Genomic Sequence Cloning and the Characteristics Analysis of Cowpea mild mottle virus Infecting Soybean from Jiangsu Province

doi:10.7668/hbnxb.20193080

Abstract

Abstract:

To characterize the genomic structure of Cowpea mild mottle virus (CpMMV)Jiangsu isolate and clarify its taxonomic status and evolutionary characteristics,4 pairs of specific primers were designed and the genomic fragments were amplified using cDNA obtained from reverse transcription of total RNA of a soybean sample collected from Jiangsu as a temple.The amplified products were cloned and sequenced,and the obtained fragments were assembled to get the full genomic sequence.Sequence analysis revealed the genome of Jiangsu isolate were 8 194 bp in length,containing 6 ORFs and 2 untranslated regions(UTRs):5' UTR(72 nt)and 3' UTR(117 nt).Among the six proteins encoded by CpMMV,CP shared highest amino acid sequence identities with other isolates(96.5%-100.0%),while RdRp(81.1%-98.2%),TGB1(81.0%-97.0%)and TGB3(80.9%-95.6%)shared relatively low sequence identities,indicating CP might be a conservative gene and RdRp,TGB1,TGB3 might show good diversity in different isolates.The phylogenetic analysis based on the whole genomic sequence revealed that Jiangsu isolate was clustered with other CpMMV isolates in a branch and it shared highest nucleotide sequence identities with 2 Chinese isolates(98.2% with Anhui isolate,96.0% with Hainan isolate),compared with other foreign isolates such as the United States,Brazil,India,Mexico,Kenya and so on(<82.7%).These results clarified the genomic structure of Jiangsu isolate,which shared similar character with other CpMMV isolates and phylogenetic analysis revealed Jiangsu isolates clustered with other isolates from China with higher sequences identities,compared with foreign isolates.

Key words: Soybean, Cowpea mild mottle virus, Genomic sequence, Clone, Characteristics analysis

JI Ying, SUN Feng, WU Shuhua, LI Shuo, TU Liqin, GAO Danna, CUI Xiaoyan, CHEN Xin, JI Yinghua, GUO Qingyun. Genomic Sequence Cloning and the Characteristics Analysis of Cowpea mild mottle virus Infecting Soybean from Jiangsu Province[J]. Acta Agriculturae Boreali-Sinica, 2022, 37(3): 200-205. doi: 10.7668/hbnxb.20193080.

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References 24

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引物 Primer	序列(5'-3') Sequence(5'-3')	退火温度/℃ Tm	片段/bp Fragment
CpMM-1F	GAAAAACAGAGCAACAATATCTAGC	53.60	2 355
CpMM-1R	CTTCATGCTTTCCACATCTTCC	52.92
CpMM-2F	GATATTGCCTGACAGGTCTGAT	52.70	2 568
CpMM-2R	TGATTCTCAATAGTGGCTGCAG	54.80
CpMM-3F	TCTCCAAGCCTCACGTTGAG	54.74	2 141
CpMM-3R	TATTGCAGACATAACAGACTCTTG	53.33
CpMM-4F	TCAACACAGRTTCCCTTTTGG	51.05	1 701
CpMM-4R	AAAACCAGGAAAATAACWTTAAAACC	50.22

引物 Primer	序列(5'-3') Sequence(5'-3')	退火温度/℃ Tm	片段/bp Fragment
CpMM-1F	GAAAAACAGAGCAACAATATCTAGC	53.60	2 355
CpMM-1R	CTTCATGCTTTCCACATCTTCC	52.92
CpMM-2F	GATATTGCCTGACAGGTCTGAT	52.70	2 568
CpMM-2R	TGATTCTCAATAGTGGCTGCAG	54.80
CpMM-3F	TCTCCAAGCCTCACGTTGAG	54.74	2 141
CpMM-3R	TATTGCAGACATAACAGACTCTTG	53.33
CpMM-4F	TCAACACAGRTTCCCTTTTGG	51.05	1 701
CpMM-4R	AAAACCAGGAAAATAACWTTAAAACC	50.22

