棉花种间导入系的构建、新型标记的开发与高密度遗传图谱的构建
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摘要
棉花在生物学分类上属于锦葵科棉属(Gossypium),是重要的经济作物和最主要的纤维作物。随着社会的不断发展进步,高产优质棉花品种的培育日益重要。但是,传统育种方法的局限性使得较多性状的同时改良难以在短期内实现。随着分子生物学及生物技术的迅猛发展,分子育种技术与常规育种技术的结合或许可以作为更有效的手段来开展棉花高产优质新品种的选育。
本研究以分子标记技术为核心技术,主要开展了以下三方面的工作,以期为棉花育种提供育种资源和理论指导。(1)棉花海陆种间导入系的构建；(2)棉花新型分子标记的开发及遗传评价；(3)棉花海陆种间高密度遗传图谱的构建。
1.棉花海陆种间导入系的构建
本研究运用分子标记辅助选择的手段进行棉花海陆种间导入系的构建,利用在种间图谱上以10cM±为标准挑选的515个标记对每一世代单株进行代换片段的检测。截至2012年,337个单株入选,其中,82.79%的单株在陆地棉Emian22(受体)的遗传背景中仅含有三个甚至更少的海岛棉3-79(供体)的代换片段。总体说来,海岛棉3-79的供体片段较好地覆盖了棉花基因组,覆盖度为77.67%。但是,海岛棉3-79的供体片段对棉花26条染色体的覆盖度大小不一,从57.14%(Chr05)到100.00%(Chr23)变化不等。另外,单株所含代换片段的数目也不尽相同,从1到10变化不等。即将构建成功的导入系会大大促进棉花数量性状方面的相关研究,也能为棉花育种工作提供新的宝贵的育种资源。
2.棉花新型分子标记的开发及遗传评价
(1)棉花纤维发育相关功能标记的开发
本研究根据棉纤维发育相关功能基因的序列,共设计了331对纤维基因引物和164对蛋白引物。经SSCP分型技术分析发现,52对引物产生了58个多态性位点,引物多态性率仅为10.51%。连锁分析发现,代表48个基因或蛋白的53个多态性标记广泛分布于棉花21条染色体上,意味着棉纤维的发育应该是受棉花整个基因组协同调控的。另外,多态性引物相关序列的GO分析结果说明了棉花纤维发育生物进程的复杂性。多态性引物对海陆棉种纤维发育各时期进行了RT-PCR和qRT-PCR分析,基因的表达模式与前人的相关报道比较吻合,而且与本实验室对海陆棉种纤维品质性状的考察结果一致。海陆种间的表达差异分析可以为陆地棉品种纤维品质的改良提供一定的理论指导,主要可以通过转基因技术将在海岛棉中特异或优势表达而又与纤维品质密切相关的功能基因转移到大面积推广的高产陆地棉品种中,以期快速有效地培育高产优质的棉花新品种。
(2)棉花SNP和IDP标记的开发
本研究利用in-silico方法共设计合成了1349对棉花SNP/IDP引物,包括455对Ghi系列引物、356对HAU-SNP系列引物、415对GhIDP系列引物和123对PCtIDP系列引物。SSCP分型技术使得137对引物产生了142个多态性位点,引物多态性率为10.16%。连锁分析显示,133个多态性标记广泛分布于棉花整个基因组,与SNPs/IDPs遍布整个基因组的事实相符,而且对提高遗传图谱的密度非常有利；另外,该部分SNP和IDP标记还能很好地补充说明棉花偏分离的相关研究。碱基变异的规律统计分析发现,棉花碱基替换中存在转换偏奇现象,碱基转换占总变异的55.78%。而碱基替换的功能聚集结果分析显示,不同功能的SNPs功能聚集情况千差万别,对不同类别的相关功能基因的研究具有一定的指导意义。最后,序列分析的实验结果表明in-silico方法是开发棉花SNP标记的一把双刃剑,因为实际的SNPs并不总是与预测的SNPs吻合。
(3)棉花亚基因组特异标记的开发
本研究共设计开发了260对At亚组特异的Ga-Gh系列引物和545对Dt亚组特异的Gr-Gh系列引物。SSCP分型技术使得25对Ga-Gh系列引物和25对Gr-Gh系列引物产生了共50个多态性位点,引物多态性率分别为9.62%和4.59%,意味着海陆棉种间的差异可能更多地源于At亚组。连锁分析显示,24个Ga-Gh多态标记中的15个定位于At亚组,另外9个定位于Dt亚组；21个Gr-Gh多态标记中的15个定位于At亚组,另外6个定位于Dt亚组。实验结果与预期结果有着巨大偏差,原因可能有以下两点：一方面是因为开发引物时数据库中序列信息的匮乏而导致的所开发引物并不真的是亚基因组特异的引物；另一方面则很大程度上是因为棉花基因组的复杂性,主要是指At和Dt亚组中同源染色体的高度同源性／共线性。
3.棉花海陆种间高密度遗传图谱的构建
利用6%变性聚丙烯酰胺凝胶电泳和8%非变性聚丙烯酰胺凝胶电泳两种分析方法,对CMD(http://www.cottonmarker.org/)中新公布的SSR标记、文献中报道的少量新型标记及本研究中开发的三大类标记等共13507对引物进行亲本间的多态性检测和群体的基因型分析,最终有2498对引物在两个作图亲本间表现出多态性,并产生了2751个多态性位点,引物多态性率为18.49%。汇总本实验室张艳欣和余渝所得的2528个多态性标记和其他人mic-RNA、靶标、转录因子、抗病基因、细胞发育等相关的共353个多态性标记,最后用Joinmap3.0作图软件对总数为5632的多态性标记进行遗传连锁分析,最终构建了一张包含5152个多态性标记、图谱全长为4696.03cM、标记间平均距离为0.91cM的高密度棉花海陆种间遗传连锁图谱。
