羊绒细度候选基因靶标miRNAs筛选和鉴定
|
摘要
为筛选和验证辽宁绒山羊粗型和细型个体羊绒细度相关候选基因ALX4、NT-3、POLD2和AKT1表达调控潜在靶标miRNAs及其表达差异,提取辽宁绒山羊细型和粗型皮肤组织DNA和RNA,先采集辽宁绒山羊粗型和细型肩胛部皮肤组织,根据NCBI上的序列克隆获得ALX4、NT-3、POLD2和AKT1基因的3'UTR,利用Primer 5.0设计并合成功能引物,通过常规PCR扩增ALX4、NT-3、POLD2和AKT1基因的3'UTR序列,在miRBase数据库中,筛选羊绒细度候选靶基因3'UTR靶标miRNAs,然后与辽宁绒山羊(LCG)和内蒙古绒山羊(MCG)品种间皮肤miRNAs高通量测序结果共聚焦,筛选和验证羊绒细度相关候选基因表达调控潜在靶标miRNAs,在NCBI中检索到miRNAs的前体序列,应用RNAfold获得二级结构图及势能图,通过qPCR验证细型辽宁绒山羊(Fine type LCG)和粗型辽宁绒山羊(Coarse type LCG)皮肤组织中羊绒细度候选基因ALX4、NT-3、POLD2和AKT1靶标miRNAs的表达量。结果表明:通过miRBase数据库预测,羊绒细度候选基因ALX4、NT-3、POLD2和AKT1分别获得104,27,19和32个靶标miRNAs,与绒山羊品种间皮肤miRNAs高通量测序结果聚焦分析,ALX4基因聚焦到3个靶标miR-m13,miR-433-5p和miR-671,其表达量是56,2和27;NT-3基因聚焦到1个靶标miR-1296,表达量是41;POLD2基因聚焦到1个靶标miR-130b,表达量是177;AKT1基因聚焦到2个靶标miR-345-3p和miR-2331-3p,其表达量是52和5。其中前体序列的二级结构表明miR-2331-3p的结构最不稳定,miR-1296最小自由能的绝对值最大,结构最为稳定。用qPCR验证miR-m13,miR-433-5p,miR-671,miR-1296,miR-345-3p和miR-2331-3p在辽宁绒山羊细型皮肤中的表达量都显著高于粗型皮肤中的表达量,可能对羊绒细度候选基因发挥重要的正调控作用,而miR-130b在LCG粗型的表达量高于LCG细型表达量,可能miR-130b对羊绒细度相关候选基因起负调控作用。
The purpose of this study was to screen and verify the expression and regulation of the potential target miRNAs and its expression difference between coarse and fine individual cashmere fineness related candidate genes ALX4, NT-3, POLD2 and AKT1 in Liaoning Cashmere Goat. DNA and RNA were extracted from fine and coarse skin tissue of Liaoning Cashmere Goat.The coarse and fine scapular skin tissues of Liaoning Cashmere Goat were collected firstly. According to the sequence of NCBI,the 3'UTRs of ALX4, NT-3, POLD2 and AKT1 genes were obtained, and functional primers were designed and synthesized byPrimer 5.0. 3'UTR sequences of ALX4, NT-3, POLD2 and AKT1 genes were amplified by conventional PCR. In miRBase database, the target miRNAs was screened by 3'UTR of cashmere fineness candidate gene and then confocal with high-throughput miRNAs sequencing results of Liaoning Cashmere Goat(LCG) and Inner Mongolia Cashmere Goat(MCG). Screening and Verification of Cashmere fineness related candidate Gene expression Regulation potential Target miRNAs, the precursor sequence of miRNAs was found in NCBI. The secondary structure diagram and potential energy map were obtained by using RNAfold. The expression of candidate gene ALX4, NT-3, POLD2 and AKT1 target miRNAs in fine Liaoning cashmere goat(Fine type LCG) and coarse Liaoning cashmere goat(Coarse type LCG) skin was detected by qPCR. The results showed that the candidate gene of cashmere fineness, ALX4, NT-3, POLD2 and AKT1, were predicted by miRBase to obtain 104, 27, 19 and 32 target miRNAs. Focusing analysis of high-throughput sequencing results of skin miRNAs between cashmere goat breeds, ALX4 gene focused on three targets miR-m13, miR-433-5 p and miR-671, the expression levels were 56, 2 and 27; The miR-1296,expression of NT-3 gene focused on one target was 41. The expression of POLD2 gene focused on one target miR-130 b, was177. The AKT1 gene was focused on two targets, miR-345-3 p and miR-2331-3 p, and the expression levels were 52 and 5. The centroid secondary structure of the precursor sequence indicates that the structure of miR-2331-3 p is the most unstable, and the absolute value of the minimum free energy of miR-1296 is the largest, and the structure is