chemical and mechanical stability and are suitable for use in a variety of complex working environments. Our study demonstrated a long-term outdoor cooling strategy that can be applied to buildings and chemical storage tanks.

Genomic imprinting is an epigenetic regulation in mammals in which a small subset of genes is monoallelically expressed dependent on their parental origin. A large imprinted domain, SGCE/PEG10 locus, is located on human chromosome 7q21s and mouse proximal chromosome 6. However, genomic imprinting of bovine SGCE/PEG10 cluster has not been systematically studied. In this study, we investigated allele expression of 14 genes of the SGCE/PEG10 locus in bovine somatic tissues and term placenta using a single nucleotide polymorphism (SNP)-based sequencing method. In addition to SGCE and PEG10, two conserved paternally expressed genes in human and mice, five other genes (TFPI2, GNG11, ASB4, PON1, and PON3) were paternally expressed. Three genes, BET1, COL1A2, and CASD1, exhibited tissue-specific monoallelic expression. CALCR showed monoallelic expression in tissues but biallelic expression in the placenta. Three genes, GNGT1, PPP1R9A, and PON2, showed biallelic expression in cattle. Five differentially methylated regions (DMRs) were found to be associated with the allelic expression of TFPI2, COL1A2, SGCE/PEG10, PON3, and ASB4 genes, respectively. The SGCE/PEG10 DMR is a maternally hypermethylated germline DMR, but TFPI2, COL1A2, PON3, and ASB4 DMRs are secondary DMRs. In summary, we identified five novel bovine imprinted genes (GNG11, BET1, COL1A2, CASD1, and PON1) and four secondary DMRs at the SGCE/PEG10 locus.

Passive daytime radiative cooling (PDRC) technology has received a great deal of attention in the field of energy efficiency and environmental protection as a sustainable technology and a large-scale and promising solution to mitigate the environmental impact of global warming. In this study, we prepared PDRC material by combining FEP with modified Al2O3 particles and using the method of spray combined with phase separation. The synergistic effect of the formed surface micronanostructures, combined with the molecular vibration of FEP and the phonon polarization resonance of Al2O3, further improves the optical performance of the PDRC coating. The PDRC coating has an average reflectivity of 0.96 in the solar spectral band (0.3-2.5 mu m) and an average emissivity of 0.963 in the atmospheric window band ((8-13 mu m). In addition, the PDRC coating had good hydrophobicity, and its water contact angle (WAC) reached 159.3 degrees. Under direct sunlight conditions, PDRC materials have a good temperature drop (4.9 degrees C) compared to ambient temperatures and radiative cooling power (81.2 W/m(2)). The prepared coating maintains superhydrophobicity and excellent cooling performance when soaked in solutions of different pH values and UV radiation, which was of great significance for sustainable applications. Our work provides a form of long-term cooling that can be effectively implemented in green and energy-efficient buildings.

Genomic imprinting is an epigenetic regulation mechanism in mammals resulting in the parentally dependent monoallelic expression of genes. Imprinting disorders in humans are associated with several congenital syndromes and cancers and remain the focus of many medical studies. Cattle is a better model organism for investigating human embryo development than mice. Imprinted genes usually cluster on chromosomes and are regulated by different methylation regions (DMRs) located in imprinting control regions that control gene expression in cis. There is an imprinted locus on human chromosome 16q24.1 associated with congenital lethal developmental lung disease in newborns. However, genomic imprinting on bovine chromosome 18, which is homologous with human chromosome 16 has not been systematically studied. The aim of this study was to analyze the allelic expressions of eight genes (CDH13, ATP2C2, TLDC1, COTL1, CRISPLD2, ZDHHC7, KIAA0513, and GSE1) on bovine chromosome 18 and to search the DMRs associated gene allelic expression. Three transcript variants of the ZDHHC7 gene (X1, X2, and X5) showed maternal imprinting in bovine placentas. In addition, the monoallelic expression of X2 and X5 was tissue-specific. Five transcripts of the KIAA0513 gene showed tissue- and isoform-specific monoallelic expression. The CDH13, ATP2C2, and TLDC1 genes exhibited tissue-specific imprinting, however, COTL1, CRISLPLD2, and GSE1 escaped imprinting. Four DMRs, established after fertilization, were found in this region. Two DMRs were located between the ZDHHC7 and KIAA0513 genes, and two were in exon 1 of the CDH13 and ATP2C2 genes, respectively. The results from this study support future studies on the molecular mechanism to regulate the imprinting of candidate genes on bovine chromosome 18.

