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浙江大学学报（农业与生命科学版）
Journal of Zhejiang University (Agriculture and Life Sciences)  2019, Vol. 45 Issue (1): 109-118    DOI: 10.3785/j.issn.1008-9209.2018.02.052
Animal sciences & veterinary medicine     
Single nucleotide polymorphism screening and bioinformatics analysis of the main family genes of FABPs in Bamei pig
Li ZHANG(),Weilun NONG,Jianxiong LU,Guohua ZHANG,Lixia LIU()
1. College of Life Science and Engineering, Northwest Minzu University, Lanzhou 730124, China
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Abstract  

Fatty acid-binding proteins belong to the intracellular lipid binding protein superfamily. All members of the FABPs family have the most basic functions to manage free fatty acid absorption and intracellular transport. In order to explore the regulation mechanism of the FABPs main family genes on the lipid deposition and metabolism in Bamei pigs, in this study, DNA pooling was constructed with Bamei pigs as test objects, and the single nucleotide polymorphisms (SNPs) of FABP1 , FABP2 , FABP3 and FABP4 genes in Bamei pigs were screened by segmented amplification and direct sequencing, and the bioinformatics analysis was carried out. The result showed that 15 SNPs were detected in the FABPs main family genes, in which there were three samesense and two missense mutation (C2 033A-Exon2-FABP1 , G3 751A-Exon3-FABP1 ). Bioinformatics analysis showed that the secondary structure of mRNA and free energy change before and after nucleotide mutation led to the change of its structural stability; there were differences in the estimation of allele frequency, mRNA secondary structure prediction, and protein secondary and tertiary structure prediction before and after the mutation. The above results indicate that the FABPs main family genes have high mutagenicity, which provides a molecular basis for the genetic effect study of intramuscular fat content in Bamei pigs.

Key words： Bamei pig      fatty acid-binding protein gene      single nucleotide polymorphisms      bioinformatics     
Received: 05 February 2018      Published: 28 March 2019
CLC:  S 831  
Corresponding Authors: Lixia LIU     E-mail: zhangli2008@aliyun.com;skllx@xbmu.edu.cn
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Li ZHANG
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Cite this article:

Li ZHANG,Weilun NONG,Jianxiong LU,Guohua ZHANG,Lixia LIU. Single nucleotide polymorphism screening and bioinformatics analysis of the main family genes of FABPs in Bamei pig. Journal of Zhejiang University (Agriculture and Life Sciences), 2019, 45(1): 109-118.

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http://www.zjujournals.com/agr/10.3785/j.issn.1008-9209.2018.02.052     OR     http://www.zjujournals.com/agr/Y2019/V45/I1/109


八眉猪FABPs主要家族基因单核苷酸多态性筛查及生物信息学分析

脂肪酸结合蛋白(fatty acid-binding protein, FABPs)属细胞内脂质结合蛋白超家族，FABPs家族所有成员都具有管理游离脂肪酸吸收和细胞内转运的最基本功能。为了探究FABPs主要家族基因在八眉猪体内脂肪沉积代谢过程中的调控机制，本研究以八眉猪为试验对象构建DNA混合池，采用分段扩增和直接测序法对八眉猪FABP1 、FABP2 、FABP3 、FABP4 基因进行单核苷酸多态性(single nucleotide polymorphisms, SNPs)快速筛查和生物信息学分析。结果表明，在八眉猪FABPs主要家族基因中共检测出15个SNPs，其中编码区有3个同义突变和2个错义突变(C2 033A-Exon2-FABP1 、G3 751A-Exon3-FABP1 )。对其进行生物信息学分析表明：核苷酸突变前后mRNA二级结构和自由能的改变导致其结构稳定性发生改变；突变前后的等位基因频率估算，mRNA二级结构预测，蛋白质二、三级结构预测分析均有差异。说明八眉猪FABPs主要家族基因具有较高的突变性，为研究八眉猪肌内脂肪含量的遗传效应提供了分子理论依据。


关键词： 八眉猪,  脂肪酸结合蛋白基因,  单核苷酸多态性,  生物信息学 

引物名称

Primer name

引物序列（5′→3′）

Primer sequences (5′→3′)