the most stable. The expression of miR-m13, miR-433-5 p, miR-671, miR-1296, miR-345-3 p and miR-2331-3 p in fine skin of Liaoning cashmere goat was significantly higher than that in coarse skin by qPCR. It may play an important role in positive regulation of cashmere fineness candidate genes, while the expression of miR-130 b in coarse type of LCG is higher than that in fine type of LCG. It may be that miR-130 b plays a negative role in controlling candidate genes related to cashmere fineness.
The purpose of this study was to screen and verify the expression and regulation of the potential target miRNAs and its expression difference between coarse and fine individual cashmere fineness related candidate genes ALX4, NT-3, POLD2 and AKT1 in Liaoning Cashmere Goat. DNA and RNA were extracted from fine and coarse skin tissue of Liaoning Cashmere Goat.The coarse and fine scapular skin tissues of Liaoning Cashmere Goat were collected firstly. According to the sequence of NCBI,the 3'UTRs of ALX4, NT-3, POLD2 and AKT1 genes were obtained, and functional primers were designed and synthesized byPrimer 5.0. 3'UTR sequences of ALX4, NT-3, POLD2 and AKT1 genes were amplified by conventional PCR. In miRBase database, the target miRNAs was screened by 3'UTR of cashmere fineness candidate gene and then confocal with high-throughput miRNAs sequencing results of Liaoning Cashmere Goat(LCG) and Inner Mongolia Cashmere Goat(MCG). Screening and Verification of Cashmere fineness related candidate Gene expression Regulation potential Target miRNAs, the precursor sequence of miRNAs was found in NCBI. The secondary structure diagram and potential energy map were obtained by using RNAfold. The expression of candidate gene ALX4, NT-3, POLD2 and AKT1 target miRNAs in fine Liaoning cashmere goat(Fine type LCG) and coarse Liaoning cashmere goat(Coarse type LCG) skin was detected by qPCR. The results showed that the candidate gene of cashmere fineness, ALX4, NT-3, POLD2 and AKT1, were predicted by miRBase to obtain 104, 27, 19 and 32 target miRNAs. Focusing analysis of high-throughput sequencing results of skin miRNAs between cashmere goat breeds, ALX4 gene focused on three targets miR-m13, miR-433-5 p and miR-671, the expression levels were 56, 2 and 27; The miR-1296,expression of NT-3 gene focused on one target was 41. The expression of POLD2 gene focused on one target miR-130 b, was177. The AKT1 gene was focused on two targets, miR-345-3 p and miR-2331-3 p, and the expression levels were 52 and 5. The centroid secondary structure of the precursor sequence indicates that the structure of miR-2331-3 p is the most unstable, and the absolute value of the minimum free energy of miR-1296 is the largest, and the structure is the most stable. The expression of miR-m13, miR-433-5 p, miR-671, miR-1296, miR-345-3 p and miR-2331-3 p in fine skin of Liaoning cashmere goat was significantly higher than that in coarse skin by qPCR. It may play an important role in positive regulation of cashmere fineness candidate genes, while the expression of miR-130 b in coarse type of LCG is higher than that in fine type of LCG. It may be that miR-130 b plays a negative role in controlling candidate genes related to cashmere fineness.