登录号 Accession No.	依赖RNA的RNA 聚合酶 RdRp	三重基因块1 TGB1	三重基因块2 TGB2	三重基因块3 TGB3	外壳蛋白 CP	核酸结合蛋白 NBP
MN908944	98.2	97.0	99.1	95.6	100.0	98.1
KY420906	96.4	93.1	97.2	95.6	100.0	93.2
KC774019	86.2	86.6	87.8	82.4	98.3	92.2
KC774020	86.2	86.1	91.5	82.4	98.6	93.2
MT473963	85.9	87.0	92.5	85.3	98.6	91.3
KC884244	85.4	86.6	90.6	80.9	97.9	92.2
KC884245	86.0	86.1	91.5	83.8	98.3	92.2
KC884246	85.2	86.1	92.5	83.8	97.6	91.3
KC884247	81.3	86.1	92.5	83.8	97.6	91.3
KC884249	83.5	86.6	92.5	82.4	98.3	93.2
MK069989	86.1	85.7	91.5	82.4	98.6	94.2
MK984605	86.3	86.1	90.6	83.8	96.5	87.4
KC884248	85.2	81.4	84.0	82.4	98.3	87.4
MK202583	85.9	81.0	85.8	82.4	99.3	91.3
MH345698	81.1	83.5	86.8	83.8	98.6	89.3

登录号 Accession No.	依赖RNA的RNA 聚合酶 RdRp	三重基因块1 TGB1	三重基因块2 TGB2	三重基因块3 TGB3	外壳蛋白 CP	核酸结合蛋白 NBP
MN908944	98.2	97.0	99.1	95.6	100.0	98.1
KY420906	96.4	93.1	97.2	95.6	100.0	93.2
KC774019	86.2	86.6	87.8	82.4	98.3	92.2
KC774020	86.2	86.1	91.5	82.4	98.6	93.2
MT473963	85.9	87.0	92.5	85.3	98.6	91.3
KC884244	85.4	86.6	90.6	80.9	97.9	92.2
KC884245	86.0	86.1	91.5	83.8	98.3	92.2
KC884246	85.2	86.1	92.5	83.8	97.6	91.3
KC884247	81.3	86.1	92.5	83.8	97.6	91.3
KC884249	83.5	86.6	92.5	82.4	98.3	93.2
MK069989	86.1	85.7	91.5	82.4	98.6	94.2
MK984605	86.3	86.1	90.6	83.8	96.5	87.4
KC884248	85.2	81.4	84.0	82.4	98.3	87.4
MK202583	85.9	81.0	85.8	82.4	99.3	91.3
MH345698	81.1	83.5	86.8	83.8	98.6	89.3

登录号 Accession No.	长度/nt Length	病毒名称 Virus	寄主 Host	分离物 Isolate	国家/地区 Country/Region	同源性/% Identities
MN908944	8 256	豇豆轻斑驳病毒	大豆	Anhui_SZ_DN1383	中国/安徽	98.2
KY420906	8 193	豇豆轻斑驳病毒	豇豆	Hainan 1	中国/海南	96.0
KC774019	8 207	豇豆轻斑驳病毒	烟粉虱	Florida_Whiteflies_2007	美国	82.7
KC774020	8 209	豇豆轻斑驳病毒	菜豆	Florida_Bean_2011	美国	82.7
MT473963	8 195	豇豆轻斑驳病毒	大豆	Casa Branca_BR	巴西	82.6
KC884244	8 196	豇豆轻斑驳病毒	大豆	CPMMV:BR:MG:09:2	巴西	82.5
KC884245	8 196	豇豆轻斑驳病毒	大豆	CPMMV:BR:MG:09:3	巴西	82.6
KC884246	8 180	豇豆轻斑驳病毒	大豆	CPMMV:BR:MT:02:1	巴西	82.6
KC884247	8 191	豇豆轻斑驳病毒	大豆	CPMMV:BR:BA:02	巴西	81.7
KC884249	8 194	豇豆轻斑驳病毒	大豆	CPMMV:BR:GO:10:5	巴西	82.2
MK069989	8 191	豇豆轻斑驳病毒	菜豆	CN2	墨西哥	82.5
MK984605	8 196	豇豆轻斑驳病毒	番木瓜	KE-Kit_01	肯尼亚	82.6
KC884248	8 198	豇豆轻斑驳病毒	大豆	CPMMV:BR:GO:01:1	巴西	80.3
MK202583	8 194	豇豆轻斑驳病毒	菜豆	CPMMV:BR:GO:14	巴西	80.9
MH345698	8 189	豇豆轻斑驳病毒	黑吉豆	CpMMV-Urd-Kanpur	印度	76.6
HQ184471	8 127	豇豆轻斑驳病毒	豇豆	Ghana	加纳	64.7
NC_011525	7 890	马铃薯潜隐病毒	马铃薯		加拿大	54.0
NC_001361	8 533	马铃薯M病毒	番茄	Russian wild	俄罗斯	51.5
NC_043081	5 218	黄瓜脉明病毒	西瓜	PV-0985	坦桑尼亚	60.8
NC_007289	8 478	马铃薯S病毒	马铃薯	Leona	德国	49.9
NC_006550	9 104	甘薯褪绿斑点病毒	甘薯	Hoima 4	乌干达	49.3

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