Construction and characterization of the transformation-competent artificial chromosome (TAC) libraries of Leymus multicaulis
Construction and characterization of the transformation-competent artificial chromosome (TAC) libraries of Leymus multicaulis
复制标题
羊草具有转化能力的人工染色体（TAC）文库的构建和表征
DOI：
10.1007/s11427-008-0079-8
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发表时间：
2008-07
期刊：
影响因子：
--
通讯作者：
Zhou YuLei
中科院分区：
文献类型：
--
作者：
Xu YueYu;Zhao MaoLin;Song LinLin;Zhang Yan;Zhou YuLei
具有转化能力的人工染色体系统能够在植物中有效地克隆和转移基因。为了从羊草中克隆对大麦黄矮病毒(BYDV)、蚜虫、干旱和盐具有高度耐受性的基因，在载体pYLTAC17和pYLTAC747H/sacB中构建了两个TAC基因组文库I和II，分别包含约165000和236000个重组克隆。由于基因组 DNA 片段的大小选择约为 9 至 300 kb，因此两个文库的基因组覆盖率总体估计约为 3-5 个单倍体基因组当量。基因组文库的克隆被收集为大量池，每个池包含 500 个左右的克隆，储存在 12 个 96 深孔板中，然后在手动转移到三个 384 孔板中后，使用 GeneTACtrade mark G3 阵列机器人将其一式三份地网格化到具有 3x3 模式的高密度菌落杂交过滤器上。同时，pYLTAC17 和 pYLTAC747H/sacB 中的 2501 和 2890 个文库克隆分别储存在 14 个 384 孔板中，然后在板复制后以 6x6 模式自动一式两份地网格化到高密度菌落杂交过滤器上。利用多茎乳杆菌谷胱甘肽还原酶基因探针检测到19个阳性克隆。这里构建的TAC文库可用于分离含有靶基因的基因组克隆，并进行基因组步移以进行定位克隆。一旦分离出目标 TAC 克隆，就可以立即使用农杆菌系统将它们转移到植物基因组中。
Transformation-competent artificial chromosome system is able to clone and transfer genes efficiently in plants. In order to clone genes highly tolerant to barley yellow dwarf virus (BYDV), Aphids, drought and salt from Leymus multicaulis, the two TAC genomic libraries I and II were constructed in vector pYLTAC17 and pYLTAC747H/sacB, which contain about 165000 and 236000 recombinant clones separately. The genome coverage of the two libraries was totally estimated to be about 3-5 haploid genome equivalents, as size selection of genomic DNA fragments was approximately from 9 to 300 kb. Clones of the genomic libraries were collected as bulked pools each containing 500 clones or so, stored in twelve 96-deep-well plates and then were gridding in triplicate onto a high-density colony hybridization filter with a 3x3 pattern using a GeneTACtrade mark G3 arraying robot after being transferred manually into three 384-well plates. Meanwhile 2501 and 2890 clones of Library in pYLTAC17 and in pYLTAC747H/sacB were stored individually in fourteen 384-well plates and then were automatically gridding in duplicate onto a high-density colony hybridization filter with a 6x6 pattern after a replication of plates. Nineteen positive clones were detected by using the probe glutahione reductase gene of L. multicaulis. TAC libraries constructed here can be used to isolate genomic clones containing target genes, and to carry out genome walking for positional cloning. Once the target TAC clones were isolated, they could be immediately transferred into plant genomes with the Agrobacterium system.
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影响因子：
7.2
作者：
Liu, YG;Nagaki, K;Ogihara, Y
通讯作者：
Ogihara, Y
DOI：
10.1073/pnas.0931425100
发表时间：
2003-05-13
影响因子：
11.1
作者：
Lin, L;Liu, YG;Li, BJ
通讯作者：
Li, BJ
影响因子：
3.1
作者：
S. Qu;G. Coaker;D. Francis;B. Zhou;Gl Wang
通讯作者：
S. Qu;G. Coaker;D. Francis;B. Zhou;Gl Wang
DOI：
--
发表时间：
2002
期刊：
Yi chuan xue bao = Acta genetica Sinica
影响因子：
--
作者：
Xiao-Ping Wang;Zengyan Zhang;Qunyu Zhang;Yaoguang Liu;Z. Xin
通讯作者：
Xiao-Ping Wang;Zengyan Zhang;Qunyu Zhang;Yaoguang Liu;Z. Xin
DOI：
--
发表时间：
2002-05
期刊：
Sheng wu gong cheng xue bao = Chinese journal of biotechnology
影响因子：
--
作者：
Genji Qin;Pei-du Chen;Yaoguang Liu;Y. Fang;Dajun Liu
通讯作者：
Genji Qin;Pei-du Chen;Yaoguang Liu;Y. Fang;Dajun Liu