角蛋白19(Keratin19,Krt19)是Ⅰ型角蛋白家族中的一员,为研究Krt19在小鼠早期妊娠过程中的表达及调控,本研究利用原位杂交、免疫组织化学、Western blot、real-time PCR等技术,结合小鼠早期妊娠模型、激素处理模型、延迟着床与激活模型和人工诱导蜕膜化模型,研究了 Krt19 mRNA以及蛋白在小鼠子宫中的表达情况.结果显示,Krt19 mRNA和蛋白在小鼠早期妊娠1～4 d的腔上皮和腺上皮中强表达,在5～8 d子宫腔上皮、腺上皮及蜕膜中表达;在延迟着床时,Krt19在子宫腔上皮和腺上皮强表达,注射雌激素激活后,Krt19在腔上皮和腺上皮的表达减弱,在激活的胚胎着床位点下基质细胞中有弱表达;雌激素上调Krt19在子宫中的表达;Krt19在人工诱导蜕膜化的子宫基质细胞中表达.结果表明,Krt19参与了胚胎着床过程,并受类固醇激素的调控,与蜕膜化过程密切相关.

为鉴定牛DSCAM基因的印记状态以及DNA甲基化修饰在调控印记表达中的作用,本研究以牛胎盘和成年牛组织(心、肝、脾、肺、肾、肌肉、脂肪和大脑)为试验材料,利用基于单核苷酸多态(SNP)的RT-PCR直接测序法分析DSCAM基因的印记状态,采用亚硫酸氢盐测序法分析基因启动子区的甲基化状态.结果发现,在DSCAM基因的第 32 个外显子上存在 1 个A/G杂合的SNP位点(rs136908595),利用该SNP位点区分亲本等位基因发现,在被检测的成年牛组织中,DSCAM基因为双等位基因表达;而在胎盘中DSCAM基因为母源等位基因表达.对牛DSCAM基因启动子区及第一个外显子处 402 bp的CpG岛甲基化进行分析,发现在单等位基因表达的胎盘中,2 条亲本链间存在差异甲基化区,而在双等位基因表达的组织中,未发现差异甲基化区.结果表明,DNA甲基化修饰参与调控牛DSCAM基因的胎盘特异性父源印记,可为深入探讨牛DSCAM基因功能和调控机制提供参考依据.

Correct reprogramming of the DLK1-DIO3 imprinted region is critical for the development of cloned animals. However, in pigs, the imprinting and regulation of the DLK1-DIO3 region has not been systematically analyzed. The objective of this study was to investigate the imprinting status and methylation regulation of the DLK1-DIO3 region in wild-type and cloned neonatal pigs. We mapped the imprinting control region, IG-DMR, by homologous alignment and validated it in sperm, oocytes, fibroblasts, and parthenogenetic embryos. Subsequently, single nucleotide polymorphism-based sequencing and bisulfite sequencing polymerase chain reaction were conducted to analyze imprinting and methylation in different types of fibroblasts, as well as wild-type and cloned neonatal pigs. The results showed that Somatic cell nuclear transfer (SCNT) resulted in hypermethylation of the IG-DMR and aberrant gene expression in the DLK1-DIO3 region. Similar to wild-type pigs, imprinted expression and methylation were observed in the surviving cloned pigs, whereas in dead cloned pigs, the IG-DMR was hypermethylated and the expression of GTL2 was nearly undetectable. Our study reveals that abnormal imprinting of the DLK1-DIO3 region occurs in cloned pigs, which provides a theoretical basis for improving the cloning efficiency by gene editing to correct abnormal imprinting.