对异源四倍体棉花遗传图谱的26条染色体进行了同源染色体间的共线性分析。结果显示,At和Dt亚组的同源染色体间具有最高的同源性。此外,其中7个同源染色体对中的棉花染色体还与同亚组或非同亚组的非同源染色体间具有较高的同源性,说明异源四倍体棉花在进化过程中可能发生了染色体的倒位或易位。所以说,本研究不仅有助于揭示异源四倍体棉花复杂的基因组结构,还可能推动异源四倍体棉花进化相关方向的研究。
以该高密度遗传图谱为基础进行的四倍体棉花与雷蒙德氏棉(Dt亚组二倍体棉种)进行的同一物种不同亚种间的比较基因组学分析显示,在较高比对标准下,仍有69.57%的标记成功比对,四倍体棉花染色体中除了Chr03和Chr04没有与雷蒙德氏棉对应的同源染色体得到最高同源度外,其余24条染色体均是与其同源染色体有着最高的同源度。与可可的物种间比较基因组学分析显示,在相对较低的比对标准下,成功比对标记比例为46.55%,与可可染色体Tc01获得最高同源度的棉花染色体最多,有8条。与拟南芥进行的物种间比较基因组学分析显示,在与可可同样的比对条件下,成功比对标记比例为13.74%,与拟南芥染色体Atl获得最高同源度的棉花染色体最多,有12条。总之,本研究中以四倍体棉种遗传图谱为基础进行的比较基因组学分析,为利用雷蒙德氏棉、可可、拟南芥全基因组序列信息来分析异源四倍体棉花的复杂基因组提供了一定程度的启示和指导。
Cotton (Malvaceae Gossypium) is an important cash crop and the uppermost source of textile fiber. Along with the society's advance, cultivation of varieties with high yield and super fiber quality is becoming increasingly important. However, it is difficult for conventional breeding methods to synchronously improve several characters in a short time. With rapid development of molecular biology and biotechnology, combination of molecular breeding technology and conventional breeding technology may be an efficient approach to cotton breeding.
Laying molecular marker technology at the core, this study launched three parts of work aiming at providing germplasm resource and theoretical direction to cotton breeding:(1) construction of introgression lines in cotton;(2) development and evaluation of new markers in cotton;(3) construction of high-density genetic linkage map in cotton.
1. Construction of introgression lines in cotton
In this study, introgression lines were constructed using molecular marker-assisted selection, with substitution segments of plants checked by515markers evenly selected from the linkage map. Up to2012,82.79%of the total337plants harbored only three or less substitution segments. Totally speaking, substitution segments from the donor parent (3-79) comparatively well covered the cotton genome, with coverage of77.67%. However, coverage in different chromosomes varied from57.14%(Chr05) to100.00%(Chr23). In addition, numbers of substitution segments in different plants also varied greatly, ranging from1to10. It is expected that ILs constructed in this study will greatly accelerate research progress of quantitative traits, and will provide new valuable resource for cotton breeding.
