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Egg tanning improves the efficiency of CRISPR/Cas9-mediated mutant locust production by enhancing defense ability after microinjection
ZHANG Ting-ting, WEN Ting-mei, YUE Yang, YAN Qiang, DU Er-xia, FAN San-hong, Siegfried ROTH, LI Sheng, ZHANG Jian-zhen, ZHANG Xue-yao, ZHANG Min
2021, 20 (10): 2716-2726.   DOI: 10.1016/S2095-3119(21)63736-X
Abstract135)      PDF in ScienceDirect      
The mutant efficiency and hatching ratio are two key factors that significantly affect the construction of genome-modified mutant insects.  In the construction of CRISPR/Cas9-mediated dsLmRNase2–/–mutant locusts, we found that the tanned eggs which experienced a 20-min contact with the oocyst exhibited a higher success rate compared to fresh newly-laid eggs that were less tanned.  However, the heritable efficiency of the dsLmRNase2 deletion to the next generation G1 progeny was similar between adults derived from the tanned or less tanned engineered eggs.  Further, the similar effective mutant ratios in the normally developed eggs and G0 adults of tanned and less tanned eggs also indicated that tanning did not reduce the absolute mutation efficiency induced by CRISPR/Cas9.  Moreover, we found that the syncytial division period, which was longer than the time for tanning, conferred a window period for microinjection treatment with efficient mutation in both tanned and less tanned eggs.  We further found that tanned eggs exhibited a higher hatching rate due to a reduced infection rate following microinjection.  Both the anti-pressure and ultrastructure analyses indicated that the tanned eggs contained compressed eggshells to withstand increased external pressure.  In summary, tanned eggs possess stronger defense responses and higher efficiency of genome editing, providing an improved model for developing Cas9-mediated gene editing procedures in locusts.
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Overexpression of GmProT1 and GmProT2 increases tolerance to drought and salt stresses in transgenic Arabidopsis
GUO Na, XUE Dong, ZHANG Wei, ZHAO Jin-ming, XUE Chen-chen, YAN Qiang, XUE Jin-yan, WANG Hai-tang, ZHANG Yu-mei, XING Han
2016, 15 (8): 1727-1743.   DOI: 10.1016/S2095-3119(15)61288-6
Abstract1738)      PDF in ScienceDirect      
   The proline transporter protein (ProT) plays an important role in protective stress responses in various plants. However, its function in abiotic stress responses in soybean (Glycine max) remains obscure. In the present study, two soybean ProT genes, namely GmProT1 and GmProT2, were isolated by homologous cloning. GmProT1 and GmProT2 encode polypeptides of 435 and 433 amino acids, respectively. The GmProT1 and GmProT2 proteins showed high similarity to other ProT proteins. GmProT1 and GmProT2 transcripts were detected in different soybean tissues including roots, stems, leaves, flowers, and developmental seeds, and during diverse developmental stages. GmProT1 was strongly expressed in seeds 35 days after flowering. Quantitative real-time PCR analysis showed that the two genes were highly expressed in leaves and could be strongly induced in response to salt and drought conditions and ABA treatment. Transgenic Arabidopsis thaliana plants overexpressing the two genes were generated, which showed that GmProT genes attenuate damage from salt and drought stress. In addition, transgenic Arabidopsis plants accumulated proline in response to salt and osmotic stress. Transcription levels of salinity-responsive gene (RD29B and S0S3) and drought-induced gene (CDPK1) were higher in the transgenic lines than that of wild type plants. Our work provides evidence that GmProT genes function in the response to abiotic stresses and may affect the synthesis and response system of proline.
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