Cloning and Expression Analysis of MiUGT79B1 and MpUGT79B1 Genes from Two Varieties of Meconopsis

doi:10.7668/hbnxb.20194846

Abstract

Abstract:

To investigate the sequence characteristics of uridine diphosphate glycosyltransferase in different flower color of Meconopsis and its relationship with the flower color of Meconopsis.The yellow-flowered Meconopsis integrifolia and the red-flowered M.punicea were used as study materials,and one UGT gene was screened from the full-length transcriptome data of each of the two flower colors of Meconopsis.Based on the results of KEGG annotations of the transcriptomes,these two genes were named MiUGT79B1 and MpUGT79B1,and they were cloned,and analyzed by bioinformatic and RT-qPCR.The results showed that the ORF length of MiUGT79B1 and MpUGT79B1 genes was 1 314,encoding 437 amino acids.Both MiUGT79B1 and MpUGT79B1 proteins were hydrophilic stabilizing proteins with their structures consisting mainly of α-helices and random coil.Phylogenetic tree and multiple sequence comparison analyses revealed that MiUGT79B1 and MpUGT79B1 were most closely related to Papaver somniferum,P.californicum, P.nudicaule,and Macleaya cordata.RT-qPCR analysis showed that MiUGT79B1 and MpUGT79B1 were highly expressed in the bud stage,MiUGT79B1 expression tended to decrease first and then increase,and MpUGT79B1 expression tended to decrease first and then remain stable.There was a significant positive correlation between the expression level of UGT gene and the accumulation pattern of total flavonoids.The results of this study provide a theoretical basis for further in-depth research on the function of MiUGT79B1 and MpUGT79B1 genes in the petal coloration of Meconopsis and its molecular breeding of flower color.

Key words: Meconopsis integrifolia, Meconopsis punicea, UGT79B, Gene cloning, Flower color

ZHOU Lin, LI Tuojian, QU Yan. Cloning and Expression Analysis of MiUGT79B1 and MpUGT79B1 Genes from Two Varieties of Meconopsis[J]. Acta Agriculturae Boreali-Sinica, 2024, 39(4): 102-109. doi: 10.7668/hbnxb.20194846.

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Figures/Tables 13

Fig.1 M.integrifolia(top)and M.punicea(bottom)in different sampling periods

Tab.1 Primer sequence

引物名称 Primer name	引物序列(5'—3') Primer sequence (5'—3')
MiUGT79B1-F	ATGGGTCATCTAACTGCTTTCC
MitUGT79B1-R	CTATAGCAGATCCTTAAGCTTGGTG
qMiUGT79B1-F	CTTAGCCACCCTTCCATCGG
qMitUGT79B1-R	CCGCTGTTTTCGTCCATCAC
MpUGT79B1-F	ATGGGTCATCTAACTGCTTTCC
MpUGT79B1-R	CTATAGCAGATCCTGAAGCTTGG
qMpUGT79B1-F	CACCCTTCCATCGGGTGTTT
qMpUGT79B1-R	TTCGTTAGCAACTCCGCCAT
Action-F	TTCCGTGTTCCTACCGTTGA
Action-R	TCCCTTCATCTTACCCTCGG

Fig.2 Electrophoresis of total RNA M.Marker;1.Total RNA of M.integrifolia;2.Total RNA of M.punicea.

Fig.3 Electrophoresis of MiUGT79B1 and MpUGT79B1 genes M.Marker;1.MiUGT79B1 gene;2.MpUGT79B1 gene.

Fig.4 Hydrophobicity analysis of MiUGT79B1 and MpUGT79B1 protein A.MiUGT79B1 protein;B.MpUGT79B1 protein.The same as Fig.5—8.

Fig.5 Transmembrane structure prediction of MiUGT79B1 and MpUGT79B1 protein

Fig.6 Prediction of MiUGT79B1 and MpUGT79B1 phosphorylation sites

Fig.7 Prediction of the secondary structure of MiUGT79B1 and MpUGT79B1 Blue line.Alpha helix;Red line.Extended strand;Green line.β-sheet;Purple line.Random coil.