引文
[1]刘海英,杨桂芹,张微,等.FGF5基因对内蒙古绒山羊绒毛性状的影响[J].遗传,2009,31(2):175-179.
[2] JIN M,WANG L,LI S,et al.Characterization and expression analysis of KAP7.1,KAP8.2 gene in Liaoning new-breeding cashmere goat hair follicle[J].Molecular Biology Reports,2011,38(5):3023-3028.
[3] ZHAO M,CHEN H,WANG X,et al.aPCR-SSCP and DNA sequencing detecting two silent SNPs at KAP8.1 gene in the cashmere goat[J].Molecular Biology Reports,2009,36(6):1387-1391.
[4] ZENG X C,CHEN H Y,JIA B,et al.Identification of SNPs within the sheep PROP1 gene and their effects on wool traits[J].Molecular Biology Reports,2011,38(4):2723-2728.
[5] ZHOU J P,ZHU X P,ZHANG W,et al.A novel single-nucleotide polymorphism in the 5’upstream region of the prolactin receptor gene is associated with fiber traits in Liaoning cashmere goats[J].Genetics and Molecular Research,2011,10(4):2511-2516.
[6]刘斌.绒山羊绒毛生长相关基因的筛选、鉴定和多态性分析[D].呼和浩特:内蒙古农业大学,2012.
[7]狄冉.中国产绒山羊微卫星和单核苷酸多态性研究[D].北京:中国农业科学院,2008.
[8]李丽娟,张利平,刘武军.三个绒山羊品种KIFI基因多态性与绒用性状相关性研究[D].兰州:甘肃农业大学,2009.
[9] QIAO X,SU R,WANG Y,et al.Genome-wide target enrichment-aided chip design:a 66K SNP chip for cashmere goat[J].Scientific Reports,2017,7(1):8621.
[10] ZHANG W,GUANG W,JIANG H,et al.A subset of skin-expressed miRNAs with possible roles in goat and sheep hair growth based on expression profiling of mammalian miRNAs[J].A Journal of Integrative Biology,2007,4(11):385-396.
[11] LIU Z,XIAO H,LI H,et al.Identification of conserved and novel microRNAs in cashmere goat skin by deep sequencing[J].Plos One,2012,7(12):e50001.
[12]于永生,曹阳,朴庆林,等.辽宁绒山羊绒直径相关基因的筛选[J].中国草食动物科学,2014(S1):147-149.
[13]张桂山.辽宁绒山羊与乾华肉用美利奴羊皮肤毛囊miRNAs筛选及靶基因验证[D].长春:吉林农业大学,2016.
[14]翟博,张立春,李旭,等.microRNA在反刍动物中的研究进展[J].动物营养学报,2016,28(7):1975-1980.
[15]金雷皓.microRNA-663对羊驼黑色素细胞黑色素合成的影响[D].晋中:山西农业大学,2014.
[16]付绍印.绒山羊绒毛周期性相关miRNAs及其靶基因的研究[D].呼和浩特:内蒙古农业大学,2014.
[17]江倩.miR-let7a在辽宁绒山羊毛囊发育周期中的差异表达及其靶基因功能鉴定[D].长春:吉林大学,2015.
[18] COSKUN E,ERCIN M,GEZGINCI-OKTAYOGLU S.The role of epigenetic regulation and pluripotency-related micrornas in differentiation of pancreatic stem cells to beta cells[J].Journal of Cellular Biochemistry,2017,119(1):455-467.
[19] SUN S,WANG X,XU X,et al.MiR-433-3p suppresses cell growth and enhances chemosensitivity by targeting CREB in human glioma[J].Oncotarget,2017,8(3):5057-5068.
[20] YU Y,WANG Z,SUN D,et al.miR-671 promotes prostate cancer cell proliferation by targeting tumor suppressor SOX6[J].European Journal of Pharmacology,2018,823:65-71.
