Selection and Characterization of a Novel Glyphosate Tolerant Upland Cotton(Gossypium hirsutum L.) Mutant(R1098)

Stepwise selection approach was adopted to obtain glyphosate-tolerant upland cotton mutant(R1098) from the embryogenic calli of Coker 312(Gossypium hirsutum L.).The calli were transferred to selection medium and multi-step selection pressure process was carried out until the

In Vitro Selection and Identification of Drought-Tolerant Mutants in Sweetpotato

In vitro selection of drought-tolerant mutants in sweetpotato cv. Lizixiang was studied by using PEG6000 as selection stress. Embryogenic suspension cultures were cultured in MS medium containing 035% PEG6000 and 2 mg L 1 2,4-dichlorophenoxyacetic acid (2,4-D). The results indicated that 30% PEG6000 can be used for the optimal selection stress of drought-tolerance. Embryogenic suspension cultures irradiated with 80 Gy gamma-ray were cultured in MS medium containing 30% PEG6000 and 2 mg L 1 2,4-D and 20 drought-tolerant cell aggregates were obtained. These cell aggregates were transferred to solid MS me- dium supplemented with 2 mg L 1 2,4-D and formed embryogenic callus with somatic embryos. The embryogenic callus with somatic embryos was further transferred to MS medium supplemented with 1 mg L 1 abscisic acid (ABA), resulting in the germination of somatic embryos. In this study a total of 18 regenerated plants were obtained. The regenerated plants were transplanted in a greenhouse and 11 lines were formed. The analysis on drought treatment of seedlings, water retaining capacity of leaves and coefficient of drought-tolerance showed that 3 lines had significant drought-tolerance in comparison with the control plants.

Induced Mutation for Developing Mutant Rice Lines Tolerant to the Parasitic Weed Striga asiatica (L.) Kuntze

This work aims to screen mutant rice lines tolerant to Striga asiatica.Two rainfed sensitive rice varieties B22 and F154 were used.Plants survival rates of the two parents were significantly lower respectively(9.74a and 11.83a)than those of mutant lines(55.36c to 74.36b);Striga plants emergence/pot were significantly higher for the parents(13.96c and14.89c)compared to the mutants(0.12a to 1.5b);the infection rate of parents(7.37b;7.86b)was higher compared to the mutants(2.27a to 2.74a);fertility rate/plant of parents was lower(20.98%b;22.29%b)but much higher than mutants(72.19%b to 78.35%b);the average panicle number/plant of parents was significantly lower(0.5a;1a)than those of mutants(1.5b to 2.4bc)and the 100 g grain weight of parents are lower(2.35a;2.56a)than those of mutants(3.19b to 3.23b).The culture of those mutant lines may increase rice production and contribute to enhancing food security in Madagascar.

Screening for Spikelet Fertility and Validation of Heat Tolerance in a Large Rice Mutant Population

A total of 10 000 M4 individuals in Jao Hom Nil (JHN) mutant population was treated with high temperature (40 ℃ to 45 ℃) during the day time (6 h) from the booting to the harvesting stages, and ambient temperature (33 ℃ to 35℃) was used as the control. The results of screening and yield trials found that the mutant line M9962 had a high spikelet fertility of 78% under heat stress. In addition, the other mutant lines, including M3181 and M7988, had a spikelet fertility of approximately 70%. However, the JHN wild type, Sin Lek, RD15 and RD33 had very low spikelet fertility of 34%, 14%, 9% and 4%, respectively. The lower spikelet fertility at an elevated temperature resulted in a dramatic decrease of filled grain and contributed to a loss in 100-grain weight. M9962 is a potential genetic stock for use in a heat tolerance breeding programme. In addition, spikelet fertility at high temperature was representative of heat tolerance and can be used as a screening trait for heat tolerance during the reproductive phase on a large scale.

Characterization of salt tolerance and Fusarium wilt resistance of a sweetpotato mutant

The variant LM1 was previously obtained using embryogenic cell suspension cultures of sweetpotato variety Lizixiang by gamma-ray induced mutation, and then its characteristics were stably inherited through six clonal generations, thus this mutant was named LM1. In this study, systematic characterization of salt tolerance and Fusarium wilt resistance were performed between Lizixiang and mutant LM1. LM1 exhibited significantly higher salt tolerance compared to Lizixiang. The content of proline and activities of superoxide dismutase（SOD） and photosynthesis were significantly increased, while malonaldehyde（MDA） and H_2O_2 contents were significantly decreased compared to that of Lizixiang under salt stress. The inoculation test with Fusarium wilt showed that its Fusarium wilt resistance was also improved. The lignin, total phenolic, jasmonic acid（JA） contents and SOD activity were significantly higher, while H_2O_2 content was significantly lower in LM1 than that in Lizixiang. The expression level of salt stress-responsive and disease resistance-related genes was significantly higher in LM1 than that in Lizixiang under salt and Fusarium wilt stresses, respectively. This result provides a novel and valuable material for improving the salt tolerance and Fusarium wilt resistance of sweetpotato.

