Isolation and Functional Characterization of a Salt-Responsive Calmodulin-Like Gene MpCML40 from Semi-Mangrove Millettia pinnata
Abstract
:1. Introduction
2. Results
2.1. MpCML40 Is an EF-Hand Motif-Containing Calmodulin-Like Protein
2.2. MpCML40 Localizes at the Plasma Membrane and in the Nucleus
2.3. MpCML40 Gene Is Highly Induced by Salt Stress in Pongamia Roots
2.4. The pYES22-MpCML40-Transformed Yeast Has Enhanced Salt Tolerance
2.5. Heterologous Expression of MpCML40 in Arabidopsis Strongly Enhances Salt and Osmotic Tolerance
3. Discussion
3.1. MpCML40 Is a Salt-Induced CML Gene
3.2. MpCML40 Improves Salt Tolerance in Both Yeast and Arabidopsis
4. Materials and Methods
4.1. Plant Materials and Growth Conditions
4.2. Full-Length cDNA Cloning, Phylogenetic Analysis, and Motif Prediction
4.3. Subcellular Localization
4.4. Quantitative Real-Time PCR
4.5. Yeast Transformation and Growth Assay
4.6. Phenotype Analysis of Wild-Type and 35S::MpCML40 Transgenic Arabidopsis Plants
4.7. Proline Content Measurement
4.8. MDA Content Measurement
4.9. DAB and NBT Staining
5. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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1 | CCACAACATA TAATAACAAC TCAATTTTCC ATTTGCATAC AAGTTACATT TTCTCCTTCT |
61 | TATTCTTCTT GTTATTGTGT ACATTAAAGA TTTGAACAAA TTACACTACA CCTTTAAGAT |
121 | AAGGAGCATT GTAACATGAT GAAGCATGCG GGTTTTGAGG GTGTTCTTCG ATATTTTGAT |
181 | GAAGATGGGG ATGGAAAGGT TTCACCTTCA GAGTTAAGGC ATGGATTGGG AATGATGGGT |
241 | GGGGAGCTTT TGATGAAAGA AGCAGAGATG GCAATTGAGG CACTGGATTC TGATGGTGAT |
301 | GGGTTGTTGA GTTTGGAGGA TTTGATTGCT TTGATGGAAG CAGGGGGAGA GGAACAAAAG |
361 | TTGAAGGATT TGAGAGAAGC TTTTGAGATG TATGACACTC AAGGGTGCGG ATTTATAACC |
421 | CCAAAGAGCT TGAAGAGGAT GCTTAAGAAG ATGGGAGAGT CCAAGTCCAT TGATGAATGC |
481 | AAATTGATGA TTAGTCAATT TGATTTGAAT GGGGATGGGA TGCTTAGCTT TGAAGAATTC |
541 | AGAATTATGA TGGAGTGAGG CCAGTATATT TGTTGATGAT ATTGTTTAGT TTGTTTGTTT |
601 | GTTTGGGAGG AAGAGGGGTA TAGTTAAGTG GATTTGATTT ATTTGTTTGC AGTTTGCACA |
661 | TGTATAAATA ACTCCTTTTG TGCTTTGCAA TACTTTTGAC AATTGATTAA CTGTTAGATT |
721 | TCTCCCAAGT TCCTACATAA AAAATTATTC AAATTTTCTT AATGGGAGTT GTATTATGAC |
781 | TATTATCATG GTTAAATATA TTTTTTATTT CGTTCCAAAA AAAAAAAAAA AAAAAAAAAA |
Primer | Primer Sequence (5′ → 3′) |
---|---|
MpActin.RtF | AGAGCAGTTCTTCAGTTGAG |
MpActin.RtR | TCCTCCAATCCAGACACTAT |
MpCML40.RtF | GCACTGGATTCTGATGGTGATGGG |
MpCML40.RtR | GCTCTTTGGGGTTATAAATCCGCA |
MpCML40.3’GSP | TGGCAATTGAGGCACTGGATTCTGATGG |
MpCML40.3’NGSP | ATGGAAGCAGGGGGAGAGGAACAA |
MpCML40.5’GSP | TCCGCACCCTTGAGTGTCATACATCTCA |
MpCML40.5’NGSP | CCTCCAAACTCAACAACCCATCACCATC |
MpCML40.yeastF | CGCGGATCCATGATGAAGCATGCGGGTTTTGAGG |
MpCML40.yeastR | CCGCTCGAGTCACTCCATCATAATTCTGAA |
MpCML40.plantF | TATGGCGCGCCCATGATGAAGCATGCGGGTTTTGA |
MpCML40.plantR | CTCCATCATAATTCTGAATTCTT |
AtACT2.RtF | GACCTTTAACTCTCCCGCTATG |
AtACT2.RtR | GAGACACACCATCACCAGAAT |
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Zhang, Y.; Huang, J.; Hou, Q.; Liu, Y.; Wang, J.; Deng, S. Isolation and Functional Characterization of a Salt-Responsive Calmodulin-Like Gene MpCML40 from Semi-Mangrove Millettia pinnata. Int. J. Mol. Sci. 2021, 22, 3475. https://doi.org/10.3390/ijms22073475
Zhang Y, Huang J, Hou Q, Liu Y, Wang J, Deng S. Isolation and Functional Characterization of a Salt-Responsive Calmodulin-Like Gene MpCML40 from Semi-Mangrove Millettia pinnata. International Journal of Molecular Sciences. 2021; 22(7):3475. https://doi.org/10.3390/ijms22073475
Chicago/Turabian StyleZhang, Yi, Jianzi Huang, Qiongzhao Hou, Yujuan Liu, Jun Wang, and Shulin Deng. 2021. "Isolation and Functional Characterization of a Salt-Responsive Calmodulin-Like Gene MpCML40 from Semi-Mangrove Millettia pinnata" International Journal of Molecular Sciences 22, no. 7: 3475. https://doi.org/10.3390/ijms22073475