Insertion mutagenesis has become one of the most popular methods for gene functions analysis.Here we report a two-element Ac/Ds transposon system containing enhancer trap and gene trap for gene tagging in rice.The exc...Insertion mutagenesis has become one of the most popular methods for gene functions analysis.Here we report a two-element Ac/Ds transposon system containing enhancer trap and gene trap for gene tagging in rice.The excision of Ds element was examined by PCR amplification.The excision frequency of Ds element varied from 0% to 40% among 20 F2 populations derived from 11 different Ds parents.Southern blot analysis revealed that more than 70% of excised Ds elements reinserted into rice genome and above 70% of the reinserted Ds elements were located at different positions of the chromosome in rice.The result of histochemical GUS analysis indicated that 28% of enhancer trap and 22% of gene trap tagging plants displayed GUS activity in leaves, roots,flowers or seeds.The GUS positive lines will be useful for identifying gene function in rice.展开更多
The Agrobacterium mediated transgenic rice ( Oryza saliva L.) population with inserts of maize transposon Activator/Dissociation (Ac/Ds) was investigated. DNA sequences flanking the T-DNA were analyzed with inverse PC...The Agrobacterium mediated transgenic rice ( Oryza saliva L.) population with inserts of maize transposon Activator/Dissociation (Ac/Ds) was investigated. DNA sequences flanking the T-DNA were analyzed with inverse PCR. Results showed that 65.4% of the T-DNA was integrated in different locations of rice genome, and some T-DNA flanking sequences were located on certain chromosomes. A number of T-DNA was found to have inserted into protein coding regions. In order to induce transposition of the inserted Ds elements, 354 crosses of Ac x Ds and Ds x Ac were constructed. The excision frequency of Ds element trans-activated by Ac transposase was 22.7% in the F-2 populations, and the transposition was confirmed with analyses of DNA sequences flanking the Ds elements. In addition to the transposition due to 'cut-paste' mechanism, Ds can replicate itself and integrate into a new locus, and inaccurate excisions were also found. A proportion of DNA segments flanking the Ds elements showed no homologies to sequences published in GenBank, of which two were registered under the accession numbers AF355153 and AF355770. The strategy of using transposon tagging for rice genomics study was discussed.展开更多
文摘Insertion mutagenesis has become one of the most popular methods for gene functions analysis.Here we report a two-element Ac/Ds transposon system containing enhancer trap and gene trap for gene tagging in rice.The excision of Ds element was examined by PCR amplification.The excision frequency of Ds element varied from 0% to 40% among 20 F2 populations derived from 11 different Ds parents.Southern blot analysis revealed that more than 70% of excised Ds elements reinserted into rice genome and above 70% of the reinserted Ds elements were located at different positions of the chromosome in rice.The result of histochemical GUS analysis indicated that 28% of enhancer trap and 22% of gene trap tagging plants displayed GUS activity in leaves, roots,flowers or seeds.The GUS positive lines will be useful for identifying gene function in rice.
文摘The Agrobacterium mediated transgenic rice ( Oryza saliva L.) population with inserts of maize transposon Activator/Dissociation (Ac/Ds) was investigated. DNA sequences flanking the T-DNA were analyzed with inverse PCR. Results showed that 65.4% of the T-DNA was integrated in different locations of rice genome, and some T-DNA flanking sequences were located on certain chromosomes. A number of T-DNA was found to have inserted into protein coding regions. In order to induce transposition of the inserted Ds elements, 354 crosses of Ac x Ds and Ds x Ac were constructed. The excision frequency of Ds element trans-activated by Ac transposase was 22.7% in the F-2 populations, and the transposition was confirmed with analyses of DNA sequences flanking the Ds elements. In addition to the transposition due to 'cut-paste' mechanism, Ds can replicate itself and integrate into a new locus, and inaccurate excisions were also found. A proportion of DNA segments flanking the Ds elements showed no homologies to sequences published in GenBank, of which two were registered under the accession numbers AF355153 and AF355770. The strategy of using transposon tagging for rice genomics study was discussed.
