多花黑麦草遗传连锁图的构建

利用多花黑麦草(Loliummu ltiflorum)细胞质雄性不育(CMS)F₁群体(124个个体),以来自父本和母本的分离位点采用SSR、AFLP、EST-CAPS(表达序列标签酶切扩增多态性)和RGA-CAPS(抗病基因类似物酶切扩增多态性)等标记方法进行多态性的选择并对父、母本分别构建遗传连锁图。母本连锁图包含362个标记位点,分别包含SSR标记240个、AFLP标记84个、EST-CAPS标记24个和RGA-CAPS标记14个,由7个连锁群组成,覆盖全长776.4cM,各连锁群长度在85.8至135cM之间,相邻标记之间的平均距离为2.14cM;父本7个连锁群,包含376个标记,其中SSR标记252个、AFLP标记82个、EST-CAPS标记29个和RGA-CAPS标记13个,连锁图全长710.0cM,相邻标记之间的平均距离为1.89cM;标记密度高、标记分布均匀、连锁图长度中等。     

Cytogenetic mapping of 11 functional genes to chicken chromosomes by fluorescence in situ hybridization

With chicken bacterial artificial chromosome (BAC) DNA as probes, 11 non‐assigned functional genes were localized to chicken chromosomes 1 or 2 by fluorescence in situ hybridization (FISH). The 11 genes and their chromosomal positions are as follows: ALVEB5, 1p26‐24; ACO2, 1p16‐14; HSP108, 1p14‐13; CD4, 1q11; HSD3B; 1q11; SOD1, 1q14‐21; LAMP1, 1q24‐31; P2Y5, 1q35‐36; EN2, 2p31‐24; NPY, 2p14‐13 and CA2, 2q31‐32. Metaphase chromosome spreads used for hybridization were prepared from embryonic chicken fibroblast cultures. The gene position was identified according to the international standardized G‐banded karyotype of chicken by measuring the relative fractional length from the telomere of the p‐arm to the hybridization signal (FLpter). The 11 genes mapped newly will enrich the cytogenetic map and serve as additional anchor markers for integrating the cytogenetic map with the genetic map of chicken.     

A male bovine linkage map for the ADR granddaughter design

The aim of this paper is to present the construction of a male genetic linkage map as a result of the bovine genome mapping project, which is a common effort of the German cattle breeding federation (ADR), four animal breeding institutes, three blood group laboratories and two animal data and breeding value evaluation centres. In total 20 grandsires with 1074 sires were provided from the German cattle population as reference families, 16 of these paternal half‐sib groups are German Holstein families (DH), three are German Simmental (ST) families, and one is a Brown Swiss family (BS). Of 265 markers included in the linkage map, 248 were microsatellite markers, five were bovine blood group systems, eight SSCP markers and four proteins and enzymes. More than 239 000 genotypes resulted from typing the offspring for the respective markers and these were used for the construction of the map. On average 478 informative meioses were provided from each marker of the map. The summarized map length over all chromosomes was 3135.1 cM with an average interval size of 13.34 cM. About 17, 35.7 and 79.1% of the map intervals showed a maximum genetic distance between the adjacent markers of 5, 10 and 20 cM, respectively. The number of loci ranged from two (pseudoautosomal region of the sex chromosome, BTAY) to 15 (BTA23) with an average of 8.8 markers per chromosome. Comparing the length of the chromosomes shows variation from 49.6 cM for BTA26 to 190.5 cM for BTA1 with a mean of 107.7 cM for all autosomes of the genetic linkage map. It was possible to identify chromosomal discrepancies in locus order and map intervals by comparison with other published maps. The map provided sufficient marker density to serve as a useful tool for a scan of segregating quantitative trait loci.     

