Genome-wide development of interspecific microsatellite markers for Saccharum officinarum and Saccharum spontaneum

Sugarcane has a large, complex, polyploid genome that has hindered the progress of genomic research and molecular marker-assisted selection. The user-friendly SSR markers have attracted considerable attention owing to their ideal genetic attributes. However, these markers were not characterized and developed at the genome-wide scale due to the previously lacking high-quality chromosome-level assembled sugarcane genomes. In this present study, 744 305and 361 638 candidate SSRs were identified fro...     

Genome-wide Association Study of Teats Number Traits in Kele Pigs

Teats number is one of the most important reproductive traits,and closely related to the economic benefit in pig industry.In order to reveal the underlying genetics of left teats number,right teats number and total teats number traits,a genome-wide association study（GWAS）was performed.Samples of DNA were collected to genotyping for 22 Kele pigs using the Illumina Porcine SNP 60K Chip.The GWAS was performed using a mixed-effects model and linear regression approach.When a genome-wide threshold was determined using the Bonferroni method（P<2.06E-5),4 single nucleotide polymorphism（SNP）markers were potentially associated with left teats number,right teats number and total teat number.However,3 SNPs were significant associated and 18 SNPs were potentially associated in chromosomes level.304 Ensembl genes were retrieved around 1 cM of the associated SNPs.The candidate genes in Wnt and Fgf signaling pathway（BTRC,FGF5,FGF8,BMP3,RASGEF1B and HMGB3）might have effect on target traits.These results provided valuable information about the selective breeding for Kele pigs.     

Genome-wide association study for spot blotch resistance in Afghan wheat germplasm

Spot blotch (SB), caused by Bipolaris sorokiniana, is a devastating disease of wheat globally, especially in South Asia and South America. Understanding the genetics of resistance to SB is important for developing breeding strategies to improve resistance. A panel of 301 genotypes from Afghanistan was phenotyped over two crop seasons using a mixture of virulent B. sorokiniana isolates and genotyped using DArTSeq to obtain genome-wide markers. Fifty genotypes (16.6%) showed disease scores less than the resistant control. Principal component analysis using the genotypic data clustered the genotypes into five different groups. Among models used for genome-wide association mapping, the multilocus mixed model, and fixed and random model circulating probability unification algorithms were most effective in identifying significant marker-trait associations (MTA). Twenty-five MTAs at p ≤ .001 were identified on chromosomes 1A, 1B, 1D, 2B, 2D, 3A, 3B, 4A, 5A, 5B, 6A, 7A, and 7D, indicating the quantitative nature of resistance to SB. Phenotypic variation explained by these markers ranged from 2.0% to 17.7%, and genomic regions on the chromosomes 1D, 2D, 3A, 3B, 4A, 5A, and 5B coincided with loci identified in previous studies. Three single nucleotide polymorphism (SNP) markers on chromosomes 1B (SNP 1113207) and 5A (SNPs 5411867 and 998276) were significant in both crop seasons as well as in the combined analysis across seasons. Marker 5411867 is close to Vrn-A1, shown to be associated with SB in previous studies. Furthermore, among known SB resistance genes, Sb2 on chromosome 5B was predicted to be significant in this panel.     

利用高密度SNP对猪血糖和糖基化血清蛋白性状的全基因组关联分析

【目的】测定苏太猪和白色杜洛克×二花脸F₂资源家系240 d血糖（glucose,GLU）和糖基化血清蛋白（glycosylated serum proteins,GSP）浓度,采用全基因组关联分析定位影响GLU和GSP的染色体位点,为最终鉴别影响该性状的因果基因奠定基础,同时为人类低血糖症和糖尿病的遗传学研究提供参考。【方法】分别将435头苏太猪和760头白色杜洛克×二花脸F2资源家系F₂个体在相同条件下饲养至240日龄进行统一屠宰,收集血液后分离血清,利用全自动生化分析仪测定GLU和GSP浓度。采集猪只耳组织提取DNA并测定DNA浓度。将质检合格的DNA样品利用Illumina porcine 60K SNP芯片判定基因型。运用PLINK软件对SNP判型结果进行质控,将合格的SNP标记用于后续的关联性分析,利用广义混合线性模型及R语言GenABEL软件包进行全基因组关联分析,定位影响苏太猪和白色杜洛克×二花脸F₂资源家系240 d血清GLU和GSP含量的染色体位点。根据全基因组关联分析结果从Ensembl或NCBI网站上分析可能的位置候选基因。【结果】全基因组关联分析共检测到5个与血清GLU和GSP达染色体显著水平相关的SNP位点。其中白色杜洛克×二花脸F₂资源群体在10号染色体（SSC10）24.67Mb处定位到与血清GSP含量显著相关的SNP（ALGA0057739,P=1.58×10^-5）,解释表型变异为3.72%。苏太猪群体共检测到2个与血清GSP显著相关的SNP（ALGA0108699和DRGA0017552,P=1.45×10^-5）,解释表型变异均为3.72%。使用猪参考基因组序列（10.2版本）,无法定位到具体的染色体位置。通过人、猪比较基因组分析,这两个SNP都位于SSC8,距STPG2基因3’端约180.0-193.0 kb。将两个群体进行Meta分析,未发现新的与GSP显著相关的SNP;在1号染色体250.32Mb处（DRGA0002016,P=2.48×10^-5）和14号染色体43.97Mb处（ASGA0062984,P=1.29×10^-5）,定位到与血清GLU显著相关的SNP。通过搜寻显著相关SNP所在染色体区域内的注释基因,发现ASPM、TRPM3和KCTD10 等基因是影响血清GSP和GLU的重要候选基因。【结论】检测到5个显著影响猪血清GLU和GSP的SNP位点。这些SNP位点所处染色体区域内的ASPM、TRPM3、STPG2和KCTD10基因是影响血清GSP和GLU的重要候选基因。     

