| Cotton is a widely grown commercial crop in the world.With the increasing demand and innovation of textile industry,cotton production needs new cotton varieties with high yield and excellent fiber quality.Therefore,improving yield and fiber quality is the longterm goal of cotton breeders.Genetic modification provides a powerful tool for improving cotton fiber traits.There are many genetic engineering effects that have significantly improved the fiber quality traits of cotton.It is a very effective mean to improve fiber quality by regulating actin.Enhance the recombination or aggregation of F-actin(downregulate GhADF1 or overexpress genes such as GhWLIM1 a,GhFIM2,and RABA4 c etc.),resulting in longer fibers in transgenic cotton;Improve fiber strength and fineness,as well as lint yield.Sucrose is one of the indispensable substances in the development of cotton fibers,and transgenic cotton overexpressing sucrose synthase genes(GhSusA1,StSUS,ZmSUS1,etc.)significantly increases fiber length and strength.In addition,there are many genes that can improve fiber quality.Various plant hormones such as indoleacetic acid,gibberellin,and brassinosteroids can promote the initiation and elongation of cotton fiber,increasing fiber length.Auxin plays an important role in promoting fiber initiation.Before,the research group created FBP7::iaaM(FM)transgenic cotton by precisely enhancing the auxin biosynthesis in the ovule epidermis to increase the auxin content in the ovule epidermis,significantly increase the lint yield and improve fiber fineness.The GTP binding protein GhROP6 in cotton has an active state of binding to GTP and an inactive state of binding to GDP.The previous research of the research group on GhROP6 found that its overexpression and activation reduced the protein abundance of GhPIN3 a on the cytoplasmic membrane of fiber by promoting the endocytosis of cotton auxin exocytosis vector GhPIN3 a,and then mediated the polar transport of auxin to promote fiber initiation.At the same time,it was found that transgenic cotton with overexpressed and activated GhROP6 can increase the length of mature fibers,indicating its potential value in improving fiber quality.Although genetic engineering has successfully changed the fiber traits of upland cotton,the effectiveness of genes used for genetic engineering improvement of cotton fiber traits in the field has been limited to date,and some results still need to be evaluated through field trials.In addition,due to the frequent negative correlation between lint yield and fiber quality,there has been little progress in achieving satisfactory fiber quality while increasing cotton yield.This study evaluated the field traits of transgenic cotton overexpressing GhROP6 proteins in different active states.Combined with the characteristics of this cotton regulating the distribution of auxin in ovule epidermis,it was hybridized with FBP7::iaaM9 transgenic cotton that increased the content of auxin in ovule epidermis,and further evaluated the agronomic traits of different hybrid combinations.Analyze the performance of all offspring of different hybrid combinations in terms of yield(seed cotton yield,lint yield,clothing percentage,etc.)and fiber quality(length,strength,and micronaire value,etc.).The main results of the paper are as follows:1.GhROP6-related transgenic cottons promoted fiber elongation,but showed poor performance in field.Observing the fiber initiation on the day of flowering of GhROP6 transgenic cotton,the initial density of 35S::CA-GhROP6(Constitutively Active,CA)transgenic fiber cells was lower than that of the control cotton(Null,separated non-transgenic cotton);And the initial density of fiber cells in 35S::DN-GhROP6(Dominantly Negative,DN)transgenic cotton was higher than that in the control.Simultaneously overexpressing 35S::GhROP6(Overexpression,OX)and CA cotton fibers elongates faster compared to the control,while DN overexpressing cotton fibers elongate shorter than the control.The overexpression or activation of GhROP6 promotes fiber initiation and elongation,while the inactivated GhROP6 inhibits fiber initiation and elongation.However,in the field experiment of GhROP6 transgenic cotton,the fiber detection results showed that the mature fiber lengths of OX and CA were close to Null,and even shorter.Except for the slightly higher lint percentage of genetically modified cotton with overexpression of DN compared to the control,there was no significant