退火温度

Annealing temperature/℃

目的片段长度

Target fragment length/bp
FABP1 -Exon1 F:GCACTTGATTGACACCCAT 62.0 448
R:GTCCAGGGAGGAATAGCAT
FABP1 -Exon2 F:CCACTGACCTTCCCTTTC 61.0 555
R:TCCCTAAGACCTTCCTCC
FABP1 -Exon3 F:CTATTCAGAGCCAGAGCACA 58.0 535
R:GCAATGATGAGGGAGTAGTG
FABP1 -Exon4 F:ACTCCCTCATCATTGCCACAG 59.0 544
R:ACGAGAATCACCCTTCAGACC
FABP2 -Exon1 F:ATCATCCTTCAATGCAGCT 56.0 391
R:CACAACAGGCACTATTCCC
FABP2 -Exon2 F:CACAACAGGCACTATTCCC 55.5 313
R:CTCCATTGGCTGCTTCAGTA
FABP2 -Exon3 F:CTCCATTGGCTGCTTCAGTA 57.5 482
R:AATCAGAAGGCATCAGAAAG
FABP2 -Exon4 F:ACACTTTCTGATGCCTTCTG 59.0 590
R:AAATGCTGCAAATTATTCGA
FABP3 -Exon1

F:TCTCCTCTAGTCTCTCATCTCTG

R:GTCGTCCTCACCAATTGACTTC

 61.5 521
FABP3 -Exon2 F:AGCCTTTGAAAATTCTTGCCCT 55.5 544
R:GACACGGAGGTCAGAATATCCT
FABP3 -Exon3 F:AGCTGGGCTGTCTGACTC 60.0 474
R:CCTCCACCCTCCACTATC
FABP3 -Exon4 F:TGAAGACCTGGTGTAAGCA 60.0 386
R:ACAACAAGAACCGGAACTG
FABP4 -Exon1 F:GGAGAACCAAAGTTGAGAAA 57.0 382
R:CAGAGTGAAAAACAGCCATA
FABP4 -Exon2 F:ACACACATACACGCATTCC 58.5 556
R:TTTTTTTCCCTCTTTTCCC
FABP4 -Exon3 F:GTATGCTGTTGCTTTTGGT 60.5 469
R:ATTGGAAGAGGTTAGGTGG
FABP4 -Exon4 F:CTTCCCACCATTGGAGAA 55.5 623
R:CCTAACACGGGCAACTTC
Table 1 Informations for primers of FABPs genes in Bamei pig
Fig. 1 Results of sequencing of the main family genes for FABPs and BLAST analysis in Bamei pig

基因类型

Gene type

突变位点

Mutation site

突变类型

Mutation type

氨基酸变化

Amino acid variation

 等位基因频率Allele frequency

突变前频率

Before mutation

突变后频率

After mutation
FABP1 A57G-Exon1-FABP1 — — 0.147 5 0.852 5
T2 006C-Exon2-FABP1 同义突变 — 0.214 3 0.785 7
A2 011C-Exon2-FABP1 同义突变 — 0.260 3 0.739 7
C2 033A-Exon2-FABP1 错义突变 L74M 0.205 5 0.794 5
G3 751A-Exon3-FABP1 错义突变 G87S 0.567 6 0.432 4
T3 943A-Intron3-FABP1 — — 0.615 4 0.384 6
T4 303C-Intron3-FABP1 — — 0.738 5 0.261 5
FABP2 A－207T-5′UTR-FABP2 — — 0.827 6 0.172 4
FABP3 G－332A-5′UTR-FABP3 — — 0.382 4 0.617 6
C807T-Intron1-FABP3 — — 0.687 5 0.312 5
C3 073T-Exon3-FABP3 同义突变 — 0.779 2 0.220 8
FABP4 G109C-Intron1-FABP4 — — 0.780 5 0.219 5
A4 474G-3′UTR-FABP4 — — 0.894 1 0.105 9
T4 487A-3′UTR-FABP4 — — 0.173 9 0.826 1
C4 536T-3′UTR-FABP4 — — 0.837 8 0.162 2
Table 2 SNPs locus mutation types and estimation of allele frequency of FABPs gene in Bamei pig
Fig. 2 Results of secondary structure change of mRNA
突变位点 突变类型