盐胁迫下甘蓝型油菜发芽期下胚轴和根长的全基因组关联分析

【目的】解析甘蓝型油菜发芽期根和下胚轴发育及耐盐性的调控位点,筛选油菜耐盐性相关的候选基因,可为油菜耐盐性改良提供依据。【方法】以317份具有代表性的甘蓝型油菜自交系为材料,在正常生长和盐胁迫条件下进行沙培鉴定,利用芸薹属60K SNP芯片和全基因组关联分析鉴定正常生长与盐胁迫下甘蓝型油菜发芽期根和下胚轴长度显著关联的SNP,并确定其连锁不平衡区间。通过区间内基因的功能注释及盐胁迫下油菜幼苗根和叶片转录组差异表达基因筛选连锁不平衡区间内的重要候选基因,并以实时荧光定量PCR分析候选基因的组织特异性和盐胁迫诱导表达模式,提高候选基因筛选的准确性。【结果】正常生长和盐胁迫下甘蓝型油菜发芽期下胚轴和根长在不同材料间变异较大,频次分布表明目标性状均为数量性状,受多基因调控。全基因组关联分析模型比较表明,MLM+P+K模型为最优模型。以此模型对目标性状进行全基因组关联分析,检测到45个显著关联SNP,其中40个与下胚轴长度显著关联,5个与根长显著关联,单个SNP解释的表型变异分别为9.12%—14.46%和7.67%—8.93%。重复检测的显著相关SNP中,值得注意的是C04染色体的rs8970,同时与4个性状显著关联,表型贡献率为7.67%—12.35%,是唯一在下胚轴长和根长间重复检测到的显著关联SNP。11个重要关联SNP中有6个位于10—442 kb的连锁不平衡区块中。转录组分析表明,11个连锁不平衡区间共包含447个基因,其中15个受盐胁迫诱导表达。转录组和基因功能注释综合分析表明,BnaSRO1、BnaPAGR2、BnaNPH3、BnaMYB124、BnaSAM-Mtase、BnaBIN2、BnaUMAMIT11、BnaEXPA7、BnaRPT3、BnaEF-hand和BnaF3H很可能为各自区间的候选基因。实时荧光定量PCR结果证实除BnaNPH3外,其他基因均在根或下胚轴中受盐胁迫诱导上调表达。组织特异性分析还发现BnaUMAMIT11、BnaPAGR2和BnaEXPA7主要在萌发的根和下胚轴中特异表达,BnaRPT3、BnaBIN2和BnaMYB124虽然呈组成型表达,但在萌发阶段的下胚轴中表达量最高,证实这些基因很可能参与油菜发芽期根和下胚轴生长发育及耐盐性的调节。【结论】全基因组关联分析共鉴定出45个控制油菜发芽期根和下胚轴发育及耐盐性的显著关联SNP。连锁不平衡、转录组和基因功能注释综合分析初步鉴定出11个重要候选基因。     

Whole Genome Methylation Variation Analysis of Longissimus Dorsi Between Tan Sheep and Hu Sheep

