Biotechnology Bulletin ›› 2022, Vol. 38 ›› Issue (8): 92-100.doi: 10.13560/j.cnki.biotech.bull.1985.2021-1499
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CHEN Guang1(), LI Jia2, DU Rui-ying1, WANG Xu1()
Received:
2021-12-03
Online:
2022-08-26
Published:
2022-09-14
Contact:
WANG Xu
E-mail:chenguang0066@126.com;wangxuguangzhou@126.com
CHEN Guang, LI Jia, DU Rui-ying, WANG Xu. pOsHAK1:OsFLN2 Expression Enhances the Drought Tolerance by Altering Sugar Metabolism in Rice[J]. Biotechnology Bulletin, 2022, 38(8): 92-100.
基因名称Gene name | 正向引物Forward primer(5'-3') | 反向引物Reverse primer(5'-3') |
---|---|---|
UBQ5 | CTCGCCGACTACAACATCCA | TCTTGGGCTTGGTGTACGTCTT |
OsFLN2 | CCGAATGGCTTCTCTTCTTCTC | GGCTCCTGATTGAGTTGGTACTACA |
SGR | GCAATGTCGCCAAATGACG | GCTCACCACACTCATTCCCTAAAG |
OsATG8a | AGCCCAGAAAAGGCCATCTT | CATCCTTGTTCTCTTCGTAGATTGC |
OsSNAC1 | GTCAAGACTGATTGGATCATGC | CCAATCATCCAACCTGAGAGA |
OsMYB2 | GAGCAGCGAGGAGGAGGT | TGTAGTTGACGAGCAGGAGGT |
Table 1 Primers used for RT-qPCR assays
基因名称Gene name | 正向引物Forward primer(5'-3') | 反向引物Reverse primer(5'-3') |
---|---|---|
UBQ5 | CTCGCCGACTACAACATCCA | TCTTGGGCTTGGTGTACGTCTT |
OsFLN2 | CCGAATGGCTTCTCTTCTTCTC | GGCTCCTGATTGAGTTGGTACTACA |
SGR | GCAATGTCGCCAAATGACG | GCTCACCACACTCATTCCCTAAAG |
OsATG8a | AGCCCAGAAAAGGCCATCTT | CATCCTTGTTCTCTTCGTAGATTGC |
OsSNAC1 | GTCAAGACTGATTGGATCATGC | CCAATCATCCAACCTGAGAGA |
OsMYB2 | GAGCAGCGAGGAGGAGGT | TGTAGTTGACGAGCAGGAGGT |
Fig. 1 Effects of drought stress on sugar metabolism in rice Rice seedlings were growing in normal IRRI solution for 6 weeks,then treated with 15% PEG for 10 d. A:Net photosynthetic rate(Pn). B:Sucrose phosphate synthase(SPS)activity. C:Rate of sucrose export(SER)from the leaf. D-E:Sucrose contents of the leaf(D)and root(E). The values are means ± SE of 5 replicates. Significant differences between normal and 15% PEG treatment are indicated with asterisks(P < 0.05). FW:Fresh weight
Fig. 2 Construction of expression vector and process of genetic transformation in rice A:Construction map of expression vector. B-G:Callus induction(B,C),sele-ction of transformed calli(D),shoot regeneration from resistant calli(E,F),and hardening of transgenic plants(G),bar in B-D=2 cm,bar in E-G=3 cm. H:Iden-tification of positive transgenic lines by GUS staining in T0 generation,bar=1 cm
Fig. 3 Differential analysis of FLN2 expression and net photosynthetic rate between T1 generation of pHAK1:FLN2 transgenic lines and WT in response to drought stress Rice seedlings were growing in normal IRRI solution for 2 weeks,then treated with 15% PEG for 7 d. A:RT-qPCR analysis of endogenous OsFLN2 in the shoots of seedlings. UBQ5 was chosen as the reference sequence. B:Net photosynthetic rate. Results for the five null segregants were combined together as a second control(null segregant),while five transgenic lines were combined together as pHAK1:FLN2. The values are means±SE. Significant differences at P < 0.05 are indicated with different letters
Fig. 4 Seedling growth of pHAK1:FLN2 transgenic lines compared with WT in response to drought stress A:Growth performance of the seedlings under normal and 15% PEG treatment,bar = 5 cm. B-C:Shoot(B)and root(C)biomass(dry weight). The values are means ± SE of 5 replicates. Significant differences between WT and transgenic lines are indicated with asterisks(P < 0.05)and ns indicates non-significant differences at that level of significance. The same below
Fig. 5 Differential analysis of sugar metabolism between pHAK1:FLN2 transgenic lines and WT in response to drought stress A:Sucrose phosphate synthase(SPS)activity. B:Rate of sucrose export(SER)from the leaf. C-D:Sucrose contents of the leaf(C)and root(D).The values are means±SE of 5 replicates
Fig. 6 Differential analysis of root system architecture be-tween pHAK1:FLN2 transgenic lines and WT in response to drought stress A:Total root length. B:Root surface area. The values are means ± SE of 5 replicates
Fig. 7 Differential analysis of water-content ability and lipid peroxidation between pHAK1:FLN2 trans-genic lines and WT in response to drought stress A:Relative water content. B:Water loss rate. C:Relative electrolyte leakage. D:Proline content. The values are means±SE of 5 replicates
Fig. 8 Differential analysis of expressions of senescence-associated genes and stress-responsive genes between pHAK1:FLN2 transgenic lines and WT in response to drought stress The genes assayed are(A)SGR,(B)ATG8a,(C)SNAC1 and(D)MYB2. UBQ5 is chosen as the reference gene. The values are means±SE of 3 replicates
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