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T-DNA Insertion Mutants Reveal The Roles Of The AtSIS In Abiotic Stresses In Arabidopsis Thaliana

Posted on:2016-07-26Degree:MasterType:Thesis
Country:ChinaCandidate:J ZhangFull Text:PDF
GTID:2180330470467344Subject:Biochemistry and Molecular Biology
Abstract/Summary:PDF Full Text Request
Intrinsically disordered protein is a kind of protein without high stable structure in physiological conditions,but it can exercise the normal biological function and exists widely in eukaryotic organism. Intrinsically disordered proteins play an important role in response to abiotic stresses in plants. The At SIS encodes the protein which is a kind of intrinsically disordered protein. This protein is related to environmental stress with unkonwn function. In this paper, we studied the characteristics of At SIS disordered protein and how At SIS gene does work in response to abiotic stresses in Arabidopsis thaliana.Bioinformatic analysis showed that At SIS protein contains rich Ser residues, and less hydrophobic amino acid contents.The protein contains 3 disordered regions, and its binding with target protein may be in disordered regions. Phosphorylation sites were mainly in Ser residues which located in the long disordered regions.With 7 different T-DNA insertion mutants of At SIS as materials, 5 T-DNA insertion homozygotes were genotyped by the three-primer method. They have different insertion sites on the At SIS locus, and 1 in the 5 ’UTR region, 3 in exon regions and 1 inserted into intron.The CDS sequence of At SIS was cloned into p CAMBIA130, construct of the plant expression vector of p1303-At SIS was made, and then transformed into Arabidopsis thaliana plants by floral-dipping method. Transformed plants were selected by growth on hygromycin-containing medium and identified by PCR.The T-DNA insertional mutants of At SIS had different sensitivity compared with wild type to abiotic stresses when they were treated with Na Cl, PEG and ABA in genmination and seedling growth periods. The mutants which T-DNA inserted into exton and 5’UTR of At SIS were less affected under abitoc stresses,while the mutant which T-DNA inserted into intron was highly sensitive to the stresses.The results of q RT-PCR showed that At SIS was an early response gene to adversity stresses. The expression pattern of At SIS was consistent with that of At1g69490 and At2g18700. However, At SIS inhibited the expression of At1g69490 and At2g18700, which implied that At SIS may be an upstream gene in At2g18700 and At1g69490 under abiotic atress conditions. Through loss-offunction analyses, At SIS is shown to may act as a negative regulator of abiotic stress tolerance in Arabidopsis.
Keywords/Search Tags:Arabidopsis thaliana, Intrinsically Disordered Proteins, AtSIS, T-DNA Insertion Mutant, Abiotic Stress
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