α-螺旋

Alpha helix/%

β-转角

Beta turn/%

无规则卷曲

Random coil/%

扩展链

Extended strand/%
C2 033A-Exon2-FABP1

突变前

Before mutation

 21.26（27） 15.75（20） 24.41（31） 38.58（49）

突变后

After mutation

 20.47（26） 15.75（20） 25.20（32） 38.58（49）
G3 751A-Exon3-FABP1

突变前

Before mutation

 20.47（26） 15.75（20） 25.20（32） 38.58（49）

突变后

After mutation

 20.47（26） 15.75（20） 25.20（32） 38.58（49）
Table 3 Analysis of prediction of secondary protein structure with different mutations
Fig. 3 Results of tertiary structure change of protein
[1]   刘丽霞,张丽,赵生国,等 .八眉猪DRA基因编码区多态性及生物信息学分析.华北农学报,2015,30(4):66-71.
LIU L X , ZHANG L , ZHAO S G , et al . Cloning and bioinformatics analysis on coding regions of DRA gene in Bamei Pigs. Acta Agricultural Borealisinica, 2015,30(4):66-71. (in Chinese with English abstract)
[2]   田鎏,张昌军,刁红录 .脂肪酸结合蛋白研究进展.生命科学,2017,29(2):164-169.
TIAN L , ZHANG C J , DIAO H L . Advances in fatty acid binding proteins research. Chinese Bulletin of Life Sciences, 2017,29(2):164-169. (in Chinese with English abstract)
[3]   CHMURZY?SKA A . The multigene family of fatty acid-binding proteins (FABPs): function, structure and polymorphism. Journal of Applied Genetics, 2006,47(1):39-48.
[4]   徐海霞,房兴堂,陈宏,等 .脂肪酸结合蛋白及其主要家族基因的研究进展.畜牧与兽医,2008,40(1):94-97.
XU H X , FANG X T , CHEN H , et al . Advances of the main family genes in fatty acid binding proteins research. Animal Husband＆Veterinary Medicine, 2008,40(1):94-97. (in Chinese)
[5]   许家磊,王宇,后猛,等 .SNP检测方法的研究进展.分子植物育种,2015,13(2):475-482.
XU J L , WANG Y , HOU M , et al . Progress on detection methods of SNP. Molecular Plant Breeding, 2015,13(2):475-482. (in Chinese with English abstract)
[6]   陈桂莲 .猪L-FABP基因的遗传多态性及其与肌内脂肪含量的相关性研究.河北，保定:河北农业大学,2008:30-31.
CHEN G L . Studies on genetic variation of L-FABP genes and the associations with intramuscular fat content in pigs. Baoding, Hebei: Agricultural University of Hebei, 2008:30-31. (in Chinese with English abstract)
[7]   刘智 . BMPER和FABP2基因启动子区变异与猪肌内脂肪含量相关性研究.南京:南京农业大学,2014:33-39.
LIU Z . Correlation between variations in promoter region of BMPER and FABP2 genes and intramuscular fat content in pigs. Nanjing: Nanjing Agricultural University, 2014:33-39. (in Chinese with English abstract)
[8]   周春宝,朱淑斌,赵旭庭,等 .苏姜猪H-FABP基因多态性及其与肉质性状的相关性.江苏农业学报,2014,30(6):1369-1374.
ZHOU C B , ZHU S B , ZHAO X T , et al . Genetic polymorphism of H-FABP gene in Sujiang swine and its correlation with meat quality. Jiangsu Journal of Agricultural Sciences, 2014,30(6):1369-1374. (in Chinese with English abstract)
[9]   朱爱文,倪黎刚,周春宝,等 .苏姜猪A-FABP基因遗传多态性及其与肉质性状的关联分析.中国畜牧杂志,2017,53(3):29-33.
ZHU A W , NI L G , ZHOU C B , et al . Genetic polymorphism of A-FABP gene and correlation analysis with meat quality traits in Sujiang swine. Chinese Journal of Animal Science, 2017,53(3):29-33. (in Chinese with English abstract)
[10]   SULIMAN H B , CARRAWAY M S , PIANTADOSI C A . Postlipopolysaccharide oxidative damage of mitochondrial DNA. American Journal of Respiratory and Critical Care Medicine, 2003,167(4):570-579.
[11]   张丽,刘丽霞,李强子,等 .天祝白牦牛MSTN基因编码区克隆及生物信息学分析.浙江农业学报,2017,29(4):618-624.
ZHANG L , LIU L X , LI Q Z , et al . Cloning and bioinformatics analysis of MSTN gene of Tianzhu white yak. Acta Agriculturae Zhejianggensis, 2017,29(4):618-624. (in Chinese with English abstract)
[12]   盘道兴,王振,杨茂林,等 .江口萝卜猪FABPs主要家族基因SNPs筛选及生物信息学分析.基因组学与应用生物学, 2015,34(1):47-52.