Fig.8 Prediction of the three dimensional structure of MiUGT79B1 and MpUGT79B1

Fig.9 Phylogenetic trees of MiUGT79B1 and MpUGT79B1 Black line.Same Genus;Red line.Same family;Blue line.Same order.

Fig.10 Results of amino acid sequence alignment between MiUGT79B1 and MpUGT79B1

Fig.11 Expression analysis of MiUGT79B1 and MpUGT79B1 genes n=3,the different lowercase letters represent significant differences(P<0.05).The same as Fig.12.

Fig.12 Expression of total flavonoids of two varieties of Meconopsis in different periods

References 23

[1]	陈丽琦, 严朋飞, 贾维嘉, 区智, 屈燕. 威氏绿绒蒿(Meconopsis wilsonii)花色相关基因MwF3H的克隆及表达分析[J]. 基因组学与应用生物学, 2022, 41(4):854—861.doi:10.13417/j.gab.041.000854.
	Chen L Q, Yan P F, Jia W J, Ou Z, Qu Y. Cloning and expression analysis of flower color related gene MwF3H from Meconopsis wilsonii[J]. Genomics and Applied Biology, 2022, 41(4):854—861.
[2]	罗军, 陈丽琦, 李拓键, 区智, 屈燕. 基于转录组分析金属离子对绿绒蒿属3种植物花瓣呈色的影响[J]. 植物资源与环境学报, 2023, 32(5):16—27.doi:10.3969/j.issn.1674-7895.2023.05.02.
	Luo J, Chen L Q, Li T J, Ou Z, Qu Y. Analysis on effects of metal ions on the petal color of three species in Meconopsis based on transcriptome[J]. Journal of Plant Resources and Environment, 2023, 32(5):16—27.
[3]	何静娟, 范燕萍. 观赏植物花色相关的类胡萝卜素组成及代谢调控研究进展[J]. 园艺学报, 2022, 49(5):1162—1172.doi:10.16420/j.issn.0513-353x.2021-0623.
	He J J, Fan Y P. Progress in composition and metabolic regulation of carotenoids related to floral color[J]. Acta Horticulturae Sinica, 2022, 49(5):1162—1172. doi: 10.16420/j.issn.0513-353x.2021-0623
[4]	Zhang A A, Zheng J, Chen X M, Shi X Y, Wang H S, Fu Q S. Comprehensive analysis of transcriptome and metabolome reveals the flavonoid metabolic pathway is associated with fruit peel coloration of melon[J]. Molecules, 2021, 26(9):2830.doi:10.3390/molecules26092830.
[5]	葛诗蓓, 张学宁, 韩文炎, 李青云, 李鑫. 植物类黄酮的生物合成及其抗逆作用机制研究进展[J]. 园艺学报, 2023, 50(1):209—224.doi:10.16420/j.issn.0513-353x.2021-1186.
	Ge S B, Zhang X N, Han W Y, Li Q Y, Li X. Research progress on plant flavonoids biosynthesis and their anti-stress mechanism[J]. Acta Horticulturae Sinica, 2023, 50(1):209—224. doi: 10.16420/j.issn.0513-353x.2021-1186
[6]	岑慧芳, 黄洁琼, 申王晖, 朱慧森, 夏方山, 许涛. 紫花苜蓿MsUGT87A1基因克隆及其对非生物胁迫的响应分析[J]. 草地学报, 2023, 31(6):1682—1692.doi:10.11733/j.issn.1007-0435.2023.06.010.
	Cen H F, Huang J Q, Shen W H, Zhu H S, Xia F S, Xu T. Cloning of MsUGT87A1 gene in alfalfa and analysis of its expression in the response to abiotic stresses[J]. Acta Agrestia Sinica, 2023, 31(6):1682—1692.