It aimed to identify the differential methylation region (DMR) and differential methylation gene (DMG) through the analysis of the genome-wide methylation difference of the longest muscle of the sheep's back of different breeds of sheep,and laid the foundation for the analysis of the differences in sheep skeletal muscle development.In this study,one-year-old sheep(Tan sheep and Hu sheep and Tan-Hu F2) were sanned by the whole genome bisulfite sequencing method(WGBS).The degree of methylation and differentially methylated region in the whole genome DNA of longissimus dorsi(LD) muscle were studied to explore the difference of DNA methylation in level in sheep.The results showed that the methylation (mC) rates of cytosine (C) were 3.55%,3.18% and 3.56% in Tan sheep,Hu sheep and Tan-Hu F2,respectively.A total of 97 731 DMRs and 10 784 related DMGs were detected.In CG,CHH and CHG sequences,the methylation levels of the three groups were not significantly different.419 GO terms and 20 related signal pathways were detected by GO and KEGG analysis,respectively,which were showed to be significantly enriched in cellular process,cell part,binding,long-term depression,and so on.Five candidate genes,ACTA2、ROCK1、CALD1、MYH3 and MYH10 were sreened out in muscle tissues.This study provided genome-wide methylation of pattern of three groups (Tan sheep and Hu sheep and Tan-Hu F2).The results provided reference information for the epigenetic study of Tan sheep and screened candidate genes related to muscle development and meat quality.     

Genome-wide Association Study on Alive Litter Size Trait in Bama Xiang Pigs

In order to identify the molecular markers related to alive litter size of Bama Xiang pigs,the genome-wide association study (GWAS) was used to map and screen the candidate genes affecting the alive litter size trait.Ear tissue samples of 297 Bama Xiang pigs with multiple parity records were collected,and DNA was extracted and genotyped by porcine 50K SNP beadchip.After quality control and genotype imputation,the alive litter size of Bama Xiang pigs were GWAS by Tassel.The results showed that the average number born alive per litter of Bama Xiang pigs increased gradually with the increasing of parity in the range of 1-9 parities.A total of 32 816 SNPs were obtained after quality control and filtration.8 SNPs related to alive litter size of Bama Xiang pigs were screened by genome-wide association analysis,which were significant at genome or chromosome level.Based on the enrichment analysis of the coding genes in the region between 500 kb upstream and downstream of the associated significant SNP loci,and the QTL regions and gene functions related to porcine reproductive traits,4 genes (CAPZB,MSH3,CITED2 and HSD17B7) were finally identified to be candidate genes related to alive litter size of Bama Xiang pigs.     

产蛋前期和产蛋高峰期鸡全基因组甲基化差异及其对肝脏转录组影响的研究

旨在通过对产蛋前期和产蛋高峰期鸡肝全基因组甲基化差异进行分析,解析基因组甲基化对不同发育阶段肝中基因表达差异的影响。本研究采用全基因组重亚硫酸盐测序（WGBS）技术对产蛋前期（20周龄）和产蛋高峰期（30周龄,各3只DNA混池）卢氏绿壳蛋鸡肝全基因组的甲基化水平进行检测,并与已有的肝mRNA转录组数据进行整合分析,探讨基因组甲基化对不同生理阶段基因表达差异的影响。结果表明,全基因组范围约有4%的胞嘧啶（C）发生了甲基化（mC）;两个生理阶段的总体甲基化水平基本一致。共检测到670个差异甲基化区域（DMRs）和356个差异甲基化基因（DMGs）。基因本体（GO）和相关信号通路（KEGG）分析发现,超甲基化DMGs显著富集在发育的正向调控、细胞形态改变的调控、VEGF信号通路、肌动蛋白细胞骨架的调控、粘着斑及间隙连接等相关过程,低甲基化DMGs显著富集在胚胎消化道形态的发生、间充质细胞增殖的正向调控、淀粉和蔗糖代谢及Wnt信号通路等相关过程。基因不同功能区域甲基化水平与基因表达水平有关,启动子（promoter）及基因体（gene body）区域甲基化水平与基因表达水平呈显著负相关,其他区域（内含子、3'UTR）的甲基化水平与基因的表达水平无明显关系。其中,与肝脂质代谢相关的候选基因RASD1、HAO1、UBE2O、MSRB3受甲基化调控。本研究绘制了不同生理时期卢氏绿壳蛋鸡全基因组甲基化图谱,结合mRNA转录组数据阐述了DNA甲基化在基因表达方面的调控作用,并鉴定出了不同生理时期受甲基化调控的基因,为深入研究表观遗传调控在不同生理时期蛋鸡肝代谢中的作用机制提供参考。     