2. Development and evaluation of new markers in cotton
(1) Development of functional markers related to cotton fiber development
A total of331gene primers and164protein primers were designed from functional sequences related to cotton fiber development. After genotyping using SSCP method,52primers (10.51%) showed polymorphism, and produced58polymorphic loci. After linkage analysis,53loci representing48genes or proteins were distributed randomly throughout the cotton genome, indicating that cotton fiber development was regulated by the whole genome. Besides, GO analysis of functional sequences also illustrated the complexity of fiber development. Both RT-PCR analysis and qRT-PCR analysis of polymorphic primers obtained similar expression tendencies with previous reports, and they also support field experimental results obtained in our laboratory. These expression patterns may provide some guidance to improve fiber quality of G. hirsutum, which could be accomplished by transforming those genes preferentially expressed in G. barbadense and controlling fiber quality obviously.
(2) Development of SNP and IDP markers in cotton
A total of1349SNP/IDP markers were developed using in-silico analysis in this study, including455Ghi-prefixed primers,356HAU-SNP-prefixed primers,415GhlDP-prefixed primers, and123PCtIDP-prefixed primers. After genotyping using SSCP analysis,137primers (10.16%) showed polymorphism, and produced142polymorphic loci. Genetic mapping showed that133polymorphic loci randomly distributed throughout the cotton genome, which is in accordance with the fact that SNPs/IDPs always pervade the whole genome, and indicates that SNP and IDP markers are valuable resource to increase density of linkage map. In addition, mapped SNPs and IDPs also could supplement previous research of segregation distortion. Statistic analysis of base substitutions showed that bias of base transitions exists in cotton genome, with the fact that base transitions occupied55.78%of the total SNPs. Functional accumulation analysis showed that numbers of SNPs in different functional categories varied obviously, which may provide some guidance to research of genes belonging to different functional categories. Generally speaking, in-silico analysis is a double-faced method to develop SNP markers in cotton revealed by sequence analysis, as the practical SNPs were not always in accordance with the predicted ones.
(3) Development of subgenome-specific markers in cotton
A total of260At-subgenome-specific primers and545Dt-subgenome-specific primers were developed in this study. SSCP analysis made25(9.62%) At-subgenome-specific primers and25(4.59%) Dt-subgenome-specific primers produce50polymorphic loci, indicating that difference between G. hirsutum and G. barbadense may mainly originate from the At-subgenome. After linkage analysis,15loci of the24mapped Ga-Gh-prefixed loci were located on the At-subgenome, and other9loci were located on the Dt-subgenome. As far as the21Gr-Gh-prefixed mapped loci,15of them were located on the At-subgenome, while only6loci were located on the Dt-subgenome. Reasons why experimental results deviated from anticipation tremendously are as follows. On the one hand, sequence shortage of cotton resulted in the fact that designed primers were not really specific to subgenome. On the other hand, unfavorable results may largely originate from complexity of the cotton genome-high homology/collinearity between homologous chromosomes of the At-subgenome and Dt-subgenome.
3. Construction of high density genetic linkage map in cotton
A total of13507primers, including new published SSRs in CMD (http://www.cottonmarker.org/), new kinds of markers reported by previous researchers, and three parts of primers designed in this study, were used to detect polymorphism between the two mapping parents using two genotyping methods. As a result,2498(18.49%) primers showed polymorphism, and produced2751polymorphic loci. Gathering the original2528polymorphic loci obtained by Yu (2011) and other353polymorphic loci related to mic-RNAs, targets, transcription factors, anti-disease genes, cell development and so on, a total of5632polymorphic loci were used for linkage analysis using Joinmap3.0. Finally, a high density interspecific linkage map including5152loci was constructed, with4696.03cM in total length and0.91cM in average distance between adjacent markers.
Colinearity analysis among cotton chromosomes showed that it harbored the highest value between homologous chromosomes. However, comparatively high colinearity appeared between non-homologous chromosomes in14chromosomes which are homologous chromosomes in pairs. It indicates that this study is useful to analyze complex genome of cotton, and is helpful to research evolution history of cotton.
Comparative analysis between allotetraploid cotton and diplontic cotton (G.raimondii) showed that most of the allotetraploid cotton chromosomes (except Chr03and Chr04) had the highest autoploidy with homologous chromosomes of the diplontic cotton. Comparative analysis between allotetraploid cotton and T. cocoa showed that percentage of successfully blasted markers was46.55%, and Tc01of T.cocoa was the most frequent chromosome which had the highest autoploidy with chromosomes of cotton. Comparative analysis between allotetraploid cotton and Arabidopsis showed that percentage of successfully blasted markers was13.74%, and Atl of Arabidopsis was the most frequent chromosome which had the highest autoploidy with chromosomes of cotton. Totally speaking, all above provides revelation and guidance to analysis of complex genome of allotetraploid cotton, through exploiting genome sequences of G.raimondii, T.cocoa and Arabidopsis.