PAN D X , WANG Z , YANG M L , et al . SNP screening and bioinformatics analysis of the main family genes of FABPs in Jiangkou Luobo Pig. Genomics and Applied Biology, 2015,34(1):47-52. (in Chinese with English abstract)
[13]   祝仁铸,尹逊河,王元虎,等 .猪肌肉组织MDH和LPL基因表达与肌内脂肪含量和脂肪酸组成关系的研究.畜牧兽医学报,2013,44(8):1182-1188.
ZHU R Z , YI X H , WANG Y H , et al . The gene expression of MDH and LPL in muscle and their association with content of intramuscular fat and composition of fatty acids in pigs. Acta Veterinaria et Zootechnica Sinica, 2013,44(8):1182-1188. (in Chinese with English abstract)
[14]   苏玉虹,熊远著,邓昌彦 .猪的肉质性状基因定位研究进展.遗传,2000,22(5):334-338.
SU Y H , XIONG Y Z , DENG C Y . The progress of mapping QTL affecting meat quality in pig. Hereditas, 2000,22(5):334-338. (in Chinese with English abstract)
[15]   HOVENIER R , KANIS E , ASSELDONK T V , et al . Genetic parameters of pig meat quality traits in a halothane negative population. Livestock Production Science, 1992,32(4):309-321.
[16]   JANSS L , ARENDONK J A M VAN , BRASCAMP P . Identification of a single gene affecting intramuscular fat in Meishan crossbreds using Gibbs sampling. Transactions-Pennsylvania Academy of Ophthalmology and Otolaryngology, 1994,18(2):25-27.
[17]   陈志辉,徐良梅,单安山 .脂肪酸结合蛋白及其基因.东北农业大学学报,2006,37(5):689-692.
CHEN Z H , XU L M , SHAN A S . Fatty acid-binding proteins (FABPs) and their genes of FABPs. Journal of Northeast Agricultural University, 2006,37(5):689-692. (in Chinese with English abstract)
[18]   WANG S C , ZHU Y T , LI X J , et al . Fatty acid binding protein regulate antimicrobial function via Toll signaling in Chinese mitten crab. Fish & Shellfish Immunology, 2017,63:9-17.
[19]   AITKEN N , SMITH S C , MORIN P , et al . Single nucleotide polymorphism (SNP) discovery in mammals: a targeted-gene approach. Molecular Ecology, 2004,13(6):1423-1431.
[20]   梁艳 .猪Myf-5基因和L-FABP基因的遗传多态性及其与胴体和肉质性状间关联性研究.四川,雅安:四川农业大学,2007:43-44.
LIANG Y . Studies on the genetics polymorphism of Myf-5 gene and L-FABP gene and the association with carcass and meat quality traits in pigs. Ya'an, Sichuan: Sichuan Agricultural University, 2007:43-44. (in Chinese with English abstract)
[21]   刘利刚,张树敏,李兆华,等 .松辽黑猪L-FABP基因C276A位点多态性及其与肉质性状的相关性分析.养猪,2010(3):73-75.
LIU L G , ZHANG S M , LI Z H , et al . C276A polymorphism analysis of L-FABP gene and correlation with meat quality traits in Songliao black pigs. Swine Production, 2010(3):73-75. (in Chinese)
[22]   徐松松,安荣荣,陈耀峰,等 .鸡FABP2基因单倍型与生长和体组成性状的相关分析.中国家禽,2016,38(8):5-9.
XU S S , AN R R , CHEN Y F , et al . Association of haplotypes of FABP2 gene with growth and body composition traits in chickens. China Poultry, 2016,38(8):5-9. (in Chinese with English abstract)
[23]   GERBENS F , RETTENBERGER G , LENSTRA J A , et al . Characterization, chromosomal localization, and genetic variation of the porcine heart fatty acid-binding protein gene. Mammalian Genome, 1997,8(5):328-332.
[24]   朱淑斌,赵旭庭,周春宝,等 .姜曲海猪A-FABP基因内含子1多态性与肉质性状的相关分析.中国畜牧杂志,2013,49(13):12-15.
ZHU S B , ZHAO X T , ZHOU C B , et al . Intron 1 polymorphism of A-FABP gene and correlation with meat quality traits in Jiangquhai pigs. Chinese Journal of Animal Science, 2013,49(13):12-15. (in Chinese)
[25]   张金耀,杨建敏,叶炎,等 .莆田黑猪A-FABP基因的群体遗传结构分析.黑龙江畜牧兽医,2015(15):94-97.
