云南微小扇头蜱线粒体基因组全序列测定与分析

doi:10.12140/j.issn.1000-7423.2022.05.018

中国寄生虫学与寄生虫病杂志 ›› 2022, Vol. 40 ›› Issue (5): 677-681.doi: 10.12140/j.issn.1000-7423.2022.05.018

云南微小扇头蜱线粒体基因组全序列测定与分析

刘雅芳¹(), 陈彬¹, 芦新焱¹, 李光华¹, 杜春红², 姜丹丹³, 杨兴¹^,^*()

1.大理大学基础医学院,大理 671000
2.云南省地方病防治所,大理 671000
3.大理大学公共卫生学院,大理 671000

收稿日期:2022-01-24 修回日期:2022-04-09 出版日期:2022-10-30 发布日期:2022-08-02
通讯作者: 杨兴
作者简介:刘雅芳（1997-）,女,硕士研究生,从事人兽共患寄生虫病研究。E-mail： fangzai_05@sina.com
基金资助:
国家自然科学基金(81760607);国家自然科学基金(U2002219);云南省自然科学基金(2017FD139);云南省教育厅科学研究基金(2022J0687)

Complete mitochondrial genome sequence of Rhipicephalus microplus

LIU Ya-fang¹(), CHEN Bin¹, LU Xin-yan¹, LI Guang-hua¹, DU Chun-hong², JIANG Dan-dan³, YANG Xing¹^,^*()

1. School of Basic Medicine, Dali University, Dali 671000, China
2. Yunnan Institute for Endemic Diseases Control and Prevention, Dali 671000, China
3. School of Public Health, Dali University, Dali 671000, China

Received:2022-01-24 Revised:2022-04-09 Online:2022-10-30 Published:2022-08-02
Contact: YANG Xing
Supported by:
National Natural Science Foundation of China(81760607);National Natural Science Foundation of China(U2002219);Yunnan Natural Science Foundation(2017FD139);Scientific Research Fund of Yunnan Education Department(2022J0687)

摘要/Abstract

摘要：

2017年9月采用布旗法在云南省临沧市耿马县采集游离的微小扇头蜱,提取蜱DNA,采用通用引物PCR扩增微小扇头蜱的16S rRNA、12S rRNA、nad1、cox3基因片段,采用长PCR扩增上述4个基因区的长片段,产物纯化后采用步移法测序,拼接获得线粒体基因组全序列。结果显示,线粒体基因组全长14 763 bp,包含13个蛋白质编码基因、2个rRNA基因、22个tRNA和1个长度为264 bp的非编码区（D-Loop区）,碱基组成为A 38.66%、G 9.02%、C 11.46%、T 40.87%,（A + T）79.53%。蛋白质编码基因起始密码子均为ATN;除COX2、cox3、NAD5使用不完整T作为终止密码子外,其余基因终止密码子均为TAA或TAG;基因重叠区有12处共95 bp,长度在1~24 bp;基因间隔区有18处共433 bp,长度在1~311 bp。22个tRNA中,除tRNA^Cys、tRNA^Phe、tRNA^Ser以外均能形成典型的三叶草结构。

关键词: 微小扇头蜱, 线粒体基因组, 序列分析

Abstract:

Rhipicephalus microplus was collected by the cloth flag method in Yunnan Province in September 2017 and was used for DNA extraction. Four long fragments were amplified by long PCR using the primers designed based on 16S rRNA, 12S rRNA, nad1 and cox3 gene sequences. The fragments were sequenced by conserved primer-walking. The complete mitochondrial genome sequence was obtained by splicing. The length of the mitochondrial genome (mtDNA) of Rhipicephalus microplus is 14 763 bp. It contained 13 protein-coding genes, 2 rRNA genes, 22 tRNA genes and a non-coding control region of 264 bp (D-Loop region). The base composition was A 38.66%, G 9.02%, C 11.46%, T 40.87%. A + T content was 79.53%. The starting codon of protein-coding genes was ATN. The stop codons were TAA or TAG, except for cox2, cox3, nad5, which used incomplete T as the stop codon. The complete mitochondrial genome have 12 gene overlaps. The length ranges from 1-24 bp. The length of 18 total intergenic regions was 433 bp ranging from 1-311 bp. A total of 22 transferring RNA were found, all of which were typical cloverleaf structures except for tRNA^Cys, tRNA^Phe, tRNA^Ser.

Key words: Rhipicephalus microplus, Mitochondrial genome, Sequence analysis

中图分类号:

R384.41

刘雅芳, 陈彬, 芦新焱, 李光华, 杜春红, 姜丹丹, 杨兴. 云南微小扇头蜱线粒体基因组全序列测定与分析[J]. 中国寄生虫学与寄生虫病杂志, 2022, 40(5): 677-681.

LIU Ya-fang, CHEN Bin, LU Xin-yan, LI Guang-hua, DU Chun-hong, JIANG Dan-dan, YANG Xing. Complete mitochondrial genome sequence of Rhipicephalus microplus[J]. Chinese Journal of Parasitology and Parasitic Diseases, 2022, 40(5): 677-681.