[21] NUNES S,SILVA I B,AMPUERO M R,et al.Integrated analysis reveals that miR-193b,miR-671,and TREM-1 correlate with a good response to treatment of human localized cutaneous leishmaniasis caused by Leishmania braziliensis[J].Frontiers in Immunology,2018,9:640.
[22]薄占东,赵劲民,杨志,等.神经营养素3基因转染神经干细胞及其表达[J].中国组织工程研究与临床康复,2008,12(12):2374-2378.
[23] ALILA H,COLEMAN M,NITTA H,et al.Expression of biologically active hu-man insulin like growth factor I following intra muscular injection of aformulated plasmid in rats[J].Hum Gene Ther,1997,8:1785-1795.
[24]郭春飞,金永德,金玉莲.神经营养素3及其受体在大鼠耳蜗螺旋神经节发育中的作用研究进展[J].临床耳鼻咽喉头颈外科杂志,2017,31(12):968-971.
[25] PHAN B,MAJID S,URSU S,et al.Tumor suppressor role of miRNAs-1296 in triple-negative breast cancer[J].Oncotarget,2016,7(15):19519-19530.
[26] BOO L,HO W Y,ALI N M,et al.MiRNAs Transcriptome profiling of spheroid-enriched cells with cancer stem cell properties in human breast MCF-7 cell line[J].Int J Biol Sci,2016,12(4):427-445.
[27] BOBOWICZ M,SKRZYPSKI M,CZAPIEWSKI P,et al.Prognostic value of 5-miRNAs based signature in T2-T3N0 colon cancer[J].Clinical&Experimental Metastasis,2016,33(8):765-773.
[28] PEREZ A,LEON A,LEE M Y.Characterization of the 5'-flanking region of the gene encoding the 50 kDa subunit of human DNA polymerase delta[J].Biochimica et Biophysica Acta(BBA)-Gene Structure and Expression,2000,1493(1-2):231-236.
[29] MATSUSHIMANISHIU M,UNOKI M,ONO K,et al.Growth and gene expression profile analyses of endometrial cancer cells expressing exogenous PTEN[J].Cancer Research,2001,61(9):3741-3749.
[30] NAORA H.The heterogeneity of epithelial ovarian cancers:reconciling old and new paradigms[J].Expert Rev Mol Med,2007,9:1-12.
[31] LANDEN C N,JR BIRRER M J,SOOD A K.Early events in the pathogenesis of epithelial ovarian cancer[J].J Clin Oncol,2008,26:995-1005.
[32] SHIH I,KURMAN R J.Ovarian tumorigenesis:a proposed model based on morphological and molecular genetic analysis[J].Am J Pathol,2004,164:1511-1518.
[33] BELL DA.Origins and molecular pathology of ovarian cancer[J].Mod Pathol,2005,18(Suppl 2):S19-S32.
[34] BREDEL M,SCHOLTENS D M,HARSH G R,et al.A network model of a cooperative genetic landscape in brain tumors[J].Jama the Journal of the American Medical Association,2009,302(3):261-275.
[35] ZHANG J,XIE S,CHENG J,et al.The second subunit of DNA-polymerase delta is required for genomic stability and epigenetic regulation[J].Plant Physiology,2016,171(2):1192-1208.
[36] YIP L,KELLY L,SHUAI Y,et al.MiRNAs signature distinguishes the degree of aggressiveness of papillary thyroid carcinoma[J].Annals of Surgical Oncology,2011,18(7):2035-2041.
[37] DONG P,KARAAYVAZ M,JIA N,et al.Mutant p53 gain-of-function induces epithelial–mesenchymal transition through modulation of the miR-130b–ZEB1 axis[J].Oncogene,2013,32(27):3286-3295.
[38] LEONE V,LANGELLA C,D'ANGELO D,et al.MiR-23b and miR-130b expression is downregulated in pituitary adenomas[J].Mol Cell Endocrinol,2014,390:1-7.