红三叶遗传图谱构建及抗白粉病基因QTL定位

本研究以感(岷山红三叶)、抗(澳大利亚红三叶品种♀Sensation×Renegade♂杂交新品系“甘农PR1”)白粉病红三叶材料为父母本杂交并种植成苗经人工接菌后筛选出抗、感白粉病的F₁群体为作图群体,利用AFLP标记构建红三叶高密度遗传图谱,并利用区间作图法对抗白粉病QTL进行了定位分析,可以为红三叶抗白粉病基因克隆和转基因等分子辅助育种奠定基础。结果表明,149个AFLP标记构建得到的遗传图谱包含7个连锁群(LG1,LG2,LG3,LG4,LG5,LG6和LG7),遗传图谱的总距离为640.5 cM。其中,LG1连锁群的遗传距离(140.6 cM)和标记间平均距离(9.4 cM)均最大;LG4连锁群的遗传距离(55.2 cM)和标记间平均距离(1.8 cM)最小。应用区间作图法对红三叶抗白粉病基因进行QTL分析定位,共检测到5个抗白粉病相关QTL位点(qrp-1,qrp-2,qrp-3,qrp-4和qrp-5),其中qrp-1、qrp-2、qrp-3和qrp-4位于LG4连锁群上,qrp-5位于LG5连锁群上。5个QTL位点对抗白粉病的贡献率为29%~90%,qrp-1对红三叶白粉病抗性的贡献率最大(90%),为主效QTL。     

基于SRAP分子标记构建的菊苣遗传连锁图谱

基于SRAP标记,以遗传关系和表型差异大的菊苣亲本PI 651947和PI 652007杂交获得的84个F₁单株为作图群体,进行连锁图谱的构建。采用Map Manager QTX b20软件进行连锁分析,分别构建了PI 651947和PI 652007的分子连锁框架图,共获得77个SRAP标记,其中父本遗传图谱涉及4个连锁群,包含19个标记,图谱总长为450.9 cM,标记间平均图距为23.7 cM。母本遗传图谱涉及13个连锁群,包含58个标记,图谱总长为1404.8 cM,标记间平均图距为24.2 cM。研究结果可为菊苣重要农艺性状QTL定位奠定基础,为菊苣分子育种研究提供了基础信息。     

对家蚕多星纹基因ms的SSR标记定位分析

对家蚕幼虫斑纹突变多星纹基因ms进行分子标记定位,有益于对多星纹性状的分子标记辅助选择和对该基因的定位克隆研究。利用家蚕雌性染色体在减数分裂过程中不发生交换的特点,采用幼虫斑纹为素斑(p)的家蚕品系C108,幼虫斑纹为多星纹(ms)的家蚕品系g02,组配正反交群体(g02×C108)F1♀×g02♂和g02♀×(g02×C108)F1♂,分别记作BC1F和BC1M,利用已经构建的家蚕SSR分子标记连锁图谱中第12连锁群上的18对SSR标记引物在亲本间进行多态性筛选。BC1F群体中的普通斑个体均表现出与(g02×C108)F1相同的杂合型带型;而所有多星纹个体的带型与亲本g02一致,为纯合型。结果筛选出S1206、S1208、S1210、C5553S3共4个与家蚕ms连锁的SSR标记。利用BC1M群体构建家蚕ms及其连锁的SSR标记遗传连锁图,连锁图的图距为34.5 cM,4个SSR标记及ms的排列次序为S1206—S1208—S1210—C5553S3—ms,ms位于34.5 cM处,与ms最近的标记为C5553S3,遗传距离为12.4 cM。依据该SSR标记遗传连锁图谱可对ms进一步精确定位。     

应用于家蚕F2代分离群体的两种连锁作图方法的作图效果比较

构建精细的家蚕分子连锁图谱需要合适的作图方法。应用Mapmaker/EXP3.0和F2lnkgsilk两种作图方法,对家蚕F2代91个个体为分离群体的300个AFLP标记和69个个体为分离群体的470个RAPD标记数据进行了比较分析。用F2lnkgsilk作图方法分析得到的分群数与Mapmaker/EXP3.0作图方法相比有所增加,而总的图距却显著增加,平均图距也有较大的增加,但产生的二联体数基本相同。就单个连锁群而言,两种作图分析方法的连锁标记及其数目80%以上相同,但是标记间的排列顺序70%以上有差异;用F2lnkgsilk作图方法分析得到的单个连锁群的总图距和平均图距也相应增大。     

