黑线仓鼠OB基因部分序列的克隆与SNP分析
摘要
黑线仓鼠是我国北方的一种主要害鼠,由于其分布范围广,繁殖力极强,严重危害农业的发展和人类正常的生产生活,所以越来越多的研究围绕其展开。本论文主要是对黑线仓鼠进行了分子生物学方面的研究。
OB基因主要在脂肪组织中表达,其表达产物瘦素(Leptin)是新近发现的一种蛋白质激素。它对动物摄食、能量代谢、脂肪存储、生殖活动、胚胎及青春期发育、造血功能、神经内分泌、免疫等都有重要的调控作用,对于维持种属的生存与延续具有重要的生物学意义。
本研究通过对OB基因的克隆和分析来探讨黑线仓鼠种群间的遗传结构。研究根据GeneBank数据库中包括大鼠、小鼠等多种近源物种的OB基因设计引物,利用PCR法首次在黑线仓鼠中克隆出OB基因1665(1664)bp,其中包含完整内含子2区段和外显子2、外显子3部分区段,外显子区段包含完整的OB基因CDS序列,翻译后得到包含167个氨基酸残基的蛋白质序列；同时,在本实验中还从沂南、临朐和曲阜三个地理种群共随机挑选103个黑线仓鼠个体分别进行扩增和序列测定,结果发现19个SNPs,1个插入／缺失位点,共定义了34种单倍型,在GenBank注册序列号为：HM100043-HM100076。将实验获得的黑线仓鼠OB基因中mRNA序列与其它多个物种的相应序列进行比对,与大部分哺乳动物在核苷酸水平有较高同源性(82.3%-100%),同时在氨基酸水平上也具有较高同源性(70.1%-100%)。用DnaSPv5.0软件分别计算三个地理种群黑线仓鼠OB基因核苷酸多样性(π)、单倍型数目(H)、单倍型多样性(Hd)、平均核苷酸差异数(K)、多态位点数(S)和基于分离位点数目的多样性(θw),发现遗传多样性水平具有临朐种群>沂南种群>曲阜种群的特点。三个地理种群间的种群间遗传分化系数(Fst)、基因流(Nm)、种群间的核苷酸歧异度(Dxy)等显示沂南、临朐和曲阜三个地理种群黑线仓鼠OB基因均具有一定遗传分化但不显著。在种群间,曲阜和临朐两个地区遗传分化最大,基因交流较小；曲阜和沂南地区次之；临朐和沂南地区遗传分化最小,基因交流较大。种群遗传分化系数(Gst)在0.00455～0.06523之间,说明有93%以上的变异来自于种群内部。实验中用MEGA4.0软件对三个地理种群黑线仓鼠个体做NJ系统进化分析,结果表明三个地理种群黑线仓鼠遗传分化不明显。本研究发现山东地区黑线仓鼠OB基因存在丰富的核苷酸多样性和单倍型多样性,沂南、临朐和曲阜三个地理种群间遗传分化程度很小。
本研究还测定了66个黑线仓鼠的体脂含量,并对其OB基因的序列进行分析,结果发现,在第三外显子编码区上发现三个同义突变C263T、A481G、A511G,OB基因编码区存在4种单倍型。经SPSS17.0分析,四种单倍型的黑线仓鼠的体脂含量均存在显著性差异(P<0.05),由于这些突变不能引起氨基酸产物序列的变化,基本不影响蛋白质活性,从而不会使动物表现出明显的性状变化,故认为此基因突变并不是性状变异的主要原因。不同单倍型黑线仓鼠体脂含量的显著性差异可能是由其他原因引起的。不同单倍型与脂肪含量之间的关系以及OB基因多态性对脂肪蓄积的基因效应还会通过扩大样本后作进一步研究。另外,关于OB基因与黑线仓鼠繁殖、免疫等功能在基因水平上的相关性也有待于进一步研究。
Cricetulus barabensis are widely distributed in China and have strong reproductive ability. The species have been considered as deleterious animals for agriculture and industry, which is a serious threat to people'normal production and life. In recent years, C. barabensis as the dominant rodent species in North of China has evoked more and more concerns among the academic circles, and many researches outcomes spring. In these experiments the author investigate from the viewpoint of molecular biology.
The obese gene (OB) mainly expressed in adipose tissue. Leptin is the expression product of obese gene which is a peptide hormone discovered recently. Leptin plays an important role in animal physiology function such as ingestion, energy metabolism, reproduction activity, fat storage, fetal growth, puberty development, tumour cell proliferation, hemopoiesis, neuroimmunoendocrine and so on. It is of the most important biological significance for the survival and reproduction.
In order to investigation the population hereditary constitution of C. barabensis, in this thesis, the partial sequence of obese gene was amplifid and sequenced.1665bp of OB of the species in Shandong Province was amplified by polymerase chain reaction for the first time. The primers were designed on the basis of the homologous species from GenBank consisting of mice, rat and human. The sequence of OB covers partial exon2, total intron2 and partial exon3. The sequence were then cloned and sequenced in 3 geographic populations in Shandong Province: Yinan, Linqu and Qufu. A total of 19 SNPs were examined, including lindels Insert/Deletion and the ratio of conversion to transversion was 15/4.34 haplotypes were defined in 103 samples, The sequence numbers submitted to the GenBank were:HM100043～HM100076. The sequence numbers submitted to the GenBank were～.The nucleotide diversity (π), number of haplotye (H), haplotype diversity (Hd), average number of nucleotide differences (K), number of polymorphic sites (S) and theta from S (θw) of OB of Yinan, Linqu and Qufu populations were calculated with DnaSPv5.0 software, which showed that the genetic diversity of the three populations arrayed as (from high to low):Linqu,Yinan, Qufu. In addition, Fst, Nm and Dxy proved the genetic differentiation among the three populations existed in a low level. The result of Gst analysis among three populations arrayed from 0.00455 to 0.06523, which meant that above 93% mutation existed in each population while few mutation existed among populations. NJ analysis showed that the genetic differentiation among the three populations was not distinct. The research showed that OB on C. barabensis from Qufu,Yinan and Linqu was rich in haplotype diversity and nucleotide diversity. The genetic differentiation among the three populations:Qufu,Yinan,Linqu of C. barabensis are small.
In the study, by analyzing the sequence of OB on C. barabensi with different body fat percent we found three cSNPs in coding section:C/T, A/G and A/G, all of them are samesense mutation. We analized the coding section of OB in 66 samples, then found 4 haplotypes. The data were analyzed statistically with SPSS17.0. The result shows that there is significant difference of body fat percent among 4 haplotypes. Because these mutations couldn't change the sequence of Amino Acids Production, the protein activity wasn't affected, and obvious trait variation didn't appeared on animals, so I think gene mutation was not the main reason for trait variation. Any other reasons may lead to the significant difference among body fat percent of different haplotype of C. barabensis. And in this study, the relationships between different haplotype and body fat percent, the genetic effect of OB on body fat deposition are needed to be further studied by enlarging the samples. Besides, concerning the correlation on gene level between OB and functions such as breeding and immunity of Striped Hamster, further research is also needed.