difference in various indicators of transgenic cotton compared to the control.Some indicators such as lint index and cotton boll weight were even lower than the control,resulting in poor performance in field traits.2.Additional auxin supply promoted fiber development in GhROP6-related transgenic cottons.Based on the field test,the fiber length becomes shorter,which may be caused by insufficient supply of auxin.We measured the level of auxin in ovules on the day of flowering.The test found that the OX and control Null levels were equivalent;CA and DN were lower than those of the control.It was speculated that GhROP6 protein could not be regulated by the endogenous mechanism of cotton due to the mutation of GhROP6 protein,which resulted in uneven distribution of auxin.The results of ovule culture with exogenous auxin showed that increasing auxin supply significantly promoted the fiber length of CA and DN,especially CA-GhROP6 ovule fiber.By integrating the auxin biosynthesis gene FBP7::iaaM,the endogenous level of auxin in ovule epidermis was increased.Fiber initiation cell density of FBP7::iaaM × 35S::CA-GhROP6(FM × CA)and control FBP7::iaaM × Null(FM × Null)is equivalent and saving the fiber initiation inhibitory effect of CA;Simultaneously promoting FBP7: iaaM × 35S::DN-GhROP6(FM × DN).The elongation of DN cotton fibers restored the fiber length to the control level,further promoting the fiber development of GhROP6 transgenic cotton.3.Endogenous integration auxin biosynthesis improved the cotton yield and fiber qualities of GhROP6-related transgenic hybrid cottons.In view of the poor performance of GhROP6 cotton in the field,and increasing auxin supply can promote the fiber development of GhROP6 cotton.Therefore,through genetic integration of the auxin biosynthesis gene FBP7::iaaM9,the endogenous auxin content in the ovule epidermis of GhROP6 cotton is increased.The analysis results indicate that except the fiber length and strength of hybrid cotton FM × DN cotton restored to control(FM × Null).All other hybrid cotton varieties have improved.The lint percentage of hybrid cotton FM × OX,FM × CA and FM × DN was further improved by 7.2%,11.4%,and 16.5%,respectively;Most importantly,the OX and CA hybrid cotton integrated with FM transgenic showed higher performance than FM ×Null.The fiber length and strength increase while the micronaire value decreases,indicating better fiber quality.Secondly,in terms of cottonseed yield,the yield of FM ×CA cotton significantly increased by 17%,while the cottonseed yield of other hybrid cotton varieties was lower than that of the control(FM × Null)is equivalent.In addition,compared to the control,the lint yield of FM × CA and FM × DN cotton significantly increased(the former increased by 25%,the latter increased by 18%),which is consistent with the higher lint percentage characteristics of all hybrid cotton.4.The enhancement of auxin biosynthesis and distribution synergistically improves cotton yield and fiber quality.In the field randomized block experiment,it was found that the yield of hybrid cotton increased and the quality improved.Analyzing the synergistic promotion effect of two genes,the changes in various indicators of hybrid cotton were greater than the sum of the effects of GhROP6 and iaaM genes alone.This indicates that the integration of two genes has a synergistic promoting effect.Synergistic promotion of yield and quality traits in hybrid cotton.In summary,the overexpression of GhROP6 cotton fiber length did not match expectations,with only an increase in clothing percentage.And the yield performance in the field is poor.Integrating the iaaM gene of auxin biosynthesis gene,the field test analysis showed that the lint percentage of all hybrid cotton was further improved;And the cottonseed yield and lint yield FM × CA,FM × DN are increased compared to the control FM × Null,especially FM × CA increased 17% in cottonseed yield compared to the control;The lint yield increased by 25.4% compared to the control.And it also has a significant improvement effect on the quality of fiber.The results showed that the integration of auxin biosynthesis and distribution,on the basis of FBP7::iaaM improving lint and lint yield,increased seed cotton yield.Meanwhile,data analysis shows that the integration of two genes has a synergistic improvement effect,achieving synergistic improvement of cotton yield and fiber quality. |
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