图/表 5

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参考文献 19

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引物	引物序列（5′→3′）	扩增长度/bp
16SR	TGTACCTTTTGCATTAGGGATTTT	1 242
16SF	TCATTGAGCAGGATTTGCATTT
12SR	AGCTAAATTTGTGCCAGCAGC	703
12SF	AAGAGTGACGGGCGATATGT
nad1R	GGGCCCAAATAAAACTGGGGT	939
nad1F	ACTTGGGCAAATCCTCGATAAGA
COⅢR	TTCACGAGAAGGGTCTTACCAAG	778
COⅢF	TTAGGGGGCCATTGACTGCC

基因	位置	长度/bp	起始密码子	终止密码子	反密码子	编码链
tRNA^Cys	1~58	58			GCA	N
tRNA^Met	61~126	66			CAT	N
nad2	127~1 083	957	ATT	TAA		N
tRNA^Trp	1 085~1 148	64			TCA	N
tRNA^Tyr	1 150~1 213	64			GTA	J
cox1	1 206~2 744	1 539	ATT	TAA		N
cox2	2 749~3 421	673	ATG	T(AA)		N
tRNA^Lys	3 422~3 487	66			CTT	N
tRNA^Asp	3 487~3 552	66			GTC	N
ATP8	3 553~3 711	159	ATA	TAA		N
ATP6	3 705~4 376	672	ATT	TAA		N
cox3	4 385~5 162	778	ATG	T(AA)		N
tRNA^Gly	5 163~5 227	65			GGA	N
nad3	5 225~5 566	342	ATA	TAA		N
tRNA^Ala	5 570~5 631	62			TGC	N
tRNA^Arg	5 633~5 694	62			TCG	N
tRNA^Asn	5 693~5 755	63			GTT	N
tRNA^Ser	5 759~5 815	57			TCT	N
tRNA^Glu	5 817~5 881	65			TTC	N
nad1	5 880~6 818	939	ATT	TAA		J
tRNA^Leu	6 819~6 883	65			TAA	J
rrnL	6 860~8 101	1 242				J
tRNA^Val	8 083~8 142	60			TAC	J
rrnS	8 135~8 837	703				J
tRNA^Ile	9 149~9 213	65			GTA	N
tRNA^Gln	9 224~9 288	65			TTG	J
tRNA^Phe	9 314~9 373	60			GAA	J
nad5	9 374~11 027	1 645	ATT	T(AA)		J
tRNA^His	11 028~11 088	61			GTG	J
nad4	11 092~12 405	1 314	ATG	TAA		J
nad4L	12 399~12 674	276	ATG	TAA		J
tRNA^Thr	12 682~12 742	61			TGT	N
tRNA^Pro	12 743~12 807	65			TGG	J
nad6	12 795~13 244	450	ATA	TAA		N
cytb	13 249~14 325	1 077	ATG	TAG		N
tRNA^Ser	14 325~14 387	63			TGA	N
tRNA^Leu	14 396~14 457	62			TAG	J
D-Loop	14 488~14 751	264				N

基因区	碱基含量/%					长度/bp
基因区	A	G	T	C	A + T	长度/bp
全基因组	38.66	9.02	40.87	11.46	79.53	14 763
蛋白质编码基因	34.04	10.14	45.22	10.61	79.25	10 830
密码子第一位点	34.57	9.75	43.32	12.35	77.89	3 610
密码子第二位点	31.77	10.78	46.68	10.78	78.45	3 610
密码子第三位点	35.76	9.89	45.65	8.70	81.41	3 610
tRNA基因	40.29	10.83	40.36	8.52	80.65	1 385
rRNA基因	43.24	10.13	39.59	7.04	82.83	1 945
D-Loop区	31.06	17.05	37.12	14.77	68.18	264

密码子（氨基酸）	使用次数	密码子（氨基酸）	使用次数	密码子（氨基酸）	使用次数	密码子（氨基酸）	使用次数
UUU(F)	386.0	UCU(S)	96.0	UAU(Y)	127.0	UGU(C)	28.0
UUC(F)	32.0	UCC(S)	12.0	UAC(Y)	23.0	UGC(C)	5.0
UUA(L)	378.0	UCA(S)	114.0	UAA(*)	9.0	UGA(W)	73.0
UUG(L)	13.0	UCG(S)	3.0	UAG(*)	1.0	UGG(W)	9.0
CUU(L)	49.0	CCU(P)	57.0	CAU(H)	54.0	CGU(R)	14.0
CUC(L)	1.0	CCC(P)	17.0	CAC(H)	8.0	CGC(R)	2.0
CUA(L)	36.0	CCA(P)	36.0	CAA(Q)	58.0	CGA(R)	28.0
CUG(L)	3.0	CCG(P)	5.0	CAG(Q)	1.0	CGG(R)	0
AUU(I)	450.0	ACU(T)	63.0	AAU(N)	180.0	AGU(S)	21.0
AUC(I)	17.0	ACC(T)	6.0	AAC(N)	24.0	AGC(S)	9.0
AUA(M)	308.0	ACA(T)	52.0	AAA(K)	138.0	AGA(S)	67.0
AUG(M)	34.0	ACG(T)	2.0	AAG(K)	8.0	AGG(S)	9.0
GUU(V)	62.0	GCU(A)	62.0	GAU(D)	46.0	GGU(G)	51.0
GUC(V)	5.0	GCC(A)	9.0	GAC(D)	7.0	GGC(G)	21.0
GUA(V)	60.0	GCA(A)	46.0	GAA(E)	72.0	GGA(G)	62.0
GUG(V)	5.0	GCG(A)	2.0	GAG(E)	5.0	GGG(G)	28.0

云南微小扇头蜱线粒体基因组全序列测定与分析

Complete mitochondrial genome sequence of Rhipicephalus microplus

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