本研究以分子标记技术为核心技术,主要开展了以下三方面的工作,以期为棉花育种提供育种资源和理论指导。(1)棉花海陆种间导入系的构建；(2)棉花新型分子标记的开发及遗传评价；(3)棉花海陆种间高密度遗传图谱的构建。
1.棉花海陆种间导入系的构建
本研究运用分子标记辅助选择的手段进行棉花海陆种间导入系的构建,利用在种间图谱上以10cM±为标准挑选的515个标记对每一世代单株进行代换片段的检测。截至2012年,337个单株入选,其中,82.79%的单株在陆地棉Emian22(受体)的遗传背景中仅含有三个甚至更少的海岛棉3-79(供体)的代换片段。总体说来,海岛棉3-79的供体片段较好地覆盖了棉花基因组,覆盖度为77.67%。但是,海岛棉3-79的供体片段对棉花26条染色体的覆盖度大小不一,从57.14%(Chr05)到100.00%(Chr23)变化不等。另外,单株所含代换片段的数目也不尽相同,从1到10变化不等。即将构建成功的导入系会大大促进棉花数量性状方面的相关研究,也能为棉花育种工作提供新的宝贵的育种资源。
2.棉花新型分子标记的开发及遗传评价
(1)棉花纤维发育相关功能标记的开发
本研究根据棉纤维发育相关功能基因的序列,共设计了331对纤维基因引物和164对蛋白引物。经SSCP分型技术分析发现,52对引物产生了58个多态性位点,引物多态性率仅为10.51%。连锁分析发现,代表48个基因或蛋白的53个多态性标记广泛分布于棉花21条染色体上,意味着棉纤维的发育应该是受棉花整个基因组协同调控的。另外,多态性引物相关序列的GO分析结果说明了棉花纤维发育生物进程的复杂性。多态性引物对海陆棉种纤维发育各时期进行了RT-PCR和qRT-PCR分析,基因的表达模式与前人的相关报道比较吻合,而且与本实验室对海陆棉种纤维品质性状的考察结果一致。海陆种间的表达差异分析可以为陆地棉品种纤维品质的改良提供一定的理论指导,主要可以通过转基因技术将在海岛棉中特异或优势表达而又与纤维品质密切相关的功能基因转移到大面积推广的高产陆地棉品种中,以期快速有效地培育高产优质的棉花新品种。
(2)棉花SNP和IDP标记的开发
本研究利用in-silico方法共设计合成了1349对棉花SNP/IDP引物,包括455对Ghi系列引物、356对HAU-SNP系列引物、415对GhIDP系列引物和123对PCtIDP系列引物。SSCP分型技术使得137对引物产生了142个多态性位点,引物多态性率为10.16%。连锁分析显示,133个多态性标记广泛分布于棉花整个基因组,与SNPs/IDPs遍布整个基因组的事实相符,而且对提高遗传图谱的密度非常有利；另外,该部分SNP和IDP标记还能很好地补充说明棉花偏分离的相关研究。碱基变异的规律统计分析发现,棉花碱基替换中存在转换偏奇现象,碱基转换占总变异的55.78%。而碱基替换的功能聚集结果分析显示,不同功能的SNPs功能聚集情况千差万别,对不同类别的相关功能基因的研究具有一定的指导意义。最后,序列分析的实验结果表明in-silico方法是开发棉花SNP标记的一把双刃剑,因为实际的SNPs并不总是与预测的SNPs吻合。
(3)棉花亚基因组特异标记的开发
本研究共设计开发了260对At亚组特异的Ga-Gh系列引物和545对Dt亚组特异的Gr-Gh系列引物。SSCP分型技术使得25对Ga-Gh系列引物和25对Gr-Gh系列引物产生了共50个多态性位点,引物多态性率分别为9.62%和4.59%,意味着海陆棉种间的差异可能更多地源于At亚组。连锁分析显示,24个Ga-Gh多态标记中的15个定位于At亚组,另外9个定位于Dt亚组；21个Gr-Gh多态标记中的15个定位于At亚组,另外6个定位于Dt亚组。实验结果与预期结果有着巨大偏差,原因可能有以下两点：一方面是因为开发引物时数据库中序列信息的匮乏而导致的所开发引物并不真的是亚基因组特异的引物；另一方面则很大程度上是因为棉花基因组的复杂性,主要是指At和Dt亚组中同源染色体的高度同源性／共线性。
3.棉花海陆种间高密度遗传图谱的构建
利用6%变性聚丙烯酰胺凝胶电泳和8%非变性聚丙烯酰胺凝胶电泳两种分析方法,对CMD(http://www.cottonmarker.org/)中新公布的SSR标记、文献中报道的少量新型标记及本研究中开发的三大类标记等共13507对引物进行亲本间的多态性检测和群体的基因型分析,最终有2498对引物在两个作图亲本间表现出多态性,并产生了2751个多态性位点,引物多态性率为18.49%。汇总本实验室张艳欣和余渝所得的2528个多态性标记和其他人mic-RNA、靶标、转录因子、抗病基因、细胞发育等相关的共353个多态性标记,最后用Joinmap3.0作图软件对总数为5632的多态性标记进行遗传连锁分析,最终构建了一张包含5152个多态性标记、图谱全长为4696.03cM、标记间平均距离为0.91cM的高密度棉花海陆种间遗传连锁图谱。
对异源四倍体棉花遗传图谱的26条染色体进行了同源染色体间的共线性分析。结果显示,At和Dt亚组的同源染色体间具有最高的同源性。此外,其中7个同源染色体对中的棉花染色体还与同亚组或非同亚组的非同源染色体间具有较高的同源性,说明异源四倍体棉花在进化过程中可能发生了染色体的倒位或易位。所以说,本研究不仅有助于揭示异源四倍体棉花复杂的基因组结构,还可能推动异源四倍体棉花进化相关方向的研究。
