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CRABP1过表达载体的构建及其对猪卵泡颗粒细胞凋亡的影响 被引量:5
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作者 吴夏梦 刘进辉 +4 位作者 王英 雷磊 周展波 王水莲 张虹亮 《中国畜牧杂志》 CAS 北大核心 2021年第6期172-177,共6页
本实验通过构建猪视黄酸结合蛋白1(Cellular Retinoic Acid-Binding Protein1,CRABP1)过表达载体并揭示其对猪卵泡颗粒细胞凋亡的效应。从猪卵巢组织中提取RNA,反转录获得cDNA样本,利用PCR克隆CRABP1基因,并使用T4连接酶将其连入pcDNA3.... 本实验通过构建猪视黄酸结合蛋白1(Cellular Retinoic Acid-Binding Protein1,CRABP1)过表达载体并揭示其对猪卵泡颗粒细胞凋亡的效应。从猪卵巢组织中提取RNA,反转录获得cDNA样本,利用PCR克隆CRABP1基因,并使用T4连接酶将其连入pcDNA3.1载体,进一步转化到感受态细胞中,通过PCR、双酶切及测序鉴定后提取去内毒素质粒。随后将构建好的CRABP1过表达质粒转染猪卵泡颗粒细胞,利用荧光定量PCR(qRT-PCR)检测其转染效率及其对凋亡相关基因的影响,再将CRABP1过表达质粒转染至猪颗粒细胞24 h后添加1 nmol/L全反式视黄酸(All-trans Retinoic Acid,ATRA),检测其凋亡相关基因的表达变化。结果显示:CRABP1扩增片段序列与参考序列一致,将CRABP1过表达质粒转染猪颗粒细胞24 h后,其CRABP1基因表达量极显著升高,说明CRABP1过表达载体构建成功;CRABP1过表达载体单独转染颗粒细胞后,其可促进猪颗粒细胞促凋亡基因Bax、Caspase-3和Fas的mRNA表达,并且CRABP1过表达可逆转ATRA对猪颗粒细胞的抑凋亡作用。结果表明猪CRABP1过表达载体构建成功,其可介导ATRA调控猪卵泡颗粒细胞凋亡。 展开更多
关键词 crabp1 载体构建 卵泡颗粒细胞 凋亡
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Chromosome Mapping, Expression and Polymorphism Analysis of CRABP1 Gene in Pigs
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作者 ZHAO Shuan-ping TANG Zhong-lin +3 位作者 ZHOU Rong QU Chang-qing ZHENG Jian-wei LI Kui 《Journal of Integrative Agriculture》 SCIE CAS CSCD 2014年第5期1051-1057,共7页
Cellular retinoic acid-binding protein 1 (CRABP1) is a well-conserved member of cytosolic lipid-binding protein family. It is an important modulator of retinoic acid signaling. Long serial analysis of gene expressi... Cellular retinoic acid-binding protein 1 (CRABP1) is a well-conserved member of cytosolic lipid-binding protein family. It is an important modulator of retinoic acid signaling. Long serial analysis of gene expression (LongSAGE) analysis suggested that CRABP1 gene was differentially expressed during prenatal skeletal muscle development in porcine. Here, we obtained the full-length coding region sequence and genomic sequence of the porcine CRABP1 gene and analyzed its genomic structures. Subsequently, we examined CRABP1 chromosome assignment using INRA-University of Minnesota 7 000 porcine radiation hybrid panel (IMpRH) and explored its tissue distribution in adult Tongcheng pigs and dynamical expression profiles in prenatal skeletal muscle (33, 65 and 90 days post coitus, dpc) from Landrace (lean-type) (described as L33, L65 and L90) and Tongcheng pigs (obese-type) (described as T33, T65 and T90). The CRABPI gene was mapped to chromosome 7ql 1-q23 and closely linked to the microsatellite marker SWR1928. Quantitative real-time PCR showed that CRABP1 mRNA was highly expressed in lung and stomach, moderately expressed in placenta and uterus, and weakly expressed in other tissues. Moreover, CRABP1 gene was down-regulated during prenatal skeletal muscle development in both Landrace and Tongcheng pigs and it was expressed much higher in T33 than L33. Two single-nucleotide polymorphisms (SNPs) were detected by sequencing and mass spectrometry methods, allele frequency analysis indicated that g. 281 (G〉A) and g. 2992 (G〉A)were deviated from Hardy-Weinberg equilibrium in the Landrace and DLY (Duroc×(Landrace×Yorkshire)) pig breeds. 展开更多
关键词 PIG crabp1 chromosome assignment expression profile allele frequency
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