A canine linkage map: 39 linkage groups

A low resolution canine marker map is an important tool in the further advancements in genetic analysis of dog breeds and the control and reduction of the frequency of inherited diseases. This study presents a genetic linkage analysis with 39 linkage groups using 222 polymorphic canine markers based on typing in the International DogMap reference families, consisting of 129 Beagle and German Shepherd dogs. Of these 39 linkage groups, 14 have been assigned to canine chromosomes by fluorescence in-situ hybridization (FISH). These results are a further refinement on the first linkage groups from the International DogMap collaboration and represent a continuing collaboration.     

Twenty-eight new microsatellite loci in chicken and their cross-species amplification in Japanese quail and helmeted guinea fowl

Twenty‐eight original chicken microsatellite markers were isolated and characterized to determine their utility as cross‐reactive markers for comparative genetic mapping in the order Galliformes. Primer pairs were typed in 12 unrelated chickens and also tested on Japanese quail and helmeted guinea fowl deoxyribonucleic acid (DNA). Polymorphism was observed in 23 (82.1%) of the markers and the average number of alleles per locus was 2.9 while the mean heterozygosity was 0.19. Eleven (39.3%) of the chicken markers cross‐reacted with Japanese quail DNA and 2 (7.1%) with helmeted guinea fowl DNA. The cross‐reactive markers described would serve as useful resources for comparative genetic mapping in poultry species belonging to the order Galliformes.     

家鸡豆冠基因定位的研究

构建家鸡1号染色体短臂部分连锁图谱及豆冠冠型基因的定位。通过测交选择豆冠冠型基因型纯合的2只吐鲁番斗鸡(♂)和6只玫瑰冠鸡(♀)为亲本,采用F2代试验设计建立F2代494只,依据己公布的家鸡遗传连锁图谱,分别在1号染色体上筛选了13对微卫星标记,运用MapMaker/EXP 3.0和MapDraw 2.1软件绘制遗传连锁图谱,并结合记录的表型性状值对冠型性状进行定位分析。经卡方检验F2代冠型符合9∶3∶3∶1的遗传定律。在连锁分析中除MCW0007位外,其余12个微卫星座位在试验群体中均表现较高的基因杂合度和多态信息含量。对冠型性状进行初步的分析,结果显示,MCW0428、ADL0319、LEI0174和MCW0058标记位点LOD值＞3,表明这4个微卫星位点与豆冠冠型基因存在连锁关系,其中LEI0174与豆冠基因遗传距离最近为0.12 cM。     

Linkage mapping of four chicken calpain genes

Calpains are intracellular Ca²⁺‐dependent proteases and enzymes that contribute to growth and meat quality. In the present study, we identified polymorphisms in four calpain genes (CAPN1, CAPN2, CAPN3, and CAPN1.5) expressed ubiquitously in chicken using polymerase chain reaction‐restriction fragment length polymorphism, and mapped them using two backcross families (East Lansing (EL) and Kobe University (KU)). CAPN2 and CAPN1.5 mapped to two locations on chromosome 3 about 30 cM apart, while CAPN3 mapped to chromosome 5. CAPN1 was linked to a previously unlinked microsatellite marker LEI0140 to form a new linkage group called E66. CAPN2 and CAPN3 extend the amount of conserved synteny between chicken chromosome 3 and human chromosome 1, and between chicken chromosome 5 and human chromosome 15, respectively. Although CAPN2, CAPN3, and CAPN1.5 were found in the University of California Santa Cruz chicken genome browser gateway, CAPN1 and LEI0140 were not in specific genomic positions.     

