| Oilseed rape(Brassica napus,AACC,2n=4x=38)shows the characteristics of long flowering time.Its pollen can be spread for long distance and survive for long time.Wild Brassica juncea(AABB,2n=4x=36),as the relative weed of oilseed rape in China,has the same A-genome with B.napus.Therefore,it is important to research the gene flow and introgression of transgene from transgenic herbicide-resistant B.napus to wild B.juncea.The likelihood of the introgression of transgene will be determined by the long-term persistence of transgene in subsequent generations and the fitness of these generations.The fitness is highly depended on the chromosome constitutions.If the heterologous chromosomes exist between transgenic crops and their relative species,the chromosomes of transgenic crops can be replaced or be added to their relative species during hybridization or backcross,resulting in cytological instability and genetic imbalance.That will significantly reduce the fitness of hybrids.It is supposed that the transgene located on the C-geneome of B.napus should be transmitted at a low frequency to the related species without the C-geneome,but there is no empiric evidence exists yet.Our previous studies found that the forward and reverse backcross generations of glufosinate resistant B.napus(resistance gene is located on the C genome)and wild B.juncea displayed resistance gene silence.It was found that there were three types of plants: plants carrying and expressing the transgene(Resistance-expression)(RE),plants carrying the transgene,but not expressing resistance(Resistance-silence)(RS)and plants without transgene(N)(The first generation of resistance segregation);and then the above three types of plants were segregated from the progeny of resistance-silence plants(The second generation of resistance segregation)(Zhang,2017).The RE BC1F2-6,BC2F2-5 and BC3F2-5 were all mixoploid.As the generations advanced,the chromosomes were stable in the type of 2n=37(AABB + C)(Yan,2016;Bian,2018;Wang,2019).However,the chromosome behavior of the resistance-silence plants in the first generation of resistance segregation and the resistance-expression and silence plants in the second generation of resistance segregation in high generation were not analyzed.To explore whether the transgene of herbicide-resistant oilseed rape(transgene pat located on C chromosome)has been successfully introgressed to the wild B.juncea,the high generation of RE BC1F7 and RS BC1F7 and the progenies of RE BC1F8 and RS BC1F8(the RE BC1F7 progenies: RE BC1F8 and RS BC1F8,the RS BC1F7 progenies: RE BC1F8 and RS BC1F8)of glufosinate-resistant B.napus×wild B.juncea were used as materials.The chromosome numbers and constitution of plants were observed,and the variation of the chromosomes number was analyzed.The meiotic behavior of the RE BC1F7 and RS BC1F7 was observed.The chromosome karyotype of 2n=36 cells and the fitness in all types of progenies was also analyzed.1.A large proportion cell of the resistance-expression BC1F7 and BC1F8 have 2n=37(AABB+C)The RE BC1F7 and RE BC1F8 were mixoploids with a large proportion having 2n=37 chromosomes.The RE BC1F8 from the RE BC1F7 displayed a higher probability to carry 37 chromosomes compared to the RE BC1F7,but the RE BC1F8 from the RS BC1F7 did not displayed significance difference with RE BC1F7.The genomic constitutions of the 37-chromosome cells of the RE BC1F7 was 20A+16B+1C,indicating the RE BC1F7 carried one additional oilseed rape C-chromosome.2.A large proportion cell of the resistance-silence BC1F7 and BC1F8 have 2n=36(AABB)The RS BC1F7 and RS BC1F8 were mixoploids with a large proportion having 2n=36 chromosomes.The RS BC1 p F8 from the RE BC1F7 displayed a higher probability to carry 36 chromosomes compared to the RS BC1F7,but the RS BC1 m F8 did not displayed significance difference with RS BC1F7.The RS BC1F8 from the RS BC1F7 displayed a higher probability to carry 36 chromosomes compared to the RS BC1F7,without the cell 2n=37.The genomic constitutions of the 36-chromosome cells of the RS BC1F7 was 20A+16B.3.Resistance-silence BC1F7 showing higher inheritance stability than the resistance-expression BC1F7 during MeiosisAbnormal meiosis behavior in pollen mother cell(PMC),including asynchronous meiosis,lagging chromosomes,chromatid bridge and micronucleus was observed in the RE and RS BC1F7.Meiotic indices of the RE BC1 m F7 and RE BC1 p F7 were among 81.33 to 99.07,which indicated that they were genetically unstable.The values for the RS BC1 m F7 and RS BC1 p F7 were from 91.62 to 97.21,which indicates inheritance stability.4.The karyotypes of the resistance-expression BC1F7 and BC1F8 differed from the resistance-silence BC1F7 and BC1F8The difference of 2n=36 chromosome karyotypes of the RE BC1 m F7 and RE BC1 p F7 differed from the RS BC1 m F7 and RS BC1 p F7 on their centromeric location.The chromosome of the RE BC1 m F7 and RE BC1 p F7 were median point chromosome,while the RS BC1 m F7 and RS BC1 p F7 had two and one pairs of submedian region chromosome,respecitively.The difference between the RE BC1F8 and RS BC1F8 also display differed on the centromeric location.These results indicated that the pairing of heterologous chromosome in backcross progenies between glufosinate-resistant B.napus and wild B.juncea was disordered,suggesting that the way of heterologous recombination of chromosomes differed between the RE and RS progenies.This could be an important reason for the silencing of resistance genes.5.The fitness of BC1F8 from the resistance-expression BC1F7 was lower than the BC1F8 from the resistance-silence BC1F7The results showed that the plant height of the RE BC1F7 and RS BC1F7 were significantly lower than the wild B.juncea,while no significant differences in other fitness components were observed.The plant height of the RE BC1F8 and RS BC1F8 from the RE BC1F7 were significantly lower than the wild B.juncea,while no significant differences in other fitness components were observed.All fitness components of the RE BC1F8 and RS BC1F8 from the RS BC1F7 showed similarities to the wild B.juncea.The results confirmed that the RE BC1F7 and RE BC1F8 showed lower fitness than RS BC1F7 and RS BC1F8 due to it carried one additional oilseed rape C-chromosomes,leading to the lower genetic stability and fitness.In summary,the high generation RE and RS BC1F7 and BC1F8 of glufosinate-resistant B.napus×wild B.juncea were all mixoploid.The cells of the RE BC1F7 and RE BC1F8 with 2n≧37 showed a higher probability to carry C chromosome,while the cells of 2n≧37 almost did not found in the RS BC1F7 and RS BC1F8.The results of karyotype analysis showed the differences in chromosome karyotypes between the RE and RS BC1F7 and BC1F8.The above results indicated that the difference of chromosome composition between RE and RS BC1F7 and BC1F8 is directly related to resistance gene silence. |
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