[39] SAND M,SKRYGAN M,SAND D,et al.Comparative microarray analysis of miRNAs expression profiles in primary cutaneous malignant melanoma,cutaneous malignant melanoma metastases,and benign melanocytic nevi[J].Cell&Tissue Research,2013,351(1):85-98.
[40] SCHEFFER A R,HOLDENRIEDER S,KRISTIANSEN G,et al.Circulating miRNAs in serum:novel biomarkers for patients with bladder cancer?[J].World Journal of Urology,2014,32(2):353-358.
[41] WU X,WENG L,LI X,et al.Identification of a 4-miRNAs signature for clear cell renal cell carcinoma metastasis and prognosis[J].Plos One,2012,7(5):e35661.
[42] ZHI C,LUO J,MA L A,et al.MiR130b-regulation of PPARγcoactivator-1αsuppresses fat metabolism in goat mammary epithelial cells[J].Plos One,2015,10(11):e0142809.
[43] SCHULTZE SM,HEMMINGS BA,NIESSEN M,et al.PI3K/AKT,MAPK and AMPK signalling:protein kinases in glucose homeostasis[J].Expert Reviews in Molecular Medicine,2012,14:e1.
[44] WANG S Y,SHIBOSKI S,BELAIR C D,et al.miR-19,miR-345,miR-519c-5p serum levels predict adverse pathology in prostate cancer patients eligible for active surveillance.[J].Plos One,2014,9(6):e98597.
[45] SIENGDEE P,TRAKOOLJUL N,MURANI E,et al.MiRNAs regulate cellular ATP levels by targeting mitochondrial energy metabolism genes during C2C12 myoblast differentiation[J].Plos One,2015,10(5):e0127850.
[46] CHANG S J,WENG S L,HSIEH J Y,et al.MicroRNA-34a modulates genes involved in cellular motility and oxidative phosphorylation in neural precursors derived from human umbilical cord mesenchymal stem cells[J].Bmc Medical Genomics,2011,4(1):1-8.
[47]李荣荣.miR-345-3p在周期性牵张力诱导大鼠骨髓间充质干细胞成骨分化中的生物学功能[D].济南:山东大学,2016.
[2] JIN M,WANG L,LI S,et al.Characterization and expression analysis of KAP7.1,KAP8.2 gene in Liaoning new-breeding cashmere goat hair follicle[J].Molecular Biology Reports,2011,38(5):3023-3028.
[3] ZHAO M,CHEN H,WANG X,et al.aPCR-SSCP and DNA sequencing detecting two silent SNPs at KAP8.1 gene in the cashmere goat[J].Molecular Biology Reports,2009,36(6):1387-1391.
[4] ZENG X C,CHEN H Y,JIA B,et al.Identification of SNPs within the sheep PROP1 gene and their effects on wool traits[J].Molecular Biology Reports,2011,38(4):2723-2728.
[5] ZHOU J P,ZHU X P,ZHANG W,et al.A novel single-nucleotide polymorphism in the 5’upstream region of the prolactin receptor gene is associated with fiber traits in Liaoning cashmere goats[J].Genetics and Molecular Research,2011,10(4):2511-2516.
[6]刘斌.绒山羊绒毛生长相关基因的筛选、鉴定和多态性分析[D].呼和浩特:内蒙古农业大学,2012.
[7]狄冉.中国产绒山羊微卫星和单核苷酸多态性研究[D].北京:中国农业科学院,2008.
[8]李丽娟,张利平,刘武军.三个绒山羊品种KIFI基因多态性与绒用性状相关性研究[D].兰州:甘肃农业大学,2009.
[9] QIAO X,SU R,WANG Y,et al.Genome-wide target enrichment-aided chip design:a 66K SNP chip for cashmere goat[J].Scientific Reports,2017,7(1):8621.
[10] ZHANG W,GUANG W,JIANG H,et al.A subset of skin-expressed miRNAs with possible roles in goat and sheep hair growth based on expression profiling of mammalian miRNAs[J].A Journal of Integrative Biology,2007,4(11):385-396.