A pig–human comparative RH map comprising 20 genes on pig chromosome 13q41 that harbours the ETEC F4ac receptor locus

The enterotoxigenic Escherichia coli (ETEC) F4ac is a major cause of diarrhoea in newborn and young pigs. The locus for the intestinal ETEC F4ac receptor (F4acR) has been mapped to pig chromosome (SSC) 13q41 with known homology to human chromosome (HSA) 3q21 and q29. However, the causative gene and mutation(s) remain unknown. The aim of this study was to characterize gene-derived markers on SSC13q41 for fine mapping of the F4acR locus, and construct a high-resolution pig–human comparative map to select positional candidate genes for F4acR. Pig-specific sequence-tagged site markers were developed for 20 genes that are located in a 6.8-Mb region on HSA3q21 and q29, and a total of 34 single-nucleotide polymorphisms (SNPs) were identified in 14 of 20 markers developed. Eighteen markers were mapped to SSC13q41, while the other two markers ( PLXNA1 and KLF15 ) were assigned to SSC13q32 and SSC7q13, respectively, by radiation hybrid mapping. This result showed that there was a small conserved segment on SSC7 corresponding to HSA3q21. A framework map comprising 18 markers on SSC13q41 was established, refining the synteny breakpoint on SSC13q41 to a region of 12.3 centiRay. The comparative radiation hybrid (RH) map revealed three interesting candidate genes for F4acR from the human genome, viz. MUC4 , MUC13 and MUC20 . Linkage analysis with six marker polymorphisms revealed that MUC4 had the most significant linkage with the F4acR locus.     

四倍体紫花苜蓿遗传图谱的初步构建

分别以早熟低产和晚熟高产苜蓿单株为父母本,通过人工杂交构建了四倍体紫花苜蓿(Medicago Sativa)F1遗传作图群体,采用单因子变量分析法,以降落式PCR和常规PCR结合的反应程序,建立了适宜于紫花苜蓿的分子标记扩增体系;应用130对SSR引物进行筛选,获得60对引物在父母本间存在多态性而被用于绘制遗传连锁图。采用PAGE电泳分析,对作图群体进行基因型分析。通过TetraploidMap软件对60个SSR标记进行连锁作图分析,有44个标记可以定位在8个连锁群上,占总标记数的33.8%,覆盖遗传距离979 cM,两标记间平均图距为22.25 cM,初步构建了四倍体紫花苜蓿遗传图谱的框架图,还需要进一步添加标记数量增大其饱和度,为重要性状的QTL定位奠定基础。     

Fine mapping a quantitative trait locus affecting ovulation rate in swine on chromosome 8

Ovulation rate is an integral component of litter size in swine, but is difficult to directly select for in commercial swine production. Because a QTL has been detected for ovulation rate at the terminal end of chromosome 8p, genetic markers for this QTL would enable direct selection for ovulation rate in both males and females. Eleven genes from human chromosome 4p16-p15, as well as one physiological candidate gene, were genetically mapped in the pig. Large insert swine genomic libraries were screened, clones were isolated and then screened for microsatellite repeats, and informative microsatellite markers were developed for seven genes (GNRHR, IDUA, MAN2B2, MSX1, PDE6B, PPP2R2C, and RGS12). Three genes (LRPAP1, GPRK2L, and FLJ20425) were mapped using genotyping assays developed from single nucleotide polymorphisms. Two genes were assigned since they were present in clones that contained mapped markers (HGFAC and HMX1). The resulting linkage map of pig chromosome 8 contains markers associated with 14 genes in the first 27 cM. One inversion spanning at least 3 Mb in the human genome was detected; all other differences could be explained by resolution of mapping techniques used. Fourteen of the most informative microsatellite markers in the first 27 cM of the map were genotyped across the entire MARC swine resource population, increasing the number of markers typed from 2 to 14 and more than doubling the number ofgenotyped animals with ovulation rate data (295 to 600). Results from the revised data set for the QTL analysis, assuming breed specific QTL alleles, indicated that the most likely position of the QTL resided at 4.85 cM on the new linkage map (F1,592 = 20.5150, genome-wide probability less than 0.015). The updated estimate of the effect of an allele substitution was -1.65 ova for the Meishan allele. The F-ratio peak was closest to markers for MAN2B2 (4.80 cM) and was flanked on the other side by markers for PPP2R2C. Two positional candidate genes included in this study are MAN2B2 and RGS12. These results validate the presence of a QTL affecting ovulation rate on chromosome 8 and facilitate selection of positional candidate genes to be evaluated.     