OB基因主要在脂肪组织中表达,其表达产物瘦素(Leptin)是新近发现的一种蛋白质激素。它对动物摄食、能量代谢、脂肪存储、生殖活动、胚胎及青春期发育、造血功能、神经内分泌、免疫等都有重要的调控作用,对于维持种属的生存与延续具有重要的生物学意义。
本研究通过对OB基因的克隆和分析来探讨黑线仓鼠种群间的遗传结构。研究根据GeneBank数据库中包括大鼠、小鼠等多种近源物种的OB基因设计引物,利用PCR法首次在黑线仓鼠中克隆出OB基因1665(1664)bp,其中包含完整内含子2区段和外显子2、外显子3部分区段,外显子区段包含完整的OB基因CDS序列,翻译后得到包含167个氨基酸残基的蛋白质序列；同时,在本实验中还从沂南、临朐和曲阜三个地理种群共随机挑选103个黑线仓鼠个体分别进行扩增和序列测定,结果发现19个SNPs,1个插入／缺失位点,共定义了34种单倍型,在GenBank注册序列号为：HM100043-HM100076。将实验获得的黑线仓鼠OB基因中mRNA序列与其它多个物种的相应序列进行比对,与大部分哺乳动物在核苷酸水平有较高同源性(82.3%-100%),同时在氨基酸水平上也具有较高同源性(70.1%-100%)。用DnaSPv5.0软件分别计算三个地理种群黑线仓鼠OB基因核苷酸多样性(π)、单倍型数目(H)、单倍型多样性(Hd)、平均核苷酸差异数(K)、多态位点数(S)和基于分离位点数目的多样性(θw),发现遗传多样性水平具有临朐种群>沂南种群>曲阜种群的特点。三个地理种群间的种群间遗传分化系数(Fst)、基因流(Nm)、种群间的核苷酸歧异度(Dxy)等显示沂南、临朐和曲阜三个地理种群黑线仓鼠OB基因均具有一定遗传分化但不显著。在种群间,曲阜和临朐两个地区遗传分化最大,基因交流较小；曲阜和沂南地区次之；临朐和沂南地区遗传分化最小,基因交流较大。种群遗传分化系数(Gst)在0.00455～0.06523之间,说明有93%以上的变异来自于种群内部。实验中用MEGA4.0软件对三个地理种群黑线仓鼠个体做NJ系统进化分析,结果表明三个地理种群黑线仓鼠遗传分化不明显。本研究发现山东地区黑线仓鼠OB基因存在丰富的核苷酸多样性和单倍型多样性,沂南、临朐和曲阜三个地理种群间遗传分化程度很小。
本研究还测定了66个黑线仓鼠的体脂含量,并对其OB基因的序列进行分析,结果发现,在第三外显子编码区上发现三个同义突变C263T、A481G、A511G,OB基因编码区存在4种单倍型。经SPSS17.0分析,四种单倍型的黑线仓鼠的体脂含量均存在显著性差异(P<0.05),由于这些突变不能引起氨基酸产物序列的变化,基本不影响蛋白质活性,从而不会使动物表现出明显的性状变化,故认为此基因突变并不是性状变异的主要原因。不同单倍型黑线仓鼠体脂含量的显著性差异可能是由其他原因引起的。不同单倍型与脂肪含量之间的关系以及OB基因多态性对脂肪蓄积的基因效应还会通过扩大样本后作进一步研究。另外,关于OB基因与黑线仓鼠繁殖、免疫等功能在基因水平上的相关性也有待于进一步研究。
Cricetulus barabensis are widely distributed in China and have strong reproductive ability. The species have been considered as deleterious animals for agriculture and industry, which is a serious threat to people'normal production and life. In recent years, C. barabensis as the dominant rodent species in North of China has evoked more and more concerns among the academic circles, and many researches outcomes spring. In these experiments the author investigate from the viewpoint of molecular biology.
The obese gene (OB) mainly expressed in adipose tissue. Leptin is the expression product of obese gene which is a peptide hormone discovered recently. Leptin plays an important role in animal physiology function such as ingestion, energy metabolism, reproduction activity, fat storage, fetal growth, puberty development, tumour cell proliferation, hemopoiesis, neuroimmunoendocrine and so on. It is of the most important biological significance for the survival and reproduction.
In order to investigation the population hereditary constitution of C. barabensis, in this thesis, the partial sequence of obese gene was amplifid and sequenced.1665bp of OB of the species in Shandong Province was amplified by polymerase chain reaction for the first time. The primers were designed on the basis of the homologous species from GenBank consisting of mice, rat and human. The sequence of OB covers partial exon2, total intron2 and partial exon3. The sequence were then cloned and sequenced in 3 geographic populations in Shandong Province: Yinan, Linqu and Qufu. A total of 19 SNPs were examined, including lindels Insert/Deletion and the ratio of conversion to transversion was 15/4.34 haplotypes were defined in 103 samples, The sequence numbers submitted to the GenBank were:HM100043～HM100076. The sequence numbers submitted to the GenBank were～.The nucleotide diversity (π), number of haplotye (H), haplotype diversity (Hd), average number of nucleotide differences (K), number of polymorphic sites (S) and theta from S (θw) of OB of Yinan, Linqu and Qufu populations were calculated with DnaSPv5.0 software, which showed that the genetic diversity of the three populations arrayed as (from high to low):Linqu,Yinan, Qufu. In addition, Fst, Nm and Dxy proved the genetic differentiation among the three populations existed in a low level. The result of Gst analysis among three populations arrayed from 0.00455 to 0.06523, which meant that above 93% mutation existed in each population while few mutation existed among populations. NJ analysis showed that the genetic differentiation among the three populations was not distinct. The research showed that OB on C. barabensis from Qufu,Yinan and Linqu was rich in haplotype diversity and nucleotide diversity. The genetic differentiation among the three populations:Qufu,Yinan,Linqu of C. barabensis are small.
In the study, by analyzing the sequence of OB on C. barabensi with different body fat percent we found three cSNPs in coding section:C/T, A/G and A/G, all of them are samesense mutation. We analized the coding section of OB in 66 samples, then found 4 haplotypes. The data were analyzed statistically with SPSS17.0. The result shows that there is significant difference of body fat percent among 4 haplotypes. Because these mutations couldn't change the sequence of Amino Acids Production, the protein activity wasn't affected, and obvious trait variation didn't appeared on animals, so I think gene mutation was not the main reason for trait variation. Any other reasons may lead to the significant difference among body fat percent of different haplotype of C. barabensis. And in this study, the relationships between different haplotype and body fat percent, the genetic effect of OB on body fat deposition are needed to be further studied by enlarging the samples. Besides, concerning the correlation on gene level between OB and functions such as breeding and immunity of Striped Hamster, further research is also needed.
引文
[1]肖治术,张知彬,王玉山.啮齿动物鉴别虫蛀种子的能力及其对坚果植物更新的潜在影响[J].兽类学报,2003,23(4)：312～320.
[2]黄秀清,冯志勇,颜世祥,叶冠良,温文荣,王伟轩.珠江三角洲作物结构变动对害鼠种群动态的影响[J].广东农业科学,2002,(4)：36～39.
[3]张知彬,王祖望.农业重要害鼠的生态学及控制对策[M].北京：海洋出版社,1998：1～19,223～226.
[4]汪诚信.我国鼠害及其防治对策[J].中国媒介生物学及控制杂志,1996,7(1)：62～65.
[5]黄文几,陈延熹,温业新.中国啮齿类[M].上海：复旦大学出版社,1995：175～177.
[6]罗泽珣,陈卫,高武等.中国动物志·兽纲(第六卷)[M].北京：科学出版社,2000：28～38.
[7]汪泳庆,张知彬.大仓鼠种群遗传多样性的季节性变化[J].动物学报,2002,48(6)：733～740.
[8]吕国强,孟光,蒋化民.黑线仓鼠发生与防治研究初报[J].植物保护,1991,17(4)：37～38.
[9]穆成旺,任维新.黑线仓鼠的生物学特性及防治研究[J].甘肃农业科技,1999,(1)：39.
[10]王淑卿,杨荷芳,郝守身,张知彬.黑线仓鼠的食物与食量[J].动物学报,1992,38(2)：155～164.
[11]卢浩泉,李玉春,张学栋.黑线仓鼠种群年龄组成及其数量季节消长的研究[J].兽类学报,1987,7(1)：28～34.
[12]张洁.北京地区黑线仓鼠年龄鉴定及种群年龄组成的研究[J].兽类学报,1985,5(2)：141～150.
[13]李玉春,卢浩泉,张学栋,徐文生.黑线仓鼠的生长指标分析与年龄指标确定[J].兽类学报,1989,9(1)：49～55.
[14]董维惠,侯希贤,林小泉,杨玉平.黑线仓鼠种群数量动态预测研究[J].生态学报,1993,13(4)：300～306.
[15]董维惠,侯希贤,杨玉平.鄂尔多斯沙地草场黑线仓鼠种群特征研究[J].中国媒介生物学及控制杂志,2003,14(2)：8～11.
[16]姜运良,卢浩泉,李玉春,王玉山,张学栋,徐文生,于之庆.山东阳谷县黑线仓鼠种群数量预测预报[J].兽类学报,1994,14(3)：195～201.
[17]武文华,付和平,武晓东,杨玉平,董维惠,徐胜利.应用马尔可夫链模型预测长爪沙鼠和黑线仓鼠种群数量[J].动物学杂志,2007,42(6)：69～78.
[18]武文华,付和平,武晓东,董维惠,徐胜利.应用时间序列分析法预测黑线仓鼠和长爪沙鼠种群数量[J].内蒙古农业大学学报,2007,28(4)：6～11.