[7]	Morita Y, Hoshino A, Kikuchi Y, Okuhara H, Ono E, Tanaka Y, Fukui Y, Saito N, Nitasaka E, Noguchi H, Iida S. Japanese morning glory dusky mutants displaying reddish-brown or purplish-gray flowers are deficient in a novel glycosylation enzyme for anthocyanin biosynthesis,UDP-glucose:anthocyanidin 3-O-glucoside-2″-O-glucosyltransferase,due to 4-bp insertions in the gene[J]. The Plant Journal, 2005, 42(3):353—363.doi:10.1111/j.1365-313X.2005.02383.x.
[8]	Li P, Li Y J, Zhang F J, Zhang G Z, Jiang X Y, Yu H M, Hou B K. The Arabidopsis UDP-glycosyltransferases UGT79B2 and UGT79B3,contribute to cold,salt and drought stress tolerance via modulating anthocyanin accumulation[J]. The Plant Journal, 2017, 89(1):85—103.doi:10.1111/tpj.13324.
[9]	Ahrazem O, Rubio-Moraga A, Jimeno M L, Gómez-GÓmez L. Structural characterization of highly glucosylated crocins and regulation of their biosynthesis during flower development in Crocus[J]. Frontiers in Plant Science, 2015, 6:971.doi:10.3389/fpls.2015.00971. pmid: 26582258
[10]	袁美静, 马誉, 巫瑞, 康晓玲, 丁传雨, 杜丽. 影响月季花瓣呈色的理化因子及花色苷组分分析[J]. 西北植物学报, 2024, 44(2):255—269.doi:10.7606/j.issn.1000-4025.20230582.
	Yuan M J, Ma Y, Wu R, Kang X L, Ding J Y, Du L. Physicochemical characteristics and anthocyanin components affecting the color of rose petals[J]. Acta Botanica Boreali-Occidentalia Sinica, 2024, 44(2):255—269.doi:10.7606/j.issn.1000-4025.20230582.
[11]	姚虎成, 肖旖旎, 任禹齐, 夏墨晗, 李梓悦, 孔德信. 天然产物糖基转移酶的分类、进化及底物预测研究进展[J]. 生命科学, 2023, 35(9):1160—1168.doi:10.13376/j.cbls/2023127.
	Yao H C, Xiao Y N, Ren Y Q, Xia M H, Li Z Y, Kong D X. Progress on the classification,evolution and substrate prediction of natural product glycosyltransferase[J]. Chinese Bulletin of Life Sciences, 2023, 35(9):1160—1168.
[12]	周琳, 邹红竹, 韩璐璐, 贾莹华, 王雁. 糖基转移酶在花瓣色泽形成中的作用研究进展[J]. 园艺学报, 2022, 49(3):687—700.doi:10.16420/j.issn.0513-353x.2021-0636.
	Zhou L, Zou H Z, Han L L, Jia Y H, Wang Y. Research progress on the role of glycosyltransferases in color formation of petals[J]. Acta Horticulturae Sinica, 2022, 49(3):687—700. doi: 10.16420/j.issn.0513-353x.2021-0636
[13]	顾佳珺, 姚宇, 落艳娇, 王圆月, 申国安, 张丽娟, 郭宝林. 植物花青素糖基转移酶研究进展[J]. 生物资源, 2023, 45(3):201—209.doi:10.14188/j.ajsh.2023.03.001.
	Gu J J, Yao Y, Luo Y J, Wang Y Y, Shen G A, Zhang L J, Guo B L. Research progress of plant anthocyanin glycosyltransferase[J]. Biotic Resources, 2023, 45(3):201—209.
[14]	姚宇, 顾佳珺, 孙超, 申国安, 郭宝林. 植物类黄酮UDP-糖基转移酶研究进展[J]. 生物技术通报, 2022, 38(12):47—57.doi:10.13560/j.cnki.biotech.bull.1985.2022-0236.
	Yao Y, Gu J J, Sun C, Shen G A, Guo B L. Advances in plant flavonoids UDP-glycosyltransferase[J]. Biotechnology Bulletin, 2022, 38(12):47—57. doi: 10.13560/j.cnki.biotech.bull.1985.2022-0236