[11] LIU Z,XIAO H,LI H,et al.Identification of conserved and novel microRNAs in cashmere goat skin by deep sequencing[J].Plos One,2012,7(12):e50001.
[12]于永生,曹阳,朴庆林,等.辽宁绒山羊绒直径相关基因的筛选[J].中国草食动物科学,2014(S1):147-149.
[13]张桂山.辽宁绒山羊与乾华肉用美利奴羊皮肤毛囊miRNAs筛选及靶基因验证[D].长春:吉林农业大学,2016.
[14]翟博,张立春,李旭,等.microRNA在反刍动物中的研究进展[J].动物营养学报,2016,28(7):1975-1980.
[15]金雷皓.microRNA-663对羊驼黑色素细胞黑色素合成的影响[D].晋中:山西农业大学,2014.
[16]付绍印.绒山羊绒毛周期性相关miRNAs及其靶基因的研究[D].呼和浩特:内蒙古农业大学,2014.
[17]江倩.miR-let7a在辽宁绒山羊毛囊发育周期中的差异表达及其靶基因功能鉴定[D].长春:吉林大学,2015.
[18] COSKUN E,ERCIN M,GEZGINCI-OKTAYOGLU S.The role of epigenetic regulation and pluripotency-related micrornas in differentiation of pancreatic stem cells to beta cells[J].Journal of Cellular Biochemistry,2017,119(1):455-467.
[19] SUN S,WANG X,XU X,et al.MiR-433-3p suppresses cell growth and enhances chemosensitivity by targeting CREB in human glioma[J].Oncotarget,2017,8(3):5057-5068.
[20] YU Y,WANG Z,SUN D,et al.miR-671 promotes prostate cancer cell proliferation by targeting tumor suppressor SOX6[J].European Journal of Pharmacology,2018,823:65-71.
[21] NUNES S,SILVA I B,AMPUERO M R,et al.Integrated analysis reveals that miR-193b,miR-671,and TREM-1 correlate with a good response to treatment of human localized cutaneous leishmaniasis caused by Leishmania braziliensis[J].Frontiers in Immunology,2018,9:640.
[22]薄占东,赵劲民,杨志,等.神经营养素3基因转染神经干细胞及其表达[J].中国组织工程研究与临床康复,2008,12(12):2374-2378.
[23] ALILA H,COLEMAN M,NITTA H,et al.Expression of biologically active hu-man insulin like growth factor I following intra muscular injection of aformulated plasmid in rats[J].Hum Gene Ther,1997,8:1785-1795.
[24]郭春飞,金永德,金玉莲.神经营养素3及其受体在大鼠耳蜗螺旋神经节发育中的作用研究进展[J].临床耳鼻咽喉头颈外科杂志,2017,31(12):968-971.
[25] PHAN B,MAJID S,URSU S,et al.Tumor suppressor role of miRNAs-1296 in triple-negative breast cancer[J].Oncotarget,2016,7(15):19519-19530.
[26] BOO L,HO W Y,ALI N M,et al.MiRNAs Transcriptome profiling of spheroid-enriched cells with cancer stem cell properties in human breast MCF-7 cell line[J].Int J Biol Sci,2016,12(4):427-445.
[27] BOBOWICZ M,SKRZYPSKI M,CZAPIEWSKI P,et al.Prognostic value of 5-miRNAs based signature in T2-T3N0 colon cancer[J].Clinical&Experimental Metastasis,2016,33(8):765-773.
[28] PEREZ A,LEON A,LEE M Y.Characterization of the 5'-flanking region of the gene encoding the 50 kDa subunit of human DNA polymerase delta[J].Biochimica et Biophysica Acta(BBA)-Gene Structure and Expression,2000,1493(1-2):231-236.
[29] MATSUSHIMANISHIU M,UNOKI M,ONO K,et al.Growth and gene expression profile analyses of endometrial cancer cells expressing exogenous PTEN[J].Cancer Research,2001,61(9):3741-3749.