谷子抽穗时间基因SiTOC1的表达与单倍型变异分析

【目的】谷子抽穗时间的适应性表现是广适性新品种选育的基础,分析抽穗时间关键基因的遗传变异和单倍型效应,为品种适应性改良提供基础信息。【方法】通过全基因组关联分析（genome-wide association study,GWAS）,定位谷子抽穗时间关键基因SiTOC1,利用多组学数据库（multi-omics database for Setaria italica,MDSi）提供的SiTOC1数字表达量,分析SiTOC1的组织时空表达特性,并利用原生质体对SiTOC1蛋白进行亚细胞定位。采用qRT-PCR在短日（10 h光照/14 h黑暗）条件下进行SiTOC1 24 h节律表达模式分析。利用有代表性的99份谷子品种,分析SiTOC1编码区和启动子区的遗传多态性、单倍型以及转录水平,并对单倍型与抽穗时间的关系进行鉴定。【结果】在第1染色体物理位置31 456 761 bp处鉴定到了一个显著的关联信号,与抽穗时间紧密相关,该位点附近存在一个拟南芥抽穗期TOC1的同源基因SiTOC1。SiTOC1在光周期响应组织（根、茎、叶等）中高表达,亚细胞定位于细胞核,在傍晚表达量上调,呈现出24 h节律性表达模式。SiTOC1在不同谷子品种中存在丰富的多态性,但REC和CCT结构域高度保守。SiTOC1编码区2种主要单倍型H-2和H-6分别与启动子单倍型Hp-591C和Hp-591A共分离,其中,启动子单倍型Hp-591C较Hp-591A的相对表达量显著上调了约2.5倍（P=0.014）,并且该单倍型在三亚市、长治市和乌鲁木齐市3个环境下的抽穗时间分别平均延迟9、11和12 d。【结论】SiTOC1启动子区第591 bp处的SNP是引起抽穗时间差异的主效位点,单倍型Hp-591A较Hp-591C早熟,可作为主效单倍型用于分子育种选择。     

芝麻产量相关性状的多位点全基因组关联分析及候选基因预测

【目的】通过对芝麻产量相关性状的全基因组关联分析,挖掘与产量性状关联的SNP位点及预测候选基因,为通过分子标记辅助选择育种等方式提高芝麻产量提供技术基础。【方法】以363份不同遗传背景和地理来源的芝麻种质资源构成的自然群体为研究对象,调查2年2点4环境下8个产量相关性状（单株产量、单株蒴数、蒴粒数、千粒重、株高、主茎果轴长、始蒴高度和表观收获指数）的表型值,借助覆盖全基因组的42 781个SNP标记,利用多位点SNP随机效应混合线性模型（multi-locus random-SNP-effect mixed linear model,mrMLM）对8个产量相关性状进行全基因组关联分析,检测与产量相关性状显著关联的SNP位点,并预测候选基因。【结果】在4个不同环境下,8个产量相关性状表现出广泛的表型变异,变异系数为6.51%—33.57%;相关性分析表明单株产量与单株蒴数、株高、主茎果轴长、表观收获指数呈极显著正相关;方差分析表明产量相关性状的基因型效应、环境效应、基因型与环境互作效应均达到了极显著水平。通过多位点全基因组关联分析共检测到210个与产量相关性状显著关联的SNP,在2018年南阳环境下检测到47个SNP,解释表型变异的1.63%—17.29%;在2019年南阳环境下检测到35个SNP,解释表型变异的1.94%—11.90%;在2018年平舆环境下检测到35个SNP,解释表型变异的2.15%—15.90%;在2019年平舆环境下检测到53个SNP,解释表型变异的1.25%—11.13%;在4个环境的综合BLUP条件下检测到75个SNP,解释表型变异的1.44%—13.58%。上述210个SNP涉及到175个位点,其中10个位点在3个及以上环境中被重复检测到。在这10个位点基因组区域内,共鉴定到214个候选基因,其中156个候选基因具有功能注释,主要涉及植物代谢、生物调控、生长发育等生物学过程。根据功能注释筛选出4个可能与芝麻产量相关的候选基因,其中SIN_1006338编码1-氨基环丙烷-1-羧酸合酶3（1-aminocyclopropane-1-carboxylate synthase 3-like）,参与乙烯的生物合成;SIN_1024330编码碱性螺旋-环-螺旋（basic helix-loop-helix）转录因子,负向调控植物细胞和器官的伸长;SIN_1014512编码吲哚-3-乙酸-酰胺合成酶GH3.6（indole-3-acetic acid-amido synthetase GH3.6）,参与调控茎和下胚轴细胞的伸长生长;SIN_1011473编码泛素受体蛋白DA1（protein DA1-like）,参与调节植物细胞增殖周期。【结论】通过多位点SNP随机效应混合线性模型的全基因组关联分析,检测到175个与产量相关性状显著关联的位点,筛选出4个可能与产量相关的重要候选基因。