对家蚕第2隐性赤蚁基因(ch-2)的SSR标记定位及连锁分析

家蚕蚁蚕体色突变之一的第2隐性赤蚁(ch-2)有作为特殊遗传系统的实用价值。采用家蚕正常黑蚁品种P50和第2隐性赤蚁品种k04为亲本组配正反交群体,回交后获得回交群体(k04×P50)×k04和k04×(k04×P50),分别记作BC1F和BC1M,基于雌性家蚕的W与Z染色体不发生交换的特点,用已构建的家蚕SSR分子标记连锁图谱对ch-2基因进行定位和连锁分析。在第18连锁群上的20个多态性标记中共筛选出7个与ch-2基因连锁的SSR标记。根据第18连锁群上已有但不表现多态性的微卫星序列,寻找其所在Scaffold上的其他SSR位点并设计引物,结果找到2个新的多态性SSR标记。BC1F群体中的所有黑蚁个体均表现出与F1(k04×P50)相同的杂合型带型,而所有赤蚁个体带型与亲本k04一致,为纯合型,说明家蚕ch-2基因位于第18连锁群。利用另一个群体BC1M构建了ch-2基因的SSR分子标记遗传连锁图,连锁图的遗传距离为70.7cM,ch-2基因位于69.6 cM处。2个与ch-2最近的SSR标记为S1814和S1819,与ch-2的距离分别为7.9、1.1 cM。     

RFLP在动物育种中应用的前景兼对数量性状基因定位的讨论

对于数量性状能否作基因定位的问题,过去认为这是不可能的。因为数量性状是受微效多基因决定。而这些基因的数目和单独效应又很难确定。下面刊出的李玉奎和吴常信两位先生撰写的《RFLP在动物育种中应用的前景》一文对这一问题的解决介绍了可供研究的途径。     

水稻饲料营养含量的QTL定位分析

应用85个SSR标记,对普通野生稻与粳稻台中65为亲本建立的F₂群体进行基因检测,构建了覆盖水稻基因组12条染色体的SSR分子标记连锁图,采用Mapmaker/QTL1.0统计软件对决定水稻饲用营养价值的粗蛋白、粗纤维、粗脂肪、粗灰分、硅酸和可溶性糖含量的基因座位进行了定位分析。结果定位了影响粗蛋白含量的3个QTLs,影响粗脂肪含量的1个QTL,影响可溶性糖含量的3个QTLs,影响硅酸含量的2个QTLs,这9个QTLs分别位于第1,2,4,7,8,9,10和11染色体上。其中主效QTL4个,分别是影响粗脂肪含量的qCEE-1(贡献率56.8%),影响可溶性糖含量的qCWSC-4(贡献率23.1%)和qCWSC-7(贡献率25.0%),影响硅酸含量的qCS-9(贡献率15.9%),其余5个为微效QTL。没有检测到影响粗纤维含量和粗灰分含量的QTL。     

微卫星标记与固始鸡及部分外来鸡种体型性状及屠体性状的相关分析

选择与鸡体型参数和屠体性状紧密相关的10个微卫星位点,用变性聚丙烯酰胺凝胶电泳法分析其与固始鸡5个系和4个非固始鸡品系共9个群体体型性状及屠体性状相关关系。结果表明:检测到4个标记(MCW0295、MCW0006、MCW0185、ADL0192)与5个性状(体重、胸宽、胸深、胫长、体斜长)存在显著相关,在各个群体中,标记MCW0295与体重相关比较显著,标记ADL0192与体斜长相关。在固始鸡群体中,标记MCW0006和MCW0185与体型性状各个性状相关比较显著。检测到3个标记分别与3个屠体性状存在不同程度的相关:在固始鸡原种、父母代中,标记MCW0295与胸肌率相关。在固始鸡原种和商品代中,标记MCW0006与全净膛率相关。在艾维茵鸡中,MCW0294与腿肌率相关。说明这些标记位点可能是控制特定性状的主效基因,或与控制性状的主效基因连锁。     