[30] NAORA H.The heterogeneity of epithelial ovarian cancers:reconciling old and new paradigms[J].Expert Rev Mol Med,2007,9:1-12.
[31] LANDEN C N,JR BIRRER M J,SOOD A K.Early events in the pathogenesis of epithelial ovarian cancer[J].J Clin Oncol,2008,26:995-1005.
[32] SHIH I,KURMAN R J.Ovarian tumorigenesis:a proposed model based on morphological and molecular genetic analysis[J].Am J Pathol,2004,164:1511-1518.
[33] BELL DA.Origins and molecular pathology of ovarian cancer[J].Mod Pathol,2005,18(Suppl 2):S19-S32.
[34] BREDEL M,SCHOLTENS D M,HARSH G R,et al.A network model of a cooperative genetic landscape in brain tumors[J].Jama the Journal of the American Medical Association,2009,302(3):261-275.
[35] ZHANG J,XIE S,CHENG J,et al.The second subunit of DNA-polymerase delta is required for genomic stability and epigenetic regulation[J].Plant Physiology,2016,171(2):1192-1208.
[36] YIP L,KELLY L,SHUAI Y,et al.MiRNAs signature distinguishes the degree of aggressiveness of papillary thyroid carcinoma[J].Annals of Surgical Oncology,2011,18(7):2035-2041.
[37] DONG P,KARAAYVAZ M,JIA N,et al.Mutant p53 gain-of-function induces epithelial–mesenchymal transition through modulation of the miR-130b–ZEB1 axis[J].Oncogene,2013,32(27):3286-3295.
[38] LEONE V,LANGELLA C,D'ANGELO D,et al.MiR-23b and miR-130b expression is downregulated in pituitary adenomas[J].Mol Cell Endocrinol,2014,390:1-7.
[39] SAND M,SKRYGAN M,SAND D,et al.Comparative microarray analysis of miRNAs expression profiles in primary cutaneous malignant melanoma,cutaneous malignant melanoma metastases,and benign melanocytic nevi[J].Cell&Tissue Research,2013,351(1):85-98.
[40] SCHEFFER A R,HOLDENRIEDER S,KRISTIANSEN G,et al.Circulating miRNAs in serum:novel biomarkers for patients with bladder cancer?[J].World Journal of Urology,2014,32(2):353-358.
[41] WU X,WENG L,LI X,et al.Identification of a 4-miRNAs signature for clear cell renal cell carcinoma metastasis and prognosis[J].Plos One,2012,7(5):e35661.
[42] ZHI C,LUO J,MA L A,et al.MiR130b-regulation of PPARγcoactivator-1αsuppresses fat metabolism in goat mammary epithelial cells[J].Plos One,2015,10(11):e0142809.
[43] SCHULTZE SM,HEMMINGS BA,NIESSEN M,et al.PI3K/AKT,MAPK and AMPK signalling:protein kinases in glucose homeostasis[J].Expert Reviews in Molecular Medicine,2012,14:e1.
[44] WANG S Y,SHIBOSKI S,BELAIR C D,et al.miR-19,miR-345,miR-519c-5p serum levels predict adverse pathology in prostate cancer patients eligible for active surveillance.[J].Plos One,2014,9(6):e98597.
[45] SIENGDEE P,TRAKOOLJUL N,MURANI E,et al.MiRNAs regulate cellular ATP levels by targeting mitochondrial energy metabolism genes during C2C12 myoblast differentiation[J].Plos One,2015,10(5):e0127850.
[46] CHANG S J,WENG S L,HSIEH J Y,et al.MicroRNA-34a modulates genes involved in cellular motility and oxidative phosphorylation in neural precursors derived from human umbilical cord mesenchymal stem cells[J].Bmc Medical Genomics,2011,4(1):1-8.
[47]李荣荣.miR-345-3p在周期性牵张力诱导大鼠骨髓间充质干细胞成骨分化中的生物学功能[D].济南:山东大学,2016.