Digging out molecular markers associated with low salinity tolerance of Nannochloropsis oceanica through bulked mutant analysis

Nannochloropsis oceanica is a marine microalgal species with both economic value and biological importance.It grows fast,contains rich oils,reproduces asexually,holds a small and haploidy genome,and is easy to be modified genetically.However,the genetic study of N.oceanica is scarce.Very less genetic bases of its traits have been deciphered,and no gene has been isolated from it with the function verified simultaneously via either genetic or reverse genetic approaches or both(de novo cloned).Changing medium salinity may aid to control harmful organisms met during large scale cultivation.As a stress,it may also facilitate the accumulation of desirable chemicals including fatty acids.In order to decipher the genetic basis of the low salinity tolerance of N.oceanica,we mutated N.oceanica with Zeocin.In total,five mutant bulks were constructed at equal number of cells,100 mutants each,which were tolerant to a discontinuous serial of salinities from that of 100%of f/2 to that of a mixture of 4%of f/2 and 94%of BG11.The bulks were genotyped through whole genome re-sequencing and analyzed with bulked mutant analysis(BMA)newly modified from bulked segregant analysis(BSA).In total,47 SNPs and 112 InDels were found to associate with the low salinity tolerance,and around them a set of low salinity tolerance associating genes were identified.A set of annotatable genes commonly found between control and different salinities indicated that the genes functioning in gene expression,energy metabolism and cellular structure may be involved in the low salinity tolerance.These associating molecular markers and genes around them were not enough for outlining the physiological mechanism underlining the tolerance;however they should aid to improve N.oceanica genetically.

Ethanol Fermentation by High-Stress-Tolerance Aquatic Yeasts and Their Mutants

Bioethanol is thought to be a renewable source of energy, because the biomasses used to make ethanol, such as sugar cane and its residual substance, molasses, are resources that can be continuously produced. But the practical use of ethanol to replace fossil fuels or atomic energy has been limited, because the production efficiencies of ethanol in relation to its substrates are not so high. Thus, for industrial production of the bioethanol, yeast fermentation would ideally be carried out in biomasses containing more highly concentrated carbohydrates. However, the environmental stresses in highly concentrated cultures might weaken the yeast’s physiological activities. From various kinds of aquatic yeast with stress tolerance, Torulaspora derbrueckii F2-11 and Wicherhamomyces anomalus AN2-64 were selected as candidates for high-sugar-tolerance yeasts as they showed remarkable growth in the YPD + sorbitol (600 g/L) medium at 25°C for 120 hrs. When the amounts and kinds of sugar alcohols in the cells of the two strains were measured in cultures containing 20 g/L or 400 g/L of D-glucose, maltose, or sucrose, the main two sugar alcohols that accumulated as the sugar concentration increased were glycerol and arabitol. Mutation by ethyl methanesulfonate of the parent strains T. derbrueckii F2-11 and W. anomalus AN2-64 induced mutants F2-11M or AN2-64M, which showed higher sugar, heat, and ethanol tolerances than their respective parents. Ethanol productivities and sugar assimilation activities of the mutants were also higher than those of the parents in the 25% (v/v) molasses.

The Potassium Transporter AtKUP12 Enhances Tolerance to Salt Stress through the Maintenance of the K^(+)/Na^(+)Ratio in Arabidopsis

Potassium(K^(+))is a necessary nutrient for plant growth and crop production.The K^(+)transporter plays crucial roles in the absorption and transport of K^(+)in plants.Most K^(+)transporters in Arabidopsis have been reported,but AtKUP12,which is a member of the KT/KUP/HAK family,has not yet been the subject of relevant in-depth research.In the present study,we demonstrated that AtKUP12 plays a crucial role in K^(+)uptake in Arabidopsis under 100μM low-K^(+)and 125 mM salt stress conditions.AtKUP12 transcripts were induced by K^(+)deficiency and salt stress.We analyzed the K^(+)uptake of AtKUP12 using the K^(+)uptake-deficient yeast R5421 and Arabidopsis mutant atkup12.Transformation with AtKUP12 rescued the growth defect of mutant yeast and atkup12 mutant plants at the low-K^(+)concentration,which suggested that AtKUP12 might be involved in high-affinity K^(+)uptake in low-K^(+)environments.In comparison to the wild-type(WT)and atkup12-AtKUP12 complementation lines,atkup12 showed a dramatic reduction in potassium concentration,K^(+)/Na^(+)ratio,and root and shoot growth on 12-day-old seedlings under the salt conditions;however,there was no significant difference between the complementation and WT lines.Taken together,these results demonstrate that AtKUP12 might participate in salt tolerance in Arabidopsis through K^(+)uptake and K^(+)/Na^(+)homeostasis.