地方鸡种与隐性白羽鸡遗传变异的AFLP指纹

利用6对AFLP引物组合对我国12个地方鸡种和引进鸡种隐性白羽鸡进行了遗传检测,统计了每个引物组合在各个品种中检测到的多态性条带和特异性条带,计算了13个鸡种的遗传相似系数和遗传距离,并据此构建了UPGMA聚类关系图,分析了所研究鸡种的遗传关系.结果表明:6对AFLP引物组合在13个鸡种中共检测到290条多态性条带,平均每个引物组合产生48.3奈多态性标记,同时在每个品种群体中还检测到了数量不等的特异性条带,其中寿光鸡和东乡黑鸡最多,为9条,旧院黑鸡、兴义矮脚鸡和隐性白羽鸡最少,为1条.13个鸡种聚为4类,其中隐性白羽鸡单独聚为一类,鸡种间的遗传相似系数及聚类结果与各个鸡种的地理分布、现实状况相吻合,从而表明AFLP指纹用于我国地方鸡种的遗传多态性分析、品种鉴定及品种间亲缘关系分析是可行的.     

Genome-wide linkage and QTL mapping in porcine F2 families generated from Pietrain, Meishan and Wild Boar crosses

Three informative pig F₂ families based on European Wild Boar (W), Meishan (M) and Pietrain (P) crosses have been used for genome‐wide linkage and quantitative trait loci (QTL) analysis. Altogether 129 microsatellites, 56 type I loci and 46 trait definitions (specific to growth, fattening, fat deposition, muscling, meat quality, stress resistance and body conformation) were included in the study. In the linkage maps of M × P, W × P and W × M families, average spacing of markers were 18.4, 19.7 and 18.8 cM, the numbers of informative meioses were 582, 534 and 625, and the total lengths of autosomes measured were 27.3, 26.0 and 26.2 Morgan units, respectively. Maternal maps were on average 1.3 times longer than paternal maps. QTLs contributing more than 3% of F₂ phenotypic variance could be identified at p < 0.05 chromosome‐wide level. Differences in the numbers and positions of QTLs were observed between families. Genome‐wide significant QTL effects were mapped for growth and fattening traits on eight chromosomes (1, 2, 4, 13, 14, 17, 18 and X), for fat deposition traits on seven chromosomes (1, 2, 3, 4, 6, 7 and X), for muscling traits on 11 chromosomes (1, 2, 3, 4, 6, 7, 8, 12, 14, 15 and X), for meat quality and stress resistance traits on seven chromosomes (2, 3, 6, 13, 16, 18 and X), and QTLs for body‐conformation traits were detected on 14 chromosomes. Closely correlated traits showed similar QTL profiles within families. Major QTL effects for meat quality and stress resistance traits were found on SSC6 in the interval RYR1‐A1BG in the W × P and M × P families, and could be attributed to segregation of the RYR1 allele T derived from Pietrain, whereas no effect in the corresponding SSC6 interval was found in family W × M, where Wild Boar and Meishan both contributed the RYR1 allele C. QTL positions were mostly similar in two of the three families for body conformation traits and for growth, fattening, fat deposition and muscling traits, especially on SSC4 (interval SW1073‐NGFB). QTLs with large effects were also mapped on SSC7 in the major histocompatibility complex (MHC) (interval CYP21A2‐S0102) and affected body length, weight of head and many other traits. The identification of DNA variants in genes causative for the QTLs requires further fine mapping of QTL intervals and a positional cloning. However, for these subsequent steps, the genome‐wide QTL mapping in F₂ families represents an essential starting point and is therefore significant for animal breeding.