ZHANG J Y , YANG J M , YE Y , et al . Analysis of population genetic structure of the A-FABP gene in Putian black pig. Heilongjiang Animal Science and Veterinary Medicine, 2015(15):94-97. (in Chinese with English abstract)
[26]   LIN Z G , LI W H . Evolution of 5′ untranslated region length and gene expression reprogramming in yeasts. Molecular Biology & Evolution, 2012,29(1):81-89.
[27]   部德群,李秋玲,王长法,等 .牛α1-抗胰蛋白酶基因5′侧翼区新SNP的鉴定及其与中国荷斯坦奶牛产奶性状的相关性.中国生物化学与分子生物学报,2009,25(7):627-632.
BU D Q , LI Q L , WANG C F , et al . Identification of new SNPs in bovine α1-antitrypsin gene 5′flanking region and their association with milk production traits in Chinese Holstein Dairy Cattle. Chinese Journal of Biochemistry and Molecular Biology, 2009,259(7):627-632. (in Chinese with English abstract)
[28]   王海震,王莹,刘定干 .真核生物mRNA 3′非翻译区的功能.生物化学与生物物理进展,2008,35(9):980-985.
WANG H Z , WANG Y , LIU D G . Functions of the 3′ untranslated region of eukaryotic mRNA. Progress in Biochemistry and Biophysics, 2008,35(9):980-985. (in Chinese with English abstract)
[29]   孙瑞青,赵严,贾蓉蓉,等 .L-FABP与脂类代谢相关疾病的研究进展.胃肠病学和肝病学杂志,2014,23(4):368-372.
SUN R Q , ZHAO Y , JIA R R , et al . Research progress of liver fatty acid binding protein and lipid metabolic disease. Chinese Journal of Gastroenterology and Hepatology, 2014,23(4):368-372. (in Chinese with English abstract)
[30]   斯琴,傅研,王旭东,等 . I-FABP2基因多态性与2型糖尿病的相关性研究.中国医药导刊,2012,14(6):930-933.
SI Q , FU Y , WANG X D , et al . Association between polymorphism of the fatty acid-binding protein 2 gene and type 2 diabetes. Chinese Journal of Medicinal Guide, 2012,14(6):930-933. (in Chinese with English abstract)
[31]   初丽丽,王启贵,关天竹,等 . I-FABP基因侧翼区多态性与鸡生长和胴体组成性状的相关研究.东北农业大学学报,2008,39(9):70-74.
CHU L L , WANG Q G , GUAN T Z , et al . Association analyses between SNPs in the flanking region of I-FABP gene and growth and carcass composition traits. Journal of Northeast Agricultural University, 2008,39(9):70-74. (in Chinese with English abstract)
[32]   张越,陆萌,朱卫华,等 .猪心脏脂肪酸结合蛋白基因多态性检测及与肉质性状的关联分析.中国畜牧杂志,2016,52(1):7-12.
ZHANG Y , LU M , ZHU W H , et al . Detection of polymorphism in heart fatty acid binding protein gene and its association with pork quality in swine. Chinese Journal of Animal Science, 2016,52(1):7-12. (in Chinese with English abstract)
[33]   高妍,张永宏,熊秋宏,等 .猪A-FABP基因5′调控区多态性及与肉质性状的相关分析.畜牧与兽医,2010,42(5):34-37.
GAO Y , ZHANG Y H , XIONG Q H , et al . 5′ untranslated region polymorphism of A-FABP gene and correlation with meat quality traits in pigs. Animal Husbandry＆Veterinary Medicine, 2010,42(5):34-37. (in Chinese)
[34]   于小燕 .松辽黑猪A-FABP基因SNPs位点及其与肉质性状的相关分析.长春:吉林大学,2008:60-64.
YU X Y . SNPs analysis of A-FABP gene and correlation with meat quality traits in Songliao black pigs. Changchun: Jilin University, 2008:60-64. (in Chinese with English abstract)
[35]   JIANG Y Z , LI X W , YANG G X . Sequence characterization,tissue-specific expression and polymorphism of the porcine(Sus scrofs) liver-type fatty acid binding protein gene. Acta Genetica Sinica, 2006,33(7):598-606.
[36]   廖秀冬,任立明,王光瑛,等 .北京鸭FABP2基因多态性与体尺和屠体性状的相关性研究.中国家禽,2012,34(17):23-26,30.
LIAO X D , REN L M ， WANG G Y , et al . Polymorphism of FABP2 gene and its association with body size and carcass traits in Peking Duck. China Poultry, 2012,34(17):23-26,30. (in Chinese with English abstract)
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