[19]杨玉平,周延林,张鹏利,侯希贤,董维惠.黑线仓鼠的生长发育的研究[J].兽类学报,1996,16(4)：309～311.
[20]张洁.北京大兴地区黑线仓鼠种群繁殖生态研究[J].兽类学报,1986,6(1)：45～56.
[21]鲍伟东,王德华,王祖望,周延林,王利民.内蒙古库布齐沙地和呼和浩特平原黑线仓鼠种群繁殖特征的比较[J].动物学杂志,2001,14(3)：15～18.
[22]李瑶,孙儒泳,王汶成,贾西西.长爪沙鼠和黑线仓鼠的温度选择比较[J].兽类学报,1988,8(1)：55～59.
[23]周延林,侯希贤,董维惠,杨玉平.黑线仓鼠肥满度的研究[J].兽类学报,1992,12(3)：207～212.
[24]柳劲松,王德华,孙儒泳.东北地区黑线仓鼠的代谢产热特征及其体温调节[J].动物学报,2003,49(4)：451～457.
[25]熊文华,张知彬.饶阳地区三种农田啮齿类头骨形态比较及性二型[J].兽类学报,2007,27(3)：280～283.
[26]王俊森,范琳琳,李云芳,张安民,徐德,黄绍斌.黑线仓鼠的染色体组型研究[J].解剖遗传,1996,2：27～30.
[27]徐承水,杨幼凤,卢浩泉.黑线仓鼠不同季节睾丸组织学研究[J].曲阜师范大学学报,1992,18(2)：60～65.
[28]徐承水.曲阜孔林不同月份黑线仓鼠睾丸组织变化的比较研究[J].东北林业大学报,2000,28(5)：137～138.
[29]何凤琴,邰发道,张育辉,丁小丽.黑线仓鼠、棕色田鼠和大林姬鼠犁鼻系统的显微结构观察[J].陕西师范大学学报,2005,33(2)：87～90.
[30]吴瑾.大仓鼠,黑线仓鼠,黑线姬鼠及社鼠线粒体DNA控制区全序列SNP分析[D].曲阜师范大学硕士学位论文,2006.
[31]李传海.黑线仓鼠微卫星引物的筛选及应用[D].曲阜师范大学硕士学位论文,2006.
[32]高倩.黑线仓鼠SSR标记及地理种群遗传多态性研究[D].曲阜师范大学硕士学位论文,2008.
[33]解学辉,董晓波,秦桢,孔繁华,徐来祥.黑线仓鼠MHCⅡ类DQA基因外显子2的克隆与序列分析.曲阜师范大学学报,2009,35(2)：98～103.
[34]Ingalls A, Dickie M,Snell G. Obese-a new mutation in the house mouse[J].Journal of Heredity,1950,41:317-318.
[35]Friedman J M, Leibeil R L. Tackling a weighty problem[J].Cell,1992,69:217-220.
[36]Zhang Y, Proenca R, Maffei M, Barone M,Leopold L, Friedman J M. Positional cloning of the mouse obese gene and its human homologue[J].Nature,1994,372:425-443.
[37]He Y,Chen H,Quon M,Reitman M. The mouse obese gene[J].Journal of Biological Chemistry,1996,270:28887～288891.
[38]Murakami T,Shima K. Cloning of rat obese cDNA and its expression in obese rats[J]. Biochemncal and Biophysical Research Communication,1995,209:94～95.
[39]Rock F L, Altmann S W, Heek van M, Kastelein R A, Bazan J F.The leptin haemopoietic cytoline fold is stabilized by intrachain disfufide bond.Horm[J]. Metabolic Research,1996,28: 628-652.
[40]Kline A D,Becker G W,Churgay L M,Landen B E,Martin D K,Muth W L, Rathnachalam R,Richardson J M,Schoner B,Ulmer M, Hale J E. Leptin is a four-helix bundle:secondary structure by NMR[J].FEBS letters,1997,407:239-242.
[41]Made JT,Boguski MS and Eisenberg D. Threading analysis suggests that the obese gene product may be a helical cytokine[J].FEBS letters,1995,373:13-18.
[42]Isse N, Ogawa Y, Tamura N, Masuzaki N, Mori K, Okazaki T, Satoh N, Shigemoto M, Yoshimasa Y, Nishi S, Hosoda K, Inazawa J, Nakao K Structural Organization and Chromosomal Assignment of the Human obese Gene[J].The Journal of Biological Chemistry, 1995,270(46):27728-27733.
[43]Gong D W,Bi S,Pratley R E,Weintraub B D. Genomic Structure and Promoter Analysis of the Human obese Gene[J]. The Journal of Biological Chemistry,1996,271(8):3971-3974.
[44]Neuenschwander S, Rettenberger G, Meiherink E, Jorg H, Stranzinger G. Partial characterization of porcine obesity gene(OBS)and its localization to chromosome 18 by somatic cell hybrids[J].Animal Genetics,1996,27(4):275-278.
[45]Ramsay T G,Yan X,Morrison C. The obestiy gene in swine; sequecand expression of porcine leptin[J].Journal of Animal Science,1998,76(2):484-490.
[46]戴茹娟,李宁,吴常信.猪Ob基因cDNA的克隆与分析[J].遗传学报,2000,27(4)：290～297.
[47]吴彦宁.猪分离群体中五个功能基因多态性及其生长性能的相关.[D].南京：南京农业大学,1999.
[48]掌子凯.猪ob基因不同基因型与胴体性状关系的研究.[D].南京：南京农业大学,2000.
[49]许玲玲,李宁,张守发,赵兴波,冯继东,连正兴,吴常信.猪肥胖基因5’调控区与内含子1的测序分析及其两个新微卫星的发现[J].遗传学报,2005,32(4)：360～365.
[50]Taouis M, Chen J-W, Davi and C, Dupont J, Derouet M, Simon J. Cloning the chicken leptin gene[J].Gene,1998,20:239-242.
[51]Ashwell C M, Murtry J P, Wang X H, Zhou Y, Vasilatos-Younken R. Effects of growth hormone and pair-feeding on leptin mRNA expression in liver and adipose tissue[J].Domestic animal endocrinology,1999,16(1):66-84.
[52]Kurokawa T,Susumu U,Suzukil T. Identication of cDNA coding for a homologue to mammalian leptin from puff erfish,Takifugu rubripes[J].Peptides,2005,26(5):745～750.
[53]戴汉川.几种鱼类肥胖基因的克隆与鲤肥胖基因表达的研究[D].武汉：华中农业大学,2005.
[54]C. J.Dyer, J.M.Simmons,R. L. Matteri,D.H.Keisler. cDNA cloning and tissue-specific gene expression of ovine leptin,NPY-Y1 receptor,and NPY-Y2 receptor[J].Domestic Animal Endocrinology,1997,14(5):295-303.
[55]Ji S,Willis G M,Scott R R,Spurlock M E. Partial cloning and expression of the bovine leptin gene[J].Animal Biotechnology,1998,9(1):1-14.
[56]Ogawa Y, Masuzaki H, Isse N, Okazaki T, Mori T, Shigemoto T, Satoh N, Tamura N, Hosoda K,Yoshimasa Y. Molecular clonging of rat obese cDNA and augmented gene expression in genetically obese Zucker fatty(fa/fa)rats[J].The Journal of Clinical Investigation,1995,96(3): 1647-1652.
[57]Montague C T, Farooqi I S, Whitehead J P, Soos M A, Rau H, Wareharn N J, Sewter C P, Digby J E, Mohamed S N, Hurst J A,Cheatham K H, Earley A R, Barnett A H, Prins J B, O'Rahilly S. Congenital leptin deficiency is associated with severe early-onset obesity in humans[J].Nature,1996,386:903-908.
[58]Masuzaki H, Ogawa Y, Isse N, Satoh N, Okazaki T, Shigemoto M, Mori K, Tamura N,Hosoda K,Yoshimasa Y. Human obese gene expression adipocyte-specific expression and regional differences in the adipocyte tissue[J]. Diabetes,1995,44(7):855-858.
[59]Mohammed Taouis,Jin-Wen Chen,Christian Daviaud,Joelle Dupont,Michel Derouet, Jean Simon. Cloning the chicken leptin gene[J].Gene,208 (2):239-242.