以该高密度遗传图谱为基础进行的四倍体棉花与雷蒙德氏棉(Dt亚组二倍体棉种)进行的同一物种不同亚种间的比较基因组学分析显示,在较高比对标准下,仍有69.57%的标记成功比对,四倍体棉花染色体中除了Chr03和Chr04没有与雷蒙德氏棉对应的同源染色体得到最高同源度外,其余24条染色体均是与其同源染色体有着最高的同源度。与可可的物种间比较基因组学分析显示,在相对较低的比对标准下,成功比对标记比例为46.55%,与可可染色体Tc01获得最高同源度的棉花染色体最多,有8条。与拟南芥进行的物种间比较基因组学分析显示,在与可可同样的比对条件下,成功比对标记比例为13.74%,与拟南芥染色体Atl获得最高同源度的棉花染色体最多,有12条。总之,本研究中以四倍体棉种遗传图谱为基础进行的比较基因组学分析,为利用雷蒙德氏棉、可可、拟南芥全基因组序列信息来分析异源四倍体棉花的复杂基因组提供了一定程度的启示和指导。
Cotton (Malvaceae Gossypium) is an important cash crop and the uppermost source of textile fiber. Along with the society's advance, cultivation of varieties with high yield and super fiber quality is becoming increasingly important. However, it is difficult for conventional breeding methods to synchronously improve several characters in a short time. With rapid development of molecular biology and biotechnology, combination of molecular breeding technology and conventional breeding technology may be an efficient approach to cotton breeding.
Laying molecular marker technology at the core, this study launched three parts of work aiming at providing germplasm resource and theoretical direction to cotton breeding:(1) construction of introgression lines in cotton;(2) development and evaluation of new markers in cotton;(3) construction of high-density genetic linkage map in cotton.
1. Construction of introgression lines in cotton
In this study, introgression lines were constructed using molecular marker-assisted selection, with substitution segments of plants checked by515markers evenly selected from the linkage map. Up to2012,82.79%of the total337plants harbored only three or less substitution segments. Totally speaking, substitution segments from the donor parent (3-79) comparatively well covered the cotton genome, with coverage of77.67%. However, coverage in different chromosomes varied from57.14%(Chr05) to100.00%(Chr23). In addition, numbers of substitution segments in different plants also varied greatly, ranging from1to10. It is expected that ILs constructed in this study will greatly accelerate research progress of quantitative traits, and will provide new valuable resource for cotton breeding.