Mutation-Screening of Pleurotus Ferulae with High Temperature Tolerance by Nitrogen Ion Implantation

In order to obtain Pleurotus ferulae with high temperature tolerance, conidiophores of wild type strain ACK were implanted with nitrogen ions in energy of 5 -15 keV and dose of 1.5×10^15 - 1.5 × 10^16 cm^-2, and a mutant CGMCC1763 was isolated subsequently through thermotolerant screening method. It was found that during riper period the surface layer mycelium of the mutant in mushroom bag wasn＇t aging neither grew tegument even above 30℃. The mycelium endurable temperature of the mutant was increased by 5℃ compared to that of the wild type strain. The fruiting bodies growth temperature of the mutant was 18 -22℃ in daytime and 8 -14℃ at night. The highest growth temperature of fruiting bodies of the mutant was increased about 7℃ w.r.t, that of original strain. Through three generations investigations, it was found that the mutant CGMCC1763 was stable with high temperature tolerance.

Maintenance of Normal Stress Tolerance in the Moss Physcomitrella patens Lacking Chloroplastic CuZn-Superoxide Dismutase

Superoxide dismutases (SODs) catalyze the dismutation of superoxide and play an important role in reducing oxidative stress in plants. Based on in-gel SOD activity staining, chloroplasts of the moss Physcomitrella patens have two CuZn-SODs as the major SOD isozymes and minor SODs, including a Fe-SOD and two Mn-SODs. To investigate the contribution of chloroplastic SODs to stress tolerance in P. patens, we generated a double mutant lacking chloroplastic CuZn-SOD genes. The mutant did not show any differences in comparison to the wild type based on the growth of protonemata on normal and high-salt media, extractable activities of the other SODs after culture on normal and high-salt media, and inhibition of Fv/Fm under stress conditions (high-salt, high-light, and high-temperature). These results indicate that chloroplastic CuZn-SODs do not play a principal role in oxidative stress tolerance in chloroplasts under the investigated conditions. These findings explain the previously reported unusual response of P. patens to copper deficiency, in which chloroplastic CuZn-SODs are preferentially inactivated but cytosolic CuZn-SODs are unaffected.

水稻组培耐盐突变体筛选体系的研究

本研究利用组培诱导突变体技术来开展水稻耐盐突变体筛选体系建立研究,为水稻育种新材料的获得提供新方法。本研究以‘日本晴’‘C418’‘C34’‘C210’四个粳稻品系为材料,利用N6+2.0 mg/L 2,4-D,附加4种不同NaCl浓度作为诱导培养基,进行水稻愈伤组织诱导和突变体筛选,同时结合再生植株生理指标测定。结果表明:随着NaCl浓度升高,水稻不同品系之间的愈伤诱导率及继代存活率总体趋势呈下降状态,在150 mmol/L达到最低;而愈伤组织分化率也呈现显著下降趋势,在150 mmol/L浓度下分化率为0%;通过再生植株叶片过氧化物酶(POD)活性测定发现该酶呈显著上升趋势,丙二醛(MDA)含量呈显著下降趋势,脯氨酸(Pro)含量呈显著上升趋势,因此综合以上结果分析认为,最佳的NaCl筛选浓度为100 mmol/L,对获得水稻耐盐突变体筛选最为有利,研究结果为水稻耐盐突变体诱导体系建立打下基础。

Differential gene expression in salt-tolerant rice mutant and its parental variety

The differential expressions of three genes rbcL, salT and rab!6 in response to ABA, NaCl, PEG and heat shock were investigated in seedlings of a salt-tolerant rice mutant 20 (mutant 20) and its parental variety Oryza sativa var. japonica 77-170(170). By Northern blot analysis it was found that ABA induced the expression of all three genes of rbcL, salT and rab16 in shoots and roots of both 170 and mutant 20 with the exceptions of rab16 in shoots of mutant 20 and rbcL in roots of 170. Lower concentrations of NaCl induced rbcL expression in shoots of mutant 20 but not 170. Higher concentrations of NaCl decreased rbcL expression but induced expressions of salT and rab16 in shoots of both 170 and mutant 20. PEG(15%) and 37℃ heat shock showed almost no effects on the expression of the three genes in mutant 20. However, they caused a decrease in rbcL expression and slight induction of the rab16 gene in 170, with salT expression unaffected. These results indicated that mutant 20 was relatively less responsive to applied hormonal and environmental factors as compared with 170, suggesting that mutant 20 might have acquired mechanisms by which the plant is less responsive to environmental stresses and hence gain a stronger ability to tolerate stresses.