[60]潘庭双,龙良启,吴小平,罗晓松,邹世平.蒙古红鲌肥胖基因cDNA的克隆与组织表达特异性研究[J].水生生物学报,2007,31(1)：94～98.
[61]Muruzabal F J,Fruhbeck G,Gornez-Ambrosi J,Archanco M,Burrell M A. Immunocytochemical detection of leptin in non-mammalian vertebrate stomach[J].General and Comparative Endocrinology,2002,128(2):149-152.
[62]Fruhbeck G,Aguado M,Martinez J A. In vitrolipolytic effect of leptin on mouse adipocytes-evidence for a possible autocrine/paracrine role of leptin[J].Biochemncal and Biophysical Research Communication,1997,240(3):590-594.
[63]Udagawa J,Hatta T,Naora H,Otani H. Expression of the long form of leptin receptor (Ob-Rb)mRNA in the brain of mouse embryos and newborn mice[J].Brain Research,2000, 868(2):251-258.
[64]Friedman J M. The alphabet of weight control[J]. Nature,1996,385:119-120.
[65]Erikson J,Hollopeter G,Palmiter R. Attenuation of the obesity syndrome of ob/ob mice by the loss neuropeptide Y[J].Science,1996,274:1704～1707.
[66]Mercer J G,Moar K M,Hoggard N. Localization of leptin receptor (ob-r) messenger ribonucleic acid in the rodent hindbrain[J].Endocrinology,1998,139(1):29～34.
[67]Cone R D, Lu D, Koppula S, Vage D I, Klungland H, Boston B, Chen W, Orth D N, Pouton C,Kesterson R A. The melanocortin receptors:agonists,antagonists,and the hormonal control of pigmentation[J].Recent Progress in Hormon Research,1996,51:287-317.
[68]Fan W,Boston B A,Kesterson R A,Hruby V J,Cone R D. Role of melanocortinergic neurons in feeding and the agouti obesity syndrome[J].Nature,1996,385:165-168.
[69]Huszar D,Lynch C,Fairchild Huntress V,Dunmore J H Fang Q,Berkemeier L R, Gu W, Kesterson R A, Boston B A, Cone R D, Smith F J, Campfield LA, Burn P, Lee F.Targete druption of the melanocortin-4 receptor result in obesity in mice[J].Cell,1997,88:131-141.
[70]Moussa N M,Claycombe K J. The yellow mouse obesity syndrome and mechanisma of agouti-induced obesity[J].Obesity Research,1999,7(5):506-514.
[71]Halaas J L, Gajiwala K S, Maffei M, Cohen S L, Chait B T, Rabinowitz D,Lallone R L, Burley S K, Friedman J M. Weight-reducing effects of the plasma protein encoded by the obese gene[J].Science,1995,269:543-545.
[72]黄之瑜,孙志娟,范少光.肥胖与神经调节[J].生理科学进展,2001,32(1)：45～51.
[73]Bennett B D,Solar G P,Yuan J Q,Mathias J,Thomas G R,Matthews W. A role for leptin and its cognate receptor in hematopoiesis [J]. Current Biology,1996,6(9):1170-1180.
[74]Howard J K, Lord G M, Matarese G, Vendetti S, Ghatei M A, Ritter M A, Lechler R I, Bloom S R. Leptin protects mice from starvation-induced lymphoid atrophy and increases thymic cellularity in ob/ob mice[J]. The Journal of Clinical Investigation,1999,104 (8): 1051-1059.
[75]Santos-Alvarez J, Goberna R, Sanchez-Margalet V. Human leptin stimulates proliferati on and activation of human circulating monocytes[J] Cellular Immunology,1999,194 (1):6-11.
[76]Zarkesh-Esfahani H, Pockley A G, Wu Z, Hellewell P G, Weetman A P, Ross R M. Leptin indirectly activates human neutrophils via induction of TNF-alpha[J]. The Journal of Immunology,2004,172(3):1809-1814.
[77]Zhao Y, Sun R,You L, Gao C,Tian Z. Expression of leptin receptors and response to leptin stimulation of human natural killer cell lines [J].Biochemncal and Biophysical Research Communication,2003,300(2):247-252.
[78]Tian Z,Sun R,Wei H,Gao B. Impaired natural killer (NK) cell activity in leptin receptor deficient mice:leptin as a critical regulat or in NK cell development and activation[J]. Biochemncal and Biophysical Research Communication,2002,298(3):297-302.
[79]Martin-Romero C,Santos-Alvarez J, Goberna R,Sanchez-Margalet V. Human leptin enhances activation and proliferation of human circulating T Lymphocytes[J].Cellular Immunology,2000,199(1):15-24.
[80]Howard J K, Lord G M, Matarese G, Vendetti S, Ghatei M A, Ritter M A, Lechler R I, Bloom S R. Leptin protects mice from starvation-induced lymphoid atrophy and increases thymic cellularity in ob/ob mice[J].The Journal of Clinical Investigation,1999,104(8): 1051-1059.
[81]Zarkesh E, Pockley G, Metcalfe R A, Bidlingmaier M, Zida W, Ajami A, Weetman A P,Strasburger C J,Richard J M. High-dose leptin activates human leukocytes via receptor expression on monocytes[J].The Journal of Immunology,2001,167(8):4593-4599.
[82]Green B B,Weiss N S, Daliug J F. Risk of ovulatory infertility in relation to body weight[J]. Fertility and Sterility,1988,50(5):721-726.
[83]Souza D,Metzger D A. Reproductive dysfunction in amenorrheic athletes and anorexic patients:a review[J].Medicine & Science in Sports & Exercise,1991,23(9):995-1006.
[84]Pelleymounter M A, Cullen M J, Baker M B, Hecht R, Winters D, Boone T, Collies F. Effects of the obese gene product on body weight regulation in ob/ob mice[J]. Science,1995, 269:540-543.
[85]Zheng D H,Wooter M H,Zhou Q,Dohm G L. The effect of exercise on ob gene expression[J].Biochemncal and Biophysical Research Communication,1996,225(3):646-650.
[86]Klein S,Coppack S W, Mohammed-Ail V,Landt M. Adipose tissue leptin production and plasma leptin kinerics in hamans[J].Diabetes,1996,45:984-986.
[87]Ahima R S,Prabacaran D,Mantzoros C,Qu D,Lowell B,Maratos-flier E,Flier J S. Roles of leptin in the neuroendocrine response to fasting[J].Nature,1996,382:250-252.
[88]Yu W H, Kimura M, Walczewska A Karanth S,McCann S M. Role of leptin in hypothalamic-pituitary function[J].Proceeding of the National Academy of Sciences of the United States of America,1996,94:1023-1028.
[89]Mizuno T M,Kleopoulos S P,Bergen H T,Roberts J L, Priest C A, Mobbs C V. Hypothalamic pro-opiomelanocortin mRNA is reduced by fasting and in ob/ob and db/db mice, but is stimulated by leptin [J].Diabetes,1998,47(2):294-297.
[90]孙长颖,李颖,王旭,马瑞.leptin在男性青春期发育中的作用[J].中华预防医学杂志,2004a,38(4)：331-333.
[91]孙长颖,李颖,王舒然,闻颖.leptin在女性青春期发育中的作用及其与胰岛素、生长激素和性激素的关系[J].中国临床康复,2004b,8(24)：5169～5117.
[92]陈赛,李尚为,唐海峰,张金虎,尹海林.leptin对小鼠着床前胚胎发育影响的体外研 究[J].四川大学学报(医学版),2004,35(6)：806～808.
[93]贲晓明,秦玉明,喻文亮,张伟利,蔡威,吴圣楣.胎盘肥胖基因表达与胎儿宫内生长发育相关性研究[J].中华围产医学杂志,2002,5(1)：60～63.
[94]荣飞,林季,颜光涛.瘦素与损伤修复[J].医学综述,2008,14(11)：1605～1067.
[95]Schafer K, Halle M, Goeschen C, Dellas C, Pynn M,J.Loskutoff D,Konstantinides S. Leptin promotes vascular remodeling and neointimal growth in mice [J].Arteriosclerosis, Thrombosis, and Vascular Biology,2004,24(1):112-117.
[96]Burcelin R,Kamohara S,Li J,Tannenbaum G S,Charron M J,Friedman J M. Acute intravenous leptin infusion increases glucose turnover but not skeletal muscle glucose uptake in ob/ob mice[J].Diabetes,1999,48(6):1264-1269.