Sno-lncRNAs are intron-derived long noncoding RNAs (lncRNAs) with snoRNA ends. Sno-lncRNAs were first discovered in the human Prader-Willi (PWS)/Angelman (AS) imprinted domain. Here, we report the identification and characterization of four sno-lncRNA types (sno-lncRNA1, sno-lncRNA2, sno-lncRNA3, and sno-lncRNA4) in the bovine PWS/AS imprinted domain. Reverse transcription-PCR first determined the cDNA sequences of the four bovine sno-lncRNAs. A gene structure analysis showed that sno-lncRNA1 lacks introns, but sno-lncRNA2 and sno-lncRNA3 have one and two introns respectively. The three sno-lncRNAs have similar snoRNA ends. Moreover, the three have similar snoRNAs at their 5' and 3' ends. The head-to-tail orientation has six sno-lncRNA copies arranged between bovine SNORD116-6 and SNORD116-12. Moreover, only a copy of sno-lncRNA4 was located between SNORD116-3 and SNORD116-4. The expression of the four sno-lncRNAs was analyzed in the bovine heart, liver, spleen, lung, kidney, muscle, fat, brain, and placenta tissues. The monoallelic expression of sno-lncRNA4 was determined in bovine tissues. The results showed that the four sno-lncRNAs are widely expressed in the nine tissues, although sno-lncRNA3 and sno-lncRNA4 were undetected in the placenta. Moreover, an informative single nucleotide polymorphism (rs448706424) revealed the allelic expression of sno-lncRNA4 in exon 2 of sno-lncRNA4. The bovine genome had six copies of sno-lncRNA1, sno-lncRNA2, and sno-lncRNA3, but their allelic expression was not identified.

Genomic imprinting is an epigenetic phenomenon that leads to genes monoallelically expressed in a parent-of-origin-specific manner and plays an important role in the embryonic development and postnatal growth of mammals. Imprinted genes usually occur in clusters in a chromosomal region and are regulated by a cis-acting imprinting control region that involves differential DNA methylation modification. Igf2r, Slc22a2 and Slc22a3 are three maternally expressed genes on mouse chromosome 17. The paternally expressed long noncoding RNA (lncRNA) Air and the nonimprinted gene Slc22a1 are also located in the imprinted region. Comparative characterization of imprinted clusters between species is useful for us to understand the biological significance and epigenetic regulating mechanism of genomic imprinting. The aim of this study was to analyze the allelic expression pattern of AIR and SLC22A1-3 genes in cattle and to determine the role of DNA methylation in regulating gene expression. Allelic expression analysis was performed in bovine adult tissues and term placenta using an SNP-based approach. We found that IGF2R, AIR and SLC22A3 were monoallelically expressed in all detected bovine somatic tissues, including heart, liver, spleen, lung, kidney, muscle, fat and brain. In bovine placenta, IGF2R and SLC22A3 are maternally expressed; however, the AIR gene is paternally expressed. Tissue-specific monoallelic expression of SLC22A2 is detected in bovines, with monoallelic expression in the spleen and brain but biallelic expression in kidney tissues. SLC22A1 is only detected in bovine liver and kidney tissues and is biallelicly expressed, which is consistent with the imprint expression in mice. To determine the possible role of DNA methylation in regulating the monoallelic/imprinted expression of bovine IGF2R, AIR, SLC22A2, and SLC22A3 genes, we analyzed the DNA methylation status of CpG islands in the first exon of SLC22A2, the promoter region of SLC22A3 and region 2 in the second intron of the IGF2R gene by bisulfite sequencing. Two differentially methylated regions (DMRs) were detected in the first exon of bovine SLC22A3 and the common regions of IGF2R and AIR. This suggests that DNA methylation is involved in the regulation of monoallelic/imprinted expression of IGF2R, AIR and SLC22A3 genes in cattle.

In mammals, imprinted genes are required for both fetal development and postnatal growth. A novel candidate imprinted locus was found on human chromosome 16, and maternal uniparental disomy of this locus can cause a lethal developmental lung disease in human newborns. The PMM2 and NARFL genes are located in this region and its homologous region in cattle is on chromosome 25. Currently, there is no report on the genomic imprinting of the PMM2 and NARFL genes. In this study, we demonstrated that PMM2 and NARFL are two paternally imprinted genes in bovines using an SNP-based method. In addition, two differentially methylated regions of paternal methylation were found in the promoter region and the third intron of the bovine NARFL gene, which may be involved in regulating its imprinted expression. However, we did not find differential methylation in the promoter region or the seventh intron of the bovine PMM2 gene.

泛素样PHD和含环指结构域的蛋白1(UHRF1)广泛表达在成人和胎儿的胸腺、胎儿的肝脏及骨髓中,其在胚胎发育、调节细胞周期、调控细胞的生长以及维持基因组的稳定性中发挥着重要的作用.本研究利用组织切片、原位杂交、免疫组织化学、Western blot、Real-time PCR等试验技术分析UHRF1的mRNA和蛋白的表达情况,来探究其在着床及蜕膜化过程中的表达规律.结果表明,UHRF1参与胚胎着床过程,其表达受到类固醇激素的调控,与蜕膜化过程密切相关.

