甘蓝型油菜超高密度遗传图谱整合 Solexa 序列
Integration of Solexa sequences on an ultradense genetic map in Brassica rapa L.
机构信息
College of Agronomy, Northwest A&F University, Yangling, Shaanxi, China.
出版信息
BMC Genomics. 2011 May 19;12:249. doi: 10.1186/1471-2164-12-249.
BACKGROUND
Sequence related amplified polymorphism (SRAP) is commonly used to construct high density genetic maps, map genes and QTL of important agronomic traits in crops and perform genetic diversity analysis without knowing sequence information. To combine next generation sequencing technology with SRAP, Illumina's Solexa sequencing was used to sequence tagged SRAP PCR products.
RESULTS
Three sets of SRAP primers and three sets of tagging primers were used in 77,568 SRAP PCR reactions and the same number of tagging PCR reactions respectively to produce a pooled sample for Illumina's Solexa sequencing. After sequencing, 1.28 GB of sequence with over 13 million paired-end sequences was obtained and used to match Solexa sequences with their corresponding SRAP markers and to integrate Solexa sequences on an ultradense genetic map. The ultradense genetic bin map with 465 bins was constructed using a recombinant inbred (RI) line mapping population in B. rapa. For this ultradense genetic bin map, 9,177 SRAP markers, 1,737 integrated unique Solexa paired-end sequences and 46 SSR markers representing 10,960 independent genetic loci were assembled and 141 unique Solexa paired-end sequences were matched with their corresponding SRAP markers. The genetic map in B. rapa was aligned with the previous ultradense genetic map in B. napus through common SRAP markers in these two species. Additionally, SSR markers were used to perform alignment of the current genetic map with other five genetic maps in B. rapa and B. napus.
CONCLUSION
We used SRAP to construct an ultradense genetic map with 10,960 independent genetic loci in B. rapa that is the most saturated genetic map ever constructed in this species. Using next generation sequencing, we integrated 1,878 Solexa sequences on the genetic map. These integrated sequences will be used to assemble the scaffolds in the B. rapa genome. Additionally, this genetic map may be used for gene cloning and marker development in B. rapa and B. napus.
背景
序列相关扩增多态性（SRAP）常用于构建作物高密度遗传图谱、定位重要农艺性状的基因和 QTL，并进行遗传多样性分析，而无需了解序列信息。为了将下一代测序技术与 SRAP 相结合，我们使用 Illumina 的 Solexa 测序技术对 SRAP-PCR 产物进行测序标记。
结果
我们分别用 3 组 SRAP 引物和 3 组标记引物对 77568 次 SRAP-PCR 反应和相同数量的标记 PCR 反应进行组合，产生一个混合样本进行 Illumina 的 Solexa 测序。测序后，我们获得了 12.8GB 的序列，包含超过 1300 万对的末端序列，用于将 Solexa 序列与对应的 SRAP 标记进行匹配，并将 Solexa 序列整合到超高密度遗传图谱上。我们利用白菜重组自交系（RI）作图群体构建了一个具有 465 个-bin 的超高密度遗传 bin 图谱。对于这个超高密度遗传 bin 图谱，我们组装了 9177 个 SRAP 标记、1737 个整合的独特 Solexa 配对末端序列和 46 个 SSR 标记，代表 10960 个独立的遗传位点，同时有 141 个独特的 Solexa 配对末端序列与对应的 SRAP 标记相匹配。通过这两个物种中共同的 SRAP 标记，我们将白菜的遗传图谱与之前在油菜中构建的超高密度遗传图谱进行了比对。此外，我们还使用 SSR 标记将当前遗传图谱与白菜和油菜的另外五个遗传图谱进行了比对。
结论
我们使用 SRAP 在白菜中构建了一个具有 10960 个独立遗传位点的超高密度遗传图谱，这是该物种中构建的最饱和的遗传图谱。我们使用下一代测序技术将 1878 个 Solexa 序列整合到遗传图谱上。这些整合的序列将用于组装白菜基因组的支架。此外，这个遗传图谱可用于白菜和油菜的基因克隆和标记开发。
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