[97]Chen H C,Farese R V. Inhibition of triglyceride synthesis as a treatment strategy for obesity:lessons from DGAT1-deficient mice[J],Arteriosclerosis, Thrombosis, and Vascular Biology,2005,25(3):482-486.
[98]肖德明,徐忠世,庚智文,卢小虎,胡新佳.血清leptin与骨质疏松关系的研究[J].中国骨肿瘤骨病,2004,3(6)：352～354.
[99]朱玉贤,李毅.现代分子生物学[M].第二版.北京：高等教育出版社,2002：371～376.
[100]William J C K, Kubelik A R,Livak K J. DNA polymorphism amplified by arbitrary primers are useful genetic markers[J].Nucleic Acids Research,1990,18(22):6531-6537.
[101]Velappam N,Sondgrass J L, Hakovirta J R. Rapid identification of pathogenic bacteria by single-enzyme amplified fragment length polymorphism analysis[J].Diagnstic Microbiology and infections Disease,2001,39(2):77-83
[102]王志林,赵树进,吴新荣.分子标记技术及其进展[J].生命的化学,2001,21(4)：39～42.
[103]黎裕,贾继增,王天宇.分子标记的种类及其发展[J].生物技术通报,1999,15(4)：19～22.
[104]Serapion J,Kucudtas H,Feng J,Liu Z. Bioinformatic mining of type I mincrosatellite from expressed sequence tags of channel catfish(Ictalurus punctatus) [J].Animal Genetics,2003,34 (6):445.
[105]张丽娟,张保军,耿社民.微卫星标记与动物遗传育种[J].黄牛杂志,2003,29(2)：49.
[106]冯龙仁,牛安欧.微卫星简单重复序列锚定PCR扩增技术(SSR-PCR)及其在寄生虫基因研究中的应用[J].实用寄生虫杂志,2000,8(1)：33～34.
[107]杨昭庆,洪坤学,褚嘉佑.单核苷酸多态性的研究进展[J].国外医学：遗传学分册,2000,23(1)：4～8.
[108]张效萌,韩金祥.单核苷酸多态性的检测及其在医药领域的应用[J].生命的化学,2001,21(4)：325～327.
[109]刘炬,徐兵河.单核苷酸多态性与乳腺癌患者的预后[J].癌症进展,2007,5(3)：242～248.
[110]刘越,吕社民.单核苷酸多态性影响基因功能的机制[J].生命的化学,2008,28(4)：214～216.
[111]唐棣,王志民.SNPs检测方法研究进展[J].上海交通大学学报(农业科学版),2007,25(4)：405～418.
[112]李艳杰,龙火生,李影,杨公社.单核苷酸多态性(SNP)的研究进展和应用[J].Animal Science Veteringany Medicine,2003:22(4):16-18.
[113]萨姆布鲁克J,弗里奇E F,曼尼阿蒂斯T.分子克隆实验指南[M].金冬雁,黎孟枫等译.第二版,北京：科学出版社,1992.
[114]Masuzaki H,Ogawa Y,Hosoda K, Kawada T,Fushiki T,Nakao K. Augmented expression of the obese gene in the adipose tissue from rat fed high-fat diet[J]. Biochemncal and Biophysical Research Communication,1995,216(1):355-358.
[115]Maffei M, Halaas J, Ravussin E, Pratley R E, Lee G H, Zhang Y, Fei H, Kim S, Lallone R, Ranganathan S, Kern P A, Friedman J M. Leptin levels in human and rodent:measurement of plasma Leptin and ob RNA in obese and weight-reduced subject[J]. Nature Medicine,1995,1:1155-1161.
[116]杨金权,胡雪莲,唐文乔,林弘都.长江口邻近水域刀鲚的线粒体控制区序列变异与遗传多样性[J].动物学杂志,2008,43(1)：8～15.
[117]Nei M. Molecular population genetics and evolution. North-Holland Publishing company, Amsterdam. Oxford American Elsevier publishing company, Inc. New York,1975.
[118]King J L. The role of mutation in evolution. Proc.6th Berkeley Symp. Statist, and Probab. 1972,69-100.
[119]Allendorf F W. Isolation,Gene flow and genetic differentiation among populations[J]. Genetics and Conservation,1983,18(3):51-65.
[120]Reid W V,Kenton R M. Keeping options alive:the scientific basis for conserving biodiversity[M].World Resouces Institute,Washington D C,1989.
[121]Castela V,Ruedi M,Excofifer L, Inanez C, Arlettaz R,Hausser J. Is the Gibraltar Strait a barrier to gene flow for the bat Myotis myotis(Chiroptera:Vespertilionidae)[J].Molecular Ecology,2000,9(11):1761-1772.
[122]Keller I,Nentwig W, Iargiader C R. Recent hahitat fragmentation due to roads can lead to significant genetic differentiation in an abundant flightless ground beetle Molecular Ecology, 2004,9:1761～1772.
[123]Su H,Qu L J,He K, Zhang Z,Wang J,Chen Z,Gu H. The Great wall of China:a physical barrier to gene flow? [J].Heredity,2003,90(3):212～219.
[124]Beh W,Seitz A. The influence of land use on the genetic structure of populations of the common frog Rana temporaria[J].Biology Conservation,1990,54(3):239-249.
[125]Kelley S T,Farrell B D,Mitton J B. Effects of specialization on genetic differentiation in sister species of bark beetles[J].Heredity,2000,84(14):219-227.
[126]Whittaker R H,Levin S A,Root R B.Niche,habitat,and ecotope[J].American.Nature, 1973,107:321-338.
[127]Reid W V. Kenton R M.Keeping options alive:the scientific basis for conserving biodiversity[M].World Resouces Institute,Winshingtn D C,1989.
[128]Jiang Z H,Gison J P.Genetic polymorphisms in the leptin gene and their association with fatness in four pig breeds[J].Mammalian Genome,1999,10(2):191～193.
[129]李毅.松辽黑猪ob基因单核苷酸多态性及其部分经济性状的相关分析[D].吉林：吉林大学,2005.
[130]Houseknecht K L, Portocarrero C P. Leptin and its receptors:regulators of whole body energy homeostasis[J].Domestic Animal Endocrinology,1998,15(6):457-475.
[131]杜清友,王会信.胰岛素样生长因子结合蛋白研究进展[J].生物化学与生物物理进展,1998,25(2)：104～105.
[132]Stephens T W, Basinski M, Bristow P K, Buevalleskey J M, Burgett S G,Craft L, Halb J, Hoffmann J, Hsiung H M, Kriauciuna A, Mackellar W, RosteckJr P R, Schoner B, Smith D,Tinsley F C,Zhang X,Heiman M. The role of neuropeptide Y in the antiobesity action of the obese gene product [J].Nature,1995,12,377(6549):530-532.
[133]刘桂琼,姜勋平,丁家桐.梅山猪ob基因多态性与繁殖性状的相关性研究[J].中国畜牧杂志,2002,38(4)：15～16.
[2]黄秀清,冯志勇,颜世祥,叶冠良,温文荣,王伟轩.珠江三角洲作物结构变动对害鼠种群动态的影响[J].广东农业科学,2002,(4)：36～39.
[3]张知彬,王祖望.农业重要害鼠的生态学及控制对策[M].北京：海洋出版社,1998：1～19,223～226.
[4]汪诚信.我国鼠害及其防治对策[J].中国媒介生物学及控制杂志,1996,7(1)：62～65.
[5]黄文几,陈延熹,温业新.中国啮齿类[M].上海：复旦大学出版社,1995：175～177.
[6]罗泽珣,陈卫,高武等.中国动物志·兽纲(第六卷)[M].北京：科学出版社,2000：28～38.
[7]汪泳庆,张知彬.大仓鼠种群遗传多样性的季节性变化[J].动物学报,2002,48(6)：733～740.
[8]吕国强,孟光,蒋化民.黑线仓鼠发生与防治研究初报[J].植物保护,1991,17(4)：37～38.
[9]穆成旺,任维新.黑线仓鼠的生物学特性及防治研究[J].甘肃农业科技,1999,(1)：39.
[10]王淑卿,杨荷芳,郝守身,张知彬.黑线仓鼠的食物与食量[J].动物学报,1992,38(2)：155～164.