2. Development and evaluation of new markers in cotton
(1) Development of functional markers related to cotton fiber development
A total of331gene primers and164protein primers were designed from functional sequences related to cotton fiber development. After genotyping using SSCP method,52primers (10.51%) showed polymorphism, and produced58polymorphic loci. After linkage analysis,53loci representing48genes or proteins were distributed randomly throughout the cotton genome, indicating that cotton fiber development was regulated by the whole genome. Besides, GO analysis of functional sequences also illustrated the complexity of fiber development. Both RT-PCR analysis and qRT-PCR analysis of polymorphic primers obtained similar expression tendencies with previous reports, and they also support field experimental results obtained in our laboratory. These expression patterns may provide some guidance to improve fiber quality of G. hirsutum, which could be accomplished by transforming those genes preferentially expressed in G. barbadense and controlling fiber quality obviously.
(2) Development of SNP and IDP markers in cotton
A total of1349SNP/IDP markers were developed using in-silico analysis in this study, including455Ghi-prefixed primers,356HAU-SNP-prefixed primers,415GhlDP-prefixed primers, and123PCtIDP-prefixed primers. After genotyping using SSCP analysis,137primers (10.16%) showed polymorphism, and produced142polymorphic loci. Genetic mapping showed that133polymorphic loci randomly distributed throughout the cotton genome, which is in accordance with the fact that SNPs/IDPs always pervade the whole genome, and indicates that SNP and IDP markers are valuable resource to increase density of linkage map. In addition, mapped SNPs and IDPs also could supplement previous research of segregation distortion. Statistic analysis of base substitutions showed that bias of base transitions exists in cotton genome, with the fact that base transitions occupied55.78%of the total SNPs. Functional accumulation analysis showed that numbers of SNPs in different functional categories varied obviously, which may provide some guidance to research of genes belonging to different functional categories. Generally speaking, in-silico analysis is a double-faced method to develop SNP markers in cotton revealed by sequence analysis, as the practical SNPs were not always in accordance with the predicted ones.
(3) Development of subgenome-specific markers in cotton
A total of260At-subgenome-specific primers and545Dt-subgenome-specific primers were developed in this study. SSCP analysis made25(9.62%) At-subgenome-specific primers and25(4.59%) Dt-subgenome-specific primers produce50polymorphic loci, indicating that difference between G. hirsutum and G. barbadense may mainly originate from the At-subgenome. After linkage analysis,15loci of the24mapped Ga-Gh-prefixed loci were located on the At-subgenome, and other9loci were located on the Dt-subgenome. As far as the21Gr-Gh-prefixed mapped loci,15of them were located on the At-subgenome, while only6loci were located on the Dt-subgenome. Reasons why experimental results deviated from anticipation tremendously are as follows. On the one hand, sequence shortage of cotton resulted in the fact that designed primers were not really specific to subgenome. On the other hand, unfavorable results may largely originate from complexity of the cotton genome-high homology/collinearity between homologous chromosomes of the At-subgenome and Dt-subgenome.
3. Construction of high density genetic linkage map in cotton
A total of13507primers, including new published SSRs in CMD (http://www.cottonmarker.org/), new kinds of markers reported by previous researchers, and three parts of primers designed in this study, were used to detect polymorphism between the two mapping parents using two genotyping methods. As a result,2498(18.49%) primers showed polymorphism, and produced2751polymorphic loci. Gathering the original2528polymorphic loci obtained by Yu (2011) and other353polymorphic loci related to mic-RNAs, targets, transcription factors, anti-disease genes, cell development and so on, a total of5632polymorphic loci were used for linkage analysis using Joinmap3.0. Finally, a high density interspecific linkage map including5152loci was constructed, with4696.03cM in total length and0.91cM in average distance between adjacent markers.
Colinearity analysis among cotton chromosomes showed that it harbored the highest value between homologous chromosomes. However, comparatively high colinearity appeared between non-homologous chromosomes in14chromosomes which are homologous chromosomes in pairs. It indicates that this study is useful to analyze complex genome of cotton, and is helpful to research evolution history of cotton.
Comparative analysis between allotetraploid cotton and diplontic cotton (G.raimondii) showed that most of the allotetraploid cotton chromosomes (except Chr03and Chr04) had the highest autoploidy with homologous chromosomes of the diplontic cotton. Comparative analysis between allotetraploid cotton and T. cocoa showed that percentage of successfully blasted markers was46.55%, and Tc01of T.cocoa was the most frequent chromosome which had the highest autoploidy with chromosomes of cotton. Comparative analysis between allotetraploid cotton and Arabidopsis showed that percentage of successfully blasted markers was13.74%, and Atl of Arabidopsis was the most frequent chromosome which had the highest autoploidy with chromosomes of cotton. Totally speaking, all above provides revelation and guidance to analysis of complex genome of allotetraploid cotton, through exploiting genome sequences of G.raimondii, T.cocoa and Arabidopsis.
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