Gene Cloning And Phenotypic Analysis Of Abnormal Mutants From Arabidopsis Thaliana

Developmental programs in plants are under the elaborate regulation of genetic interaction networks at organs,tissues and cellular levels.Our lab is interested in dissecting the genetic basis and molecular mechanism of the plant developmental traits formation responsing to abiotic stress by the forward genetic method.Previous work has isolated a dominant Arabidopsis mutant abs3-1D(abnormal shoot3-1 dominant)from activation tagged mutant pools.abs3-1D displays short petiole,small plant type,compact yellow leaves,early flower,clustal secondary inflorescence,dwarf stem,short pods and poor fertility.In the dark induced senescence conditions,mutant abs3-1D showed a characteristic of accelerated leaf senescence.In order to further study the role of ABS3 in plant development process,abs3-1D was mutagenized with EMS(ethyl-methane sulphonate).The screened mutant F08-69 reverse the early flower of abs3-1D.F08-69 showed late flowering phenotype and F08-69 single has a prolonged vegetative growth period and slow down leaf senescence in dark condition.Whole genome sequencing of F08-69 single found that G changed to A in the +678bp position of At3g10390,leading to glycine(G)mutation into serine(S).In addition,a large number of mutants involved in leaf color,leaf shape and trichome decelopment were screened in the mutant library.An Arabidopsis recessive mutant abt3-1(aberrantly branched trichome 3)was isolated from EMS mutagenesis mutant pools.abt3-1 has reduced stature,darker green and wider leaves.abt3-1 appeareed a high proportion of multibranched trichomes and a defective primary root.The growth curves showed that mutant root growth rate was only one third of the wild-type.The meristematic zone of abt3-1 was reduced by measuring the number and the length of cortical cells in it.In this study,we found the G to A transition in the +33 position of exon 8 of At1g66510 by sequencing the candidate genes in the chromosome interval which was determined using the position cloning strategy,which resulted in a premature translation termination.Genetic complementation experiments further proved that the mutant At1g66510(ABT3)leads to the phenotype in abt3-1.ABT3 encodes a currently unknown AAR2(a1-?2 repression 2)family proteins.The subcellular localization of ABT3 protein was found in cytoplasm and nucleus using the protoplast transient expression system.