[11]卢浩泉,李玉春,张学栋.黑线仓鼠种群年龄组成及其数量季节消长的研究[J].兽类学报,1987,7(1)：28～34.
[12]张洁.北京地区黑线仓鼠年龄鉴定及种群年龄组成的研究[J].兽类学报,1985,5(2)：141～150.
[13]李玉春,卢浩泉,张学栋,徐文生.黑线仓鼠的生长指标分析与年龄指标确定[J].兽类学报,1989,9(1)：49～55.
[14]董维惠,侯希贤,林小泉,杨玉平.黑线仓鼠种群数量动态预测研究[J].生态学报,1993,13(4)：300～306.
[15]董维惠,侯希贤,杨玉平.鄂尔多斯沙地草场黑线仓鼠种群特征研究[J].中国媒介生物学及控制杂志,2003,14(2)：8～11.
[16]姜运良,卢浩泉,李玉春,王玉山,张学栋,徐文生,于之庆.山东阳谷县黑线仓鼠种群数量预测预报[J].兽类学报,1994,14(3)：195～201.
[17]武文华,付和平,武晓东,杨玉平,董维惠,徐胜利.应用马尔可夫链模型预测长爪沙鼠和黑线仓鼠种群数量[J].动物学杂志,2007,42(6)：69～78.
[18]武文华,付和平,武晓东,董维惠,徐胜利.应用时间序列分析法预测黑线仓鼠和长爪沙鼠种群数量[J].内蒙古农业大学学报,2007,28(4)：6～11.
[19]杨玉平,周延林,张鹏利,侯希贤,董维惠.黑线仓鼠的生长发育的研究[J].兽类学报,1996,16(4)：309～311.
[20]张洁.北京大兴地区黑线仓鼠种群繁殖生态研究[J].兽类学报,1986,6(1)：45～56.
[21]鲍伟东,王德华,王祖望,周延林,王利民.内蒙古库布齐沙地和呼和浩特平原黑线仓鼠种群繁殖特征的比较[J].动物学杂志,2001,14(3)：15～18.
[22]李瑶,孙儒泳,王汶成,贾西西.长爪沙鼠和黑线仓鼠的温度选择比较[J].兽类学报,1988,8(1)：55～59.
[23]周延林,侯希贤,董维惠,杨玉平.黑线仓鼠肥满度的研究[J].兽类学报,1992,12(3)：207～212.
[24]柳劲松,王德华,孙儒泳.东北地区黑线仓鼠的代谢产热特征及其体温调节[J].动物学报,2003,49(4)：451～457.
[25]熊文华,张知彬.饶阳地区三种农田啮齿类头骨形态比较及性二型[J].兽类学报,2007,27(3)：280～283.
[26]王俊森,范琳琳,李云芳,张安民,徐德,黄绍斌.黑线仓鼠的染色体组型研究[J].解剖遗传,1996,2：27～30.
[27]徐承水,杨幼凤,卢浩泉.黑线仓鼠不同季节睾丸组织学研究[J].曲阜师范大学学报,1992,18(2)：60～65.
[28]徐承水.曲阜孔林不同月份黑线仓鼠睾丸组织变化的比较研究[J].东北林业大学报,2000,28(5)：137～138.
[29]何凤琴,邰发道,张育辉,丁小丽.黑线仓鼠、棕色田鼠和大林姬鼠犁鼻系统的显微结构观察[J].陕西师范大学学报,2005,33(2)：87～90.
[30]吴瑾.大仓鼠,黑线仓鼠,黑线姬鼠及社鼠线粒体DNA控制区全序列SNP分析[D].曲阜师范大学硕士学位论文,2006.
[31]李传海.黑线仓鼠微卫星引物的筛选及应用[D].曲阜师范大学硕士学位论文,2006.
[32]高倩.黑线仓鼠SSR标记及地理种群遗传多态性研究[D].曲阜师范大学硕士学位论文,2008.
[33]解学辉,董晓波,秦桢,孔繁华,徐来祥.黑线仓鼠MHCⅡ类DQA基因外显子2的克隆与序列分析.曲阜师范大学学报,2009,35(2)：98～103.
[34]Ingalls A, Dickie M,Snell G. Obese-a new mutation in the house mouse[J].Journal of Heredity,1950,41:317-318.
[35]Friedman J M, Leibeil R L. Tackling a weighty problem[J].Cell,1992,69:217-220.
[36]Zhang Y, Proenca R, Maffei M, Barone M,Leopold L, Friedman J M. Positional cloning of the mouse obese gene and its human homologue[J].Nature,1994,372:425-443.
[37]He Y,Chen H,Quon M,Reitman M. The mouse obese gene[J].Journal of Biological Chemistry,1996,270:28887～288891.
[38]Murakami T,Shima K. Cloning of rat obese cDNA and its expression in obese rats[J]. Biochemncal and Biophysical Research Communication,1995,209:94～95.
[39]Rock F L, Altmann S W, Heek van M, Kastelein R A, Bazan J F.The leptin haemopoietic cytoline fold is stabilized by intrachain disfufide bond.Horm[J]. Metabolic Research,1996,28: 628-652.
[40]Kline A D,Becker G W,Churgay L M,Landen B E,Martin D K,Muth W L, Rathnachalam R,Richardson J M,Schoner B,Ulmer M, Hale J E. Leptin is a four-helix bundle:secondary structure by NMR[J].FEBS letters,1997,407:239-242.
[41]Made JT,Boguski MS and Eisenberg D. Threading analysis suggests that the obese gene product may be a helical cytokine[J].FEBS letters,1995,373:13-18.
[42]Isse N, Ogawa Y, Tamura N, Masuzaki N, Mori K, Okazaki T, Satoh N, Shigemoto M, Yoshimasa Y, Nishi S, Hosoda K, Inazawa J, Nakao K Structural Organization and Chromosomal Assignment of the Human obese Gene[J].The Journal of Biological Chemistry, 1995,270(46):27728-27733.
[43]Gong D W,Bi S,Pratley R E,Weintraub B D. Genomic Structure and Promoter Analysis of the Human obese Gene[J]. The Journal of Biological Chemistry,1996,271(8):3971-3974.
[44]Neuenschwander S, Rettenberger G, Meiherink E, Jorg H, Stranzinger G. Partial characterization of porcine obesity gene(OBS)and its localization to chromosome 18 by somatic cell hybrids[J].Animal Genetics,1996,27(4):275-278.
[45]Ramsay T G,Yan X,Morrison C. The obestiy gene in swine; sequecand expression of porcine leptin[J].Journal of Animal Science,1998,76(2):484-490.
[46]戴茹娟,李宁,吴常信.猪Ob基因cDNA的克隆与分析[J].遗传学报,2000,27(4)：290～297.
[47]吴彦宁.猪分离群体中五个功能基因多态性及其生长性能的相关.[D].南京：南京农业大学,1999.
[48]掌子凯.猪ob基因不同基因型与胴体性状关系的研究.[D].南京：南京农业大学,2000.
[49]许玲玲,李宁,张守发,赵兴波,冯继东,连正兴,吴常信.猪肥胖基因5’调控区与内含子1的测序分析及其两个新微卫星的发现[J].遗传学报,2005,32(4)：360～365.
[50]Taouis M, Chen J-W, Davi and C, Dupont J, Derouet M, Simon J. Cloning the chicken leptin gene[J].Gene,1998,20:239-242.
[51]Ashwell C M, Murtry J P, Wang X H, Zhou Y, Vasilatos-Younken R. Effects of growth hormone and pair-feeding on leptin mRNA expression in liver and adipose tissue[J].Domestic animal endocrinology,1999,16(1):66-84.
[52]Kurokawa T,Susumu U,Suzukil T. Identication of cDNA coding for a homologue to mammalian leptin from puff erfish,Takifugu rubripes[J].Peptides,2005,26(5):745～750.
[53]戴汉川.几种鱼类肥胖基因的克隆与鲤肥胖基因表达的研究[D].武汉：华中农业大学,2005.
[54]C. J.Dyer, J.M.Simmons,R. L. Matteri,D.H.Keisler. cDNA cloning and tissue-specific gene expression of ovine leptin,NPY-Y1 receptor,and NPY-Y2 receptor[J].Domestic Animal Endocrinology,1997,14(5):295-303.
[55]Ji S,Willis G M,Scott R R,Spurlock M E. Partial cloning and expression of the bovine leptin gene[J].Animal Biotechnology,1998,9(1):1-14.
