Study On Gene Flow From Transgenic Rice T1c-19 With Stacked Crylc*/Bar Genes To Weedy Rice And The Fitness Of Hybrids Betweem Them

Releasing of transgenic rice(Oryza sativa L.)could produce economic and social benefits,however,risk of transgene flow into weedy rice,which may induce weedy rice with transgenic resistant gene,is of major concern.Compared to mono-trait crop varieties,stacked transgenic crops may be more prone to become weeds due to the additional advantage traits that could confer advantage under certain environment conditions.Thus,safety of stacked transgenic rice should be evaluated more comprehensively.Transgenic rice Tlc-19 with stacked Cry1C*/Bar genes was cultivated by Huazhong Agricultural University.It was obtained by transforming Minghui 63,an elite CMS-restorer line used in the production of several widely grown rice hybrids,with both the insect resistance Cry1C*gene and the glufosinate resistance Bar gene.T1c-19 displayed a high level of resistance to its target insects and glufosinate,and did not significantly affect non-target organisms.T1c-19 has potential to gain regulatory approval.Hence,the current study concentrated on gene flow from T1c-19 to weedy rice and the fitness of hybrids between them under differnet environment.The results derived from this study should be useful to regulatory authorities in their risk assessment and decision making related to authorizing the release of this stacked transgenic rice.Gene flow from Tlc-19 to three weedy rice(O.sativa)collected from Maoming Guangdong(WRMM),Taizhou Jiangsu(WRTZ)and Yiyang Hunan(WRYY),and the receptor rice(O.sativaL.)MH63 was researched using two experimental designs(alternating and adjacent cultivations).It was shown that the maximum gene flow frequencies from Tlc-19 to WRYY was 0.164%and 0.230%,to WRTZ was 0.106%and 0.211%when they were planted in alternating and adjacent cultivation,respectively.However,there was no gene flow detected from T1c-19 to WRMM in both cultivation conditions.Gene flow frequency from Tlc-19 to the reciptor rice of MH63 was 0.919%in the alternating cultivation experiment.Overall,gene flow frequency had significant positive correlation with flowering overlap,and significant negative correlation with plant height(p<0.05).The gene flow occurred from Tlc-19 to WRYY with high frequency and WRTZ with low frequency.RF1+ was obtained using Tlc-19 as maternal plants and WRMM,WRTZ and WRYY as paternal plants.RF1-was obtained using MH63 as maternal plants and three weedy rices as paternal plants.Composite fitness of RF1+ and RF1-under two insect pressures(18.4%natural and no insect pressure)and two herbicides(405 g ai/L glufonisate and 18.75 g ai/ha Ricer)were evaluated.The results demonstrated that,under natutural insect pressure,composite fitness of three types of RF1+ were similar compared with their respective RF1-,while greater than their respective weedy rices due to 39-73%more tillers and 2-3 times greater yield per plant.Under no insect pressure,RF1MM+ and RF1TZ+ had similar composite fitness,while RF1YY+ had lower one compared with their respective RF1-.Compared with respective weedy rice,RF1TZ+ performed greater composite fitness due to 60%more full gains per panicle and 1.3 times greater yield per plant,and the composite fitness of RF1MM+ and RF1YY+ was similar with that of WRMM and WRYY.Furthermore,three types of RF1+ performed similar under glufosinate and Ricer application.The seed shattering of three types of RF1+ was similar with their respective RF1-,but significantly lower than their respective weedy rice parents.The above results demonstrated that of all RF1+ had greater composite fitness under insect pressure,and had greater or similar composite fitness compared to their respective weedy rice without insect pressure.In most cases,RF1+ had similar composite fitness to their respective RF1-.Therefore,gene flow from weedy rice to transgenic rice should be paid more attention during releasing stacked transgenic rice T1c-19 due to similar composite fitness of RF1+with respective weedy rice.F2 hybrids were obtain by self-polliantion of F1 hybrids.The transmission of insect and glufosinate resistant genes from F1 to F2 was tested by seed bioassay and PCR test.Fitness of F2+ and RF2+ under two level of insect pressure(16.3%-20.3%natural and no insect pressure),two herbicides(405 g ai/L glufosinate and 18.75 g ai/ha Ricer application)and two planting methods(pure and mixed planting(F2+/RF2+:WR = 1:1))were researched.The results showed that the segregation for insect and glufosinate resistance from F1+ to F2+ and from RF1+ to RF2+ fitted 3:1 Mendelian segregation.Research on fitness showed that,compared to F2-and RF2-,majority F2+ and RF2+ performed similar composite fitness undernatural or no insect pressure,except for F2YY+,which performed greater composite fitness than F2YY-under natural insect pressure.However,compared to weedy rice parents,in some cases,F2+ or RF2+ performed greater composite fitness than their weedy rice parents,respectively,including F2MM+ and RF2MM+ in pure planting with natural insect pressure,RF2TZ+ in mixed planting under natural insect pressure,RF2MM+ in pure planting without insect pressure,F2TZ+ and RF2TZ+ in mixed planting without insect pressure,whilein other cases,F2+ and RF2+ performed similar with their weedy rice parents.The present research showed that the stacked resistant transgenes could inherit steadily in the hybrids once gene exchanged between Tlc-19 and weedy rice.Overall,F2+ and RF2+ performed similar or greater composite fitness than F2-and RF2-under natural or no insect pressure,and performed greater or similar fitness with their weedy rice parents under natural or no insect pressure,in pure or mixed planting.Stacked Cry1C*/Bar transgenes could flow from Tlc-19 to weedy rice,and F1 and F2 hybrids with Cry1C*/Bar genes performed similar or greater fitness than the respective hybrids with receptor rice as parent and weedy rice parents in agro-ecosystem.Therefore,some measures should be adopted to prevent gene exchange between Tlc-19 and weedy rice during releasing stacked transgenic rice Tlc-19.Furthermore,the present research implied that the fitness of hybrids with stacked CrylC*/Bar genes was closely related to the insect pressure and genotype of weedy rice.