[15]	蔡启忠, 周良云, 刘露, 刘长征, 王浩, 钟磊, 杨全. 何首乌糖基转移酶基因的克隆、生物信息学及表达分析[J]. 中草药, 2021, 52(19):6013—6022.doi:10.7501/j.issn.0253-2670.2021.19.024.
	Cai Q Z, Zhou L Y, Liu L, Liu C Z, Wang H, Zhong L, Yang Q. Cloning,bioinformatic,and expression analysis of glycosyltransferase gene from Polygonum multiflorum Thunb[J]. Chinese Traditional and Herbal Drugs, 2021, 52(19):6013—6022.
[16]	Ju Z G, Sun W, Meng X Y, Liang L J, Li Y Q, Zhou T T, Shen H, Gao X, Wang L. Isolation and functional characterization of two 5-O-glucosyltransferases related to anthocyanin biosynthesis from Freesia hybrida[J]. Plant Cell,Tissue and Organ Culture, 2018, 135(1):99—110.doi:10.1007/s11240-018-1447-0.
[17]	Morita Y, Ishiguro K, Tanaka Y, Iida S, Hoshino A. Spontaneous mutations of the UDP-glucose:flavonoid 3-O-glucosyltransferase gene confers pale-and dull-colored flowers in the Japanese and common morning glories[J]. Planta, 2015, 242(3):575—587.doi:10.1007/s00425-015-2321-5.
[18]	Yang Y, Li B, Feng C Y, Wu Q, Wang Q Y, Li S S, Yu X N, Wang L S. Chemical mechanism of flower color microvariation in Paeonia with yellow flowers[J]. Horticultural Plant Journal, 2020, 6(3):179—190.doi:10.1016/j.hpj.2020.04.002.
[19]	Qu Y, Ou Z, Wang S. Coloration differences in three Meconopsis species:M.punicea,M.integrifolia and M.wilsonii[J]. South African Journal of Botany, 2022, 150:171—177.doi:10.1016/j.sajb.2022.07.016.
[20]	Yokoyama K, Yangzom R, Mizuno T, Murai Y, Dorji K, Wangmo C, Gyeltshen C, Iwashina T. Flavonol glycosides in the flowers of the Himalayan Meconopsis paniculata and Meconopsis integrifolia as yellow pigments[J]. Biochemical Systematics and Ecology, 2018, 81:102—104.doi:10.1016/j.bse.2018.10.006.
[21]	王卓为, 戴梦怡, 程少禹, 王小德, 王亚玲, 申亚梅, 张超. 紫玉兰红元宝Ml3GT1基因的克隆及表达分析[J]. 广西植物, 2022, 42(8):1417—1425.doi:10.11931/guihaia.gxzw202012045.
	Wang Z W, Dai M Y, Cheng S Y, Wang X D, Wang Y L, Shen Y M, Zhang C. Cloning and expression analysis of Ml3GT1 in Magnolia liliflora Hongyuanbao[J]. Guihaia, 2022, 42(8):1417—1425.
[22]	招雪晴, 杨静, 沈雨, 苑兆和. 石榴PgUGT基因表达特性与重组表达分析[J]. 西北植物学报, 2022, 42(3):390—397.doi:10.7606/j.issn.1000-4025.2022.03.0390.
	Zhao X Q, Yang J, Shen Y, Yuan Z H. Expression profiles and recombinant expression of PgUGT in pomegranate[J]. Acta Botanica Boreali-Occidentalia Sinica, 2022, 42(3):390—397.
[23]	梁沛雯, 娄倩, 刘雅莉. 葡萄风信子MaGT1基因的克隆及其表达分析[J]. 西北植物学报, 2019, 39(6):1001—1008.doi:10.7606/j.issn.1000-4025.2019.06.1001.
	Liang P W, Lou Q, Liu Y L. Cloning and expression analysis of MaGT1 gene from Muscari ameniacum[J]. Acta Botanica Boreali-Occidentalia Sinica, 2019, 39(6):1001—1008.

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