[56]Ogawa Y, Masuzaki H, Isse N, Okazaki T, Mori T, Shigemoto T, Satoh N, Tamura N, Hosoda K,Yoshimasa Y. Molecular clonging of rat obese cDNA and augmented gene expression in genetically obese Zucker fatty(fa/fa)rats[J].The Journal of Clinical Investigation,1995,96(3): 1647-1652.
[57]Montague C T, Farooqi I S, Whitehead J P, Soos M A, Rau H, Wareharn N J, Sewter C P, Digby J E, Mohamed S N, Hurst J A,Cheatham K H, Earley A R, Barnett A H, Prins J B, O'Rahilly S. Congenital leptin deficiency is associated with severe early-onset obesity in humans[J].Nature,1996,386:903-908.
[58]Masuzaki H, Ogawa Y, Isse N, Satoh N, Okazaki T, Shigemoto M, Mori K, Tamura N,Hosoda K,Yoshimasa Y. Human obese gene expression adipocyte-specific expression and regional differences in the adipocyte tissue[J]. Diabetes,1995,44(7):855-858.
[59]Mohammed Taouis,Jin-Wen Chen,Christian Daviaud,Joelle Dupont,Michel Derouet, Jean Simon. Cloning the chicken leptin gene[J].Gene,208 (2):239-242.
[60]潘庭双,龙良启,吴小平,罗晓松,邹世平.蒙古红鲌肥胖基因cDNA的克隆与组织表达特异性研究[J].水生生物学报,2007,31(1)：94～98.
[61]Muruzabal F J,Fruhbeck G,Gornez-Ambrosi J,Archanco M,Burrell M A. Immunocytochemical detection of leptin in non-mammalian vertebrate stomach[J].General and Comparative Endocrinology,2002,128(2):149-152.
[62]Fruhbeck G,Aguado M,Martinez J A. In vitrolipolytic effect of leptin on mouse adipocytes-evidence for a possible autocrine/paracrine role of leptin[J].Biochemncal and Biophysical Research Communication,1997,240(3):590-594.
[63]Udagawa J,Hatta T,Naora H,Otani H. Expression of the long form of leptin receptor (Ob-Rb)mRNA in the brain of mouse embryos and newborn mice[J].Brain Research,2000, 868(2):251-258.
[64]Friedman J M. The alphabet of weight control[J]. Nature,1996,385:119-120.
[65]Erikson J,Hollopeter G,Palmiter R. Attenuation of the obesity syndrome of ob/ob mice by the loss neuropeptide Y[J].Science,1996,274:1704～1707.
[66]Mercer J G,Moar K M,Hoggard N. Localization of leptin receptor (ob-r) messenger ribonucleic acid in the rodent hindbrain[J].Endocrinology,1998,139(1):29～34.
[67]Cone R D, Lu D, Koppula S, Vage D I, Klungland H, Boston B, Chen W, Orth D N, Pouton C,Kesterson R A. The melanocortin receptors:agonists,antagonists,and the hormonal control of pigmentation[J].Recent Progress in Hormon Research,1996,51:287-317.
[68]Fan W,Boston B A,Kesterson R A,Hruby V J,Cone R D. Role of melanocortinergic neurons in feeding and the agouti obesity syndrome[J].Nature,1996,385:165-168.
[69]Huszar D,Lynch C,Fairchild Huntress V,Dunmore J H Fang Q,Berkemeier L R, Gu W, Kesterson R A, Boston B A, Cone R D, Smith F J, Campfield LA, Burn P, Lee F.Targete druption of the melanocortin-4 receptor result in obesity in mice[J].Cell,1997,88:131-141.
[70]Moussa N M,Claycombe K J. The yellow mouse obesity syndrome and mechanisma of agouti-induced obesity[J].Obesity Research,1999,7(5):506-514.
[71]Halaas J L, Gajiwala K S, Maffei M, Cohen S L, Chait B T, Rabinowitz D,Lallone R L, Burley S K, Friedman J M. Weight-reducing effects of the plasma protein encoded by the obese gene[J].Science,1995,269:543-545.
[72]黄之瑜,孙志娟,范少光.肥胖与神经调节[J].生理科学进展,2001,32(1)：45～51.
[73]Bennett B D,Solar G P,Yuan J Q,Mathias J,Thomas G R,Matthews W. A role for leptin and its cognate receptor in hematopoiesis [J]. Current Biology,1996,6(9):1170-1180.
[74]Howard J K, Lord G M, Matarese G, Vendetti S, Ghatei M A, Ritter M A, Lechler R I, Bloom S R. Leptin protects mice from starvation-induced lymphoid atrophy and increases thymic cellularity in ob/ob mice[J]. The Journal of Clinical Investigation,1999,104 (8): 1051-1059.
[75]Santos-Alvarez J, Goberna R, Sanchez-Margalet V. Human leptin stimulates proliferati on and activation of human circulating monocytes[J] Cellular Immunology,1999,194 (1):6-11.
[76]Zarkesh-Esfahani H, Pockley A G, Wu Z, Hellewell P G, Weetman A P, Ross R M. Leptin indirectly activates human neutrophils via induction of TNF-alpha[J]. The Journal of Immunology,2004,172(3):1809-1814.
[77]Zhao Y, Sun R,You L, Gao C,Tian Z. Expression of leptin receptors and response to leptin stimulation of human natural killer cell lines [J].Biochemncal and Biophysical Research Communication,2003,300(2):247-252.
[78]Tian Z,Sun R,Wei H,Gao B. Impaired natural killer (NK) cell activity in leptin receptor deficient mice:leptin as a critical regulat or in NK cell development and activation[J]. Biochemncal and Biophysical Research Communication,2002,298(3):297-302.
[79]Martin-Romero C,Santos-Alvarez J, Goberna R,Sanchez-Margalet V. Human leptin enhances activation and proliferation of human circulating T Lymphocytes[J].Cellular Immunology,2000,199(1):15-24.
[80]Howard J K, Lord G M, Matarese G, Vendetti S, Ghatei M A, Ritter M A, Lechler R I, Bloom S R. Leptin protects mice from starvation-induced lymphoid atrophy and increases thymic cellularity in ob/ob mice[J].The Journal of Clinical Investigation,1999,104(8): 1051-1059.
[81]Zarkesh E, Pockley G, Metcalfe R A, Bidlingmaier M, Zida W, Ajami A, Weetman A P,Strasburger C J,Richard J M. High-dose leptin activates human leukocytes via receptor expression on monocytes[J].The Journal of Immunology,2001,167(8):4593-4599.
[82]Green B B,Weiss N S, Daliug J F. Risk of ovulatory infertility in relation to body weight[J]. Fertility and Sterility,1988,50(5):721-726.
[83]Souza D,Metzger D A. Reproductive dysfunction in amenorrheic athletes and anorexic patients:a review[J].Medicine & Science in Sports & Exercise,1991,23(9):995-1006.
[84]Pelleymounter M A, Cullen M J, Baker M B, Hecht R, Winters D, Boone T, Collies F. Effects of the obese gene product on body weight regulation in ob/ob mice[J]. Science,1995, 269:540-543.
[85]Zheng D H,Wooter M H,Zhou Q,Dohm G L. The effect of exercise on ob gene expression[J].Biochemncal and Biophysical Research Communication,1996,225(3):646-650.
[86]Klein S,Coppack S W, Mohammed-Ail V,Landt M. Adipose tissue leptin production and plasma leptin kinerics in hamans[J].Diabetes,1996,45:984-986.
[87]Ahima R S,Prabacaran D,Mantzoros C,Qu D,Lowell B,Maratos-flier E,Flier J S. Roles of leptin in the neuroendocrine response to fasting[J].Nature,1996,382:250-252.
[88]Yu W H, Kimura M, Walczewska A Karanth S,McCann S M. Role of leptin in hypothalamic-pituitary function[J].Proceeding of the National Academy of Sciences of the United States of America,1996,94:1023-1028.
[89]Mizuno T M,Kleopoulos S P,Bergen H T,Roberts J L, Priest C A, Mobbs C V. Hypothalamic pro-opiomelanocortin mRNA is reduced by fasting and in ob/ob and db/db mice, but is stimulated by leptin [J].Diabetes,1998,47(2):294-297.