The epigenetic process of genomic imprinting results in the monoallelic expression of genes based on their parental origin. Comparative analysis of imprinted genes between species is useful for investigating the biological significance and regulatory mechanisms of genomic imprinting. Mouse Impact is an imprinted gene, but its human ortholog IMPACT escapes genomic imprinting. Hrh4 and Osbpl1a are the two nearest neighbors of the Impact located in distal and proximal regions, respectively. This study aims to assess the allelic expression of bovine IMPACT, OSBPL1A and HRH4 genes and examine the differentially methylated regions (DMRs) associated with these three genes. Based on an expressed single-nucleotide polymorphism (SNP) approach, we found that both the IMPACT and OSBPL1A genes exhibit isoform-specific monoallelic expression in bovine adult tissues. In the seven detected bovine IMPACT transcripts, only one transcript variant (X6) is monoallelically expressed in bovine adult tissues and paternally expressed in the placenta. However, no DMR was found in the promoter region of the IMPACT gene. We obtained five transcript variants (V1-V5) of the bovine OSBPL1A gene of different lengths that start transcription from distinct alternative promoters by RT-PCR. Only the longest variant V1 was found to be expressed monoallelically in bovine adult tissues and a DMR was identified in its promoter region using the bisulfite sequencing method. Thus, the DMR in OSBPL1A V1 promoter region may contribute to its isoform-specific monoallelic expression. The bovine HRH4 gene is expressed biallelically. Hypermethylation was observed in brains without HRH4 expression, while hypomethylation was found in the spleens with HRH4 expression, so and the level of DNA methylation in the promoter seemed to be related to its expression in tissues.

Polyaniline (PANI) was covalently linked with hexagonal boron nitride (BN) using p-aminobenzoic acid (ABA) to obtain a PANI-BN powder. The PANI-BN/EP coating was applied on hot-dip galvanized steel, which exhibited remarkable corrosion resistance. Electrochemical test results showed that the impedance modulus in the low frequency region of PANI-BN/EP was maximum after soaking in NaCl solution (3.5 wt.%) for 72 h compared with other coatings. The outstanding corrosion resistance of the PANI-BN/EP coating was primarily attributed to the synergistic effect of BN and PANI.

长非编码RNA(lncRNA)作为一种重要的调节因子,参与调控细胞的各种正常生理过程,并在肿瘤的发生发展中发挥作用.母源等位基因表达的MEG3基因(maternally expressed gene 3),位于人(Homo sapiens)14号染色体q32.2的DLK1-DIO3印记区域内,34.9 kb的基因编码长度大约为1.6 kb的lncRNA.MEG3基因在多种癌变的组织和细胞中表达缺失或下调,表现出抑癌功能.MEG3通过调控肿瘤抑制基因和癌基因,如与p53、RB、MYC和TGF-β相互作用,从而调控肿瘤的发生.MEG3基因还通过调控Wnt-β-catenin信号通路,影响上皮-间质的转化.此外,位于MEG3基因位点的甲基化修饰和多态位点也与患癌的风险以及药物的治疗效果有关.本文针对MEG3基因在肿瘤发生中的作用机制,以及MEG3基因在作为癌症诊断和预后的生物标志物,以及作为治疗的靶标中的价值进行了论述.本文为后续研究MEG3在人类肿瘤治疗中的作用提供基础.

为揭示牛AQP1（aquaporin 1）基因在不同组织及胎盘中的印记状态，以及DNA甲基化修饰在印记中的调控机制，本研究采用基于SNP的PCR产物直接测序的方法，对32头健康雌性成年荷斯坦奶牛心组织及15个自然分娩后的胎盘试验样本进行检测，确定了5头杂合子个体牛和3个杂合子胎盘，对其组织（心、肝、脾、肺、肾、肌肉和脂肪）和胎盘进行AQP1等位基因表达分析及印记状态分析，利用亚硫酸氢盐测序法分析AQP1基因位于启动子和第一个外显子区的CpG岛在牛心、肝组织、2个胎盘和对应精子中的DNA甲基化状态。结果发现，在杂合子牛被检测的7个组织中，AQP1基因呈现双等位基因表达；而在胎盘中，AQP1基因为单等位基因表达。通过分析杂合子胎盘对应的亲本基因型，发现AQP1基因为母源等位基因表达，即父源印记。进一步比较分析AQP1基因启动子区CpG岛在牛组织、胎盘及对应精子中的甲基化状态，在双等位基因表达的心脏、肝脏组织中，该区域未发现差异甲基化区（differentially methylated regions，DMR）；而在单等位基因表达的胎盘中，存在差异甲基化区，同时父源等位基因精子中为重甲基化状态。以上结果说明，牛AQP1基因为胎盘特异性单等位基因表达的父源印记基因，且AQP1基因位于启动子和第一个外显子区的CpG岛甲基化修饰参与调控牛胎盘的印记表达；在被检测的组织中为双等位基因表达。

