Preparation and Application of cRNA Probe Labeled with Digoxingenin for Gene F64 of Rice Field Eel

doi:10.13560/j.cnki.biotech.bull.1985.2016.03.022

Biotechnology Bulletin ›› 2016, Vol. 32 ›› Issue (3): 137-141.doi: 10.13560/j.cnki.biotech.bull.1985.2016.03.022

• Research report • Previous Articles Next Articles

Preparation and Application of cRNA Probe Labeled with Digoxingenin for Gene F64 of Rice Field Eel

JIANG Jiao-yun¹, TIAN Wen-fei³, FENG Long⁴, QU Xian-cheng²

1. College of Life Sciences, Guangxi Normal University, Guilin 541004；
2. College of Fisheries and Life Sciences, Shanghai Ocean University, Shanghai 201306；
3. Bowen College of Management, Guilin University of Technology, Guilin 541006；
4. Siyang Fisheries Technical Service Center, Siyang 223700

Received:2015-06-10 Online:2016-03-24 Published:2016-03-25

Abstract

Abstract: The specific gene primers were designed based on the sequences of gene F64 of rice field eel, then the templates for synthesizing the cRNA probe were amplified, then recombinant F64/pGM-T plasmids were constructed and linearized by restriction enzymes, further the sense and anti-sense cRNA probes were synthesized by in vitro transcription with RNA polymerase, and finally labeled with Digoxigenin. The expression changes of gene F64 during the gonadal development of rice field eel were detected by in situ hybridization. According to our results, the expression was only detected by the anti-sense F64 cRNA probe, and the earliest signal was observed in stage V of ovarian, but no more in early stage of ovarian formation. Our study indicated that the cRNA probes can be synthesized using in vitro transcription, and the cRNA probes prepared in this study can be used for accurately detecting the spatial and temporal expression of gene F64.

Key words: gene F64 of rice field eel, gonadal development, cRNA probe, in situ hybridization

JIANG Jiao-yun, TIAN Wen-fei, FENG Long, QU Xian-cheng. Preparation and Application of cRNA Probe Labeled with Digoxingenin for Gene F64 of Rice Field Eel[J]. Biotechnology Bulletin, 2016, 32(3): 137-141.

References

[1]Liu CK. Rudimentary hermaphroditism in the symbranchoid eel, Monopterus javanens is[J]. Sinensia, 1944, 15：1-8.
[2]周秋白, 吴华东, 吴红翔, 等. 产卵与黄鳝性转化关联的研究[J]. 经济动物学报, 2004, 8(2)：89-91.
[3]郭威, 胡青, 高宇, 等. 黄鳝不同性腺发育期间血清甲状腺素、性激素和皮质醇水平的差异[J]. 淡水渔业, 2014, 44：13-17.
[4]Simpson ER, Mahendroo MS, Means GD, et al. Aromatase cytochrome P450, the enzyme responsible for estrogen biosynthesis[J]. Endocr Rev, 1994, 15(3)：342-355.
[5]Zhang Y, Zhang WM, Yang HY. Two cytochrome P450 aromatase genes in the hermaphrodite rice field eel Monopterus albus：mRNA expression during ovarian development and sex change[J]. Endocrinology, 2008, 199(2)：317-331.
[6]Yu JH, Tang YK, Li JL. Cloning, structure, and expression pattern of the P450 aromatase gene in rice field eel(Monopterus albus)[J]. Biochem Genet, 2008, 46：267-280.
[7]Liu JF, Guiguen Y, Liu SJ. Aromatase(P450arom)and 1lβ-hydrexylase(P45011β)genes are differentially expressed during the sex change process of the protogynous rice field eel, Monoptems albus[J]. Fish Physiol Biochem, 2009, 35(3)：511-518.
[8]Xiao YM, Chen LL, Liu J, et al. Contrast expression patterns of JNK1 during sex reversal of the rice field eel[J]. Journal of Experimental Zoology, 2010, 314B：242-256.
[9]胡青, 杨娇艳, 高宇, 等. 黄鳝WT1基因序列分析及在性腺发育过程中的表达[J]. 华中农业大学学报, 2014, 33：73-79.
[10] 何智, 李依雪, 张碧鱼, 等. 黄鳝卵巢初次发育成熟及相关基因表达分析[J]. 水产学报, 2014, 37(4)：536-544.
[11]曲宪成, 尚晓莉, 程翠, 等. 黄鳝性腺抑制差减文库的构建和分析[J]. 中国水产科学, 2011, 18(1)：23-28.
[12]蒋骄云, 冯龙, 曲宪成. 黄鳝性逆转相关基因F4的克隆和表达分析[J]. 广西师范大学学报, 2013, 31(4)：121-127.
[13]冯龙, 许世杰, 李园园, 等. 黄鳝性逆转相关基因的表达及定位分析[J]. 广东农业科学, 2014, 5：189-193.
[14]蒋骄云, 冯龙, 许世杰, 等. 黄鳝性逆转相关基因的克隆与表达[J]. 广东农业科学, 2012, 39(2)：106-109.
[15]Matsuda M, Nagahama Y, Shinomiya A, et al. DMY is a Y-specific DM-domain gene required for male development in the medaka fish[J]. Nature, 2002, 417：559-563.
[16]路畅, 苏利娜, 朱邦科. 鱼类性别决定及分化相关基因研究进展[J]. 湖北农业科学, 2014, 53(13)：2981-2985.
[17]武晓敏, 黄静, 魏燕燕, 等. 小鼠Brd2基因探针的制备及其在荧光原位杂交实验中的应用[J]. 神经解剖学杂志, 2014, 30(3)：335-339.
[18]曹玉文, 陆天才, 李锋, 等. 体外转录法制备地高辛标记cx43cRNA探针[J]. 农垦医学, 2004, 26(1)：9-10.
[19]Qu XC, Jiang JY, Shang XL, et al. Construction and analysis of gonad suppression subtractive hybridization libraries for the rice field eel, Monopterus albus[J]. Gene, 2014, 540(1)：20-25.

Preparation and Application of cRNA Probe Labeled with Digoxingenin for Gene F64 of Rice Field Eel

PDF (PC)

Knowledge

Abstract

Cite this article

share this article

References

Related Articles 2

Recommended Articles

Metrics

Comments

[1]	LI Yu, CHEN Gang, MA Qian, KUANG Jie-hua, CHEN You-ming. Cloning of nanos1 Gene of Rachycentron canadum and Its Expression Analysis in the Embryonic and Gonadal Development [J]. Biotechnology Bulletin, 2022, 38(5): 175-182.
[2]	HUANG Tian-qing, WANG Xing-ran ,WANG Yan-na, SUN Hui-zhi, HAN Ying. Expression Analysis of lc3b Gene in Gonadal Development of Triploid Female Rainbow Trout [J]. Biotechnology Bulletin, 2017, 33(7): 179-184.