[90]孙长颖,李颖,王旭,马瑞.leptin在男性青春期发育中的作用[J].中华预防医学杂志,2004a,38(4)：331-333.
[91]孙长颖,李颖,王舒然,闻颖.leptin在女性青春期发育中的作用及其与胰岛素、生长激素和性激素的关系[J].中国临床康复,2004b,8(24)：5169～5117.
[92]陈赛,李尚为,唐海峰,张金虎,尹海林.leptin对小鼠着床前胚胎发育影响的体外研 究[J].四川大学学报(医学版),2004,35(6)：806～808.
[93]贲晓明,秦玉明,喻文亮,张伟利,蔡威,吴圣楣.胎盘肥胖基因表达与胎儿宫内生长发育相关性研究[J].中华围产医学杂志,2002,5(1)：60～63.
[94]荣飞,林季,颜光涛.瘦素与损伤修复[J].医学综述,2008,14(11)：1605～1067.
[95]Schafer K, Halle M, Goeschen C, Dellas C, Pynn M,J.Loskutoff D,Konstantinides S. Leptin promotes vascular remodeling and neointimal growth in mice [J].Arteriosclerosis, Thrombosis, and Vascular Biology,2004,24(1):112-117.
[96]Burcelin R,Kamohara S,Li J,Tannenbaum G S,Charron M J,Friedman J M. Acute intravenous leptin infusion increases glucose turnover but not skeletal muscle glucose uptake in ob/ob mice[J].Diabetes,1999,48(6):1264-1269.
[97]Chen H C,Farese R V. Inhibition of triglyceride synthesis as a treatment strategy for obesity:lessons from DGAT1-deficient mice[J],Arteriosclerosis, Thrombosis, and Vascular Biology,2005,25(3):482-486.
[98]肖德明,徐忠世,庚智文,卢小虎,胡新佳.血清leptin与骨质疏松关系的研究[J].中国骨肿瘤骨病,2004,3(6)：352～354.
[99]朱玉贤,李毅.现代分子生物学[M].第二版.北京：高等教育出版社,2002：371～376.
[100]William J C K, Kubelik A R,Livak K J. DNA polymorphism amplified by arbitrary primers are useful genetic markers[J].Nucleic Acids Research,1990,18(22):6531-6537.
[101]Velappam N,Sondgrass J L, Hakovirta J R. Rapid identification of pathogenic bacteria by single-enzyme amplified fragment length polymorphism analysis[J].Diagnstic Microbiology and infections Disease,2001,39(2):77-83
[102]王志林,赵树进,吴新荣.分子标记技术及其进展[J].生命的化学,2001,21(4)：39～42.
[103]黎裕,贾继增,王天宇.分子标记的种类及其发展[J].生物技术通报,1999,15(4)：19～22.
[104]Serapion J,Kucudtas H,Feng J,Liu Z. Bioinformatic mining of type I mincrosatellite from expressed sequence tags of channel catfish(Ictalurus punctatus) [J].Animal Genetics,2003,34 (6):445.
[105]张丽娟,张保军,耿社民.微卫星标记与动物遗传育种[J].黄牛杂志,2003,29(2)：49.
[106]冯龙仁,牛安欧.微卫星简单重复序列锚定PCR扩增技术(SSR-PCR)及其在寄生虫基因研究中的应用[J].实用寄生虫杂志,2000,8(1)：33～34.
[107]杨昭庆,洪坤学,褚嘉佑.单核苷酸多态性的研究进展[J].国外医学：遗传学分册,2000,23(1)：4～8.
[108]张效萌,韩金祥.单核苷酸多态性的检测及其在医药领域的应用[J].生命的化学,2001,21(4)：325～327.
[109]刘炬,徐兵河.单核苷酸多态性与乳腺癌患者的预后[J].癌症进展,2007,5(3)：242～248.
[110]刘越,吕社民.单核苷酸多态性影响基因功能的机制[J].生命的化学,2008,28(4)：214～216.
[111]唐棣,王志民.SNPs检测方法研究进展[J].上海交通大学学报(农业科学版),2007,25(4)：405～418.
[112]李艳杰,龙火生,李影,杨公社.单核苷酸多态性(SNP)的研究进展和应用[J].Animal Science Veteringany Medicine,2003:22(4):16-18.
[113]萨姆布鲁克J,弗里奇E F,曼尼阿蒂斯T.分子克隆实验指南[M].金冬雁,黎孟枫等译.第二版,北京：科学出版社,1992.
[114]Masuzaki H,Ogawa Y,Hosoda K, Kawada T,Fushiki T,Nakao K. Augmented expression of the obese gene in the adipose tissue from rat fed high-fat diet[J]. Biochemncal and Biophysical Research Communication,1995,216(1):355-358.
[115]Maffei M, Halaas J, Ravussin E, Pratley R E, Lee G H, Zhang Y, Fei H, Kim S, Lallone R, Ranganathan S, Kern P A, Friedman J M. Leptin levels in human and rodent:measurement of plasma Leptin and ob RNA in obese and weight-reduced subject[J]. Nature Medicine,1995,1:1155-1161.
[116]杨金权,胡雪莲,唐文乔,林弘都.长江口邻近水域刀鲚的线粒体控制区序列变异与遗传多样性[J].动物学杂志,2008,43(1)：8～15.
[117]Nei M. Molecular population genetics and evolution. North-Holland Publishing company, Amsterdam. Oxford American Elsevier publishing company, Inc. New York,1975.
[118]King J L. The role of mutation in evolution. Proc.6th Berkeley Symp. Statist, and Probab. 1972,69-100.
[119]Allendorf F W. Isolation,Gene flow and genetic differentiation among populations[J]. Genetics and Conservation,1983,18(3):51-65.
[120]Reid W V,Kenton R M. Keeping options alive:the scientific basis for conserving biodiversity[M].World Resouces Institute,Washington D C,1989.
[121]Castela V,Ruedi M,Excofifer L, Inanez C, Arlettaz R,Hausser J. Is the Gibraltar Strait a barrier to gene flow for the bat Myotis myotis(Chiroptera:Vespertilionidae)[J].Molecular Ecology,2000,9(11):1761-1772.
[122]Keller I,Nentwig W, Iargiader C R. Recent hahitat fragmentation due to roads can lead to significant genetic differentiation in an abundant flightless ground beetle Molecular Ecology, 2004,9:1761～1772.
[123]Su H,Qu L J,He K, Zhang Z,Wang J,Chen Z,Gu H. The Great wall of China:a physical barrier to gene flow? [J].Heredity,2003,90(3):212～219.
[124]Beh W,Seitz A. The influence of land use on the genetic structure of populations of the common frog Rana temporaria[J].Biology Conservation,1990,54(3):239-249.
[125]Kelley S T,Farrell B D,Mitton J B. Effects of specialization on genetic differentiation in sister species of bark beetles[J].Heredity,2000,84(14):219-227.
[126]Whittaker R H,Levin S A,Root R B.Niche,habitat,and ecotope[J].American.Nature, 1973,107:321-338.
[127]Reid W V. Kenton R M.Keeping options alive:the scientific basis for conserving biodiversity[M].World Resouces Institute,Winshingtn D C,1989.
[128]Jiang Z H,Gison J P.Genetic polymorphisms in the leptin gene and their association with fatness in four pig breeds[J].Mammalian Genome,1999,10(2):191～193.
[129]李毅.松辽黑猪ob基因单核苷酸多态性及其部分经济性状的相关分析[D].吉林：吉林大学,2005.
[130]Houseknecht K L, Portocarrero C P. Leptin and its receptors:regulators of whole body energy homeostasis[J].Domestic Animal Endocrinology,1998,15(6):457-475.
[131]杜清友,王会信.胰岛素样生长因子结合蛋白研究进展[J].生物化学与生物物理进展,1998,25(2)：104～105.
[132]Stephens T W, Basinski M, Bristow P K, Buevalleskey J M, Burgett S G,Craft L, Halb J, Hoffmann J, Hsiung H M, Kriauciuna A, Mackellar W, RosteckJr P R, Schoner B, Smith D,Tinsley F C,Zhang X,Heiman M. The role of neuropeptide Y in the antiobesity action of the obese gene product [J].Nature,1995,12,377(6549):530-532.
[133]刘桂琼,姜勋平,丁家桐.梅山猪ob基因多态性与繁殖性状的相关性研究[J].中国畜牧杂志,2002,38(4)：15～16.