弹性蛋白(Elastin)是一种关键的细胞外基质蛋白,对大动脉、肺、韧带、肌腱、皮肤和弹性软骨等许多脊椎动物组织的弹性和复原力至关重要.本实验旨在利用原位杂交、荧光定量PCR方法,研究Elastin mRNA在小鼠早期妊娠、假孕及人工诱导蜕膜化模型子宫中的表达.结果 显示:在小鼠早期妊娠1～4 d子宫中未检测到Elastin mRNA表达;随着妊娠进行,在妊娠第5天子宫肌层检测到Elastin mRNA微弱表达;在妊娠第6天的子宫壁肌层与第7、8天的肌层及蜕膜区Elastin mRNA表达逐渐增强.在小鼠假孕1～5 d子宫中,Elastin mRNA不表达.在人工诱导蜕膜化模型中,Elastin mRNA在子宫壁肌层及系膜侧蜕膜区均有表达.以上表明Elastin可能参与小鼠早期妊娠子宫壁肌层弹性的调控与蜕膜化过程.

基因组印记(genomic imprinting)作为表观遗传现象,是指依赖亲本来源特异性的单等位基因表达.印记基因在哺乳动物胚胎和胎盘发育以及个体生长中具有重要作用.ZC3H12C(zinc finger CCCH-type containing 12C)基因编码具有CCCH-type锌指结构的转录因子,参与调控人类(Homo sapiens)多种免疫性疾病.ZC3H12C首先在人类胎盘中被发现为印记基因,在牛(Bos taurus)中是否发生印记尚不明确.本研究首先分析牛ZC3H12C基因结构,进而基于SNP比较基因组DNA和cDNA扩增产物的方法分析牛ZC3H12C基因在组织和胎盘中的等位基因表达状态,发现ZC3H12C基因在心脏、肝脏、脾脏、肺脏、肾脏、大脑等6个组织均表现为单等位基因表达,在胎盘中表现为双等位基因表达.进一步利用亚硫酸盐测序法分析ZC3H12C基因的甲基化状态,发现在单等位基因表达的肝脏和肾脏中,X2剪接体的第1个内含子上含有一段差异甲基化区,而在双等位基因表达的胎盘中,该区域呈现轻甲基化.上述结果提示,DNA甲基化可能参与调控牛ZC3H12C基因的单等位基因表达.本研究结果可为深入研究牛ZC3H12C基因的功能提供参考依据.

Genomic imprinting is the epigenetic mechanism of transcriptional regulation that involves differential DNA methylation modification. Comparative analysis of imprinted genes between species can help us to investigate the biological significance and regulatory mechanisms of genomic imprinting. MKRN3, MAGEL2 and NDN are three maternally imprinted genes identified in the human PWS/AS imprinted locus. This study aimed to assess the allelic expression of MKRN3, MAGEL2 and NDN and to examine the differentially methylated regions (DMRs) of bovine PWS/AS imprinted domains. An expressed single-nucleotide polymorphism (SNP)-based approach was used to investigate the allelic expression of MKRN3, MAGEL2 and NDN genes in bovine adult tissues and placenta. Consistent with the expression in humans and mice, we found that the MKRN3, MAGEL2 and NDN genes exhibit monoallelic expression in bovine somatic tissues and the paternal allele expressed in the bovine placenta. Three DMRs, PWS-IC, MKRN3 and NDN DMR, were identified in the bovine PWS/AS imprinted region by analysis of the DNA methylation status in bovine tissues using the bisulfite sequencing method and were located in the promoter and exon 1 of the SNRPN gene, NDN promoter and 5’ untranslated region (5’UTR) of MKRN3 gene, respectively. The PWS-IC DMR is a primary DMR inherited from the male or female gamete, but NDN and MKRN3 DMR are secondary DMRs that occurred after fertilization by examining the methylation status in gametes.