Analysis of genes specifically expressed under salt stress in salt-tolerant mutant of rice by using DDRT-PCR technique

Differential display reverse transcription-PCR (DDRT-PCR) technique was used to identify those genes that are expressed differentially between wild type rice variety 77-170 (Oryza Sativa vas Japonica) and its salt-tolerant mutant (M-20) under salt stress. Totally 13 salt-inducible cDNA fragments of 200-600 bp were identified and doned, and were designated as SIGR1 - SIGR13 (salt-induced gene in rice). Northern blot analysis showed that expression of SIGR6 and SIGR8 was salt-inducible in both wild type and mutant, and expression of SIGR12 in M-20 was much higher than that in 77-170 under salt stress. It was also shown that expression of SIGR3, SIGR4, SIGR7, SIGRIO and SIGR13 was salt-inducible, and the genes were highly homologous with Rab1d which was an ABA-inducible gene of rice. The great potential application of DDRT-PCR technique in plant molecular biology research may promote the investigation of expression of salt-induced protein in rice.

Enhanced H^+ transport activity of tonoplast vesicles isolated from roots of salt-tolerant mutant of wheat under NaCl stress

Both enhanced were H^+ transport activities of tonoplast vesicles isolated from roots of the salt-tolerant mutant and wild type of wheat with treatment of NaCI, but the activity of the mutant was significantly higher than that of wild type. H^+ transport activity was indicated as the stable value of fluorescence quenching per mg membrane proteins. The H^+ transport activities dependent on ATP of the mutant and wild type were 1099 and 558 respectively and their activities dependent on PPi were 358 and 228 separately.

Mutant Selection of Glyphosate-tolerant Soybean

Over 10% loss of agricultural incomes results from the weeds in the crop fields. But the cost of developing a new herbicide is 20-100 times as much as that of breeding a new cultivar of herbicide-tolerant crop. So such crops are bred abroad and there appear an Atrazine-tolerant cultivar of rape by hybridization breeding, an Imazapyr-tolerant line of maize selected from tissue culture, a Chlorsulfuron-tolerant soybean

Reduced Na^+ and K^+ permeability of K^+ channel in plasma membrane isolated from roots of salt-tolerant mutant of wheat

The Na^+ and K^+ permeability of K^+ channel in plasma membrane, isolated from roots of the salt-tolerant mutant of ,wheat, was lower than that of wild type in 100 mmol/L KCI and NaCI solution. The opening frequency of K^+ channel of the mutant reduced more significantly than that of wild type in two kinds of solution mentioned above. It is assumed that the reduction of opening frequency mainly contributes to the Na^+ and K^+ permeability of K^+ channel of the mutant. The electric conductance of single-channel of the mutant was similar to that of wild type and the main difference between them was exhibited as the opening frequency. Their K^+/Ka^+ selectivity of K^+ channel had no significant difference. The K^+/Na^+ selectivity of the mutant and wild type was 3.35 and 3. 18 respectively.

甘薯耐旱突变体的离体筛选与鉴定

以甘薯品种栗子香为材料 ,用PEG6 0 0 0作为耐旱性突变体离体筛选的选择剂 ,将栗子香胚性悬浮细胞培养在含有 0～ 35 % (w/v)PEG6 0 0 0和 2 .0mg·L-12 ,4 D的液体MS培养基中。结果表明 ,甘薯耐旱性离体筛选的适宜选择压为 30 %PEG6 0 0 0。采用多步选择法 ,在含有 30 %PEG6 0 0 0和 2 .0mg·L-12 ,4 D的液体MS培养基中对经 80Gyγ射线辐照的栗子香胚性悬浮细胞进行离体筛选 ,获得了 2 0个耐旱性小细胞团。将这些小细胞团转移到添加2 .0mg·L-12 ,4 D的固体MS培养基上 ,获得了耐旱胚性愈伤组织 ,并形成体细胞胚。将具有体细胞胚的胚性愈伤组织进一步转移到添加 1.0mg·L-1ABA的MS培养基上 ,体细胞胚发芽形成完整植株 ,共获得 18株再生植株。将获得的 18株再生植株移栽到温室 ,获得 11个株系。用苗期干旱处理、叶片保水力及耐旱系数等指标对这 11个株系进行耐旱性分析 ,其中 3个株系的耐旱性显著高于对照。

小麦耐盐突变体的分子生物学鉴定

利用Ｆ１花药培养、ＥＭＳ诱变和耐盐性反复筛选后已稳定９ 代的小麦耐盐突变体ＲＨ８７０６－ ４９、Ｈ８７０６－ ３４、Ｈ８７０６－ ４４、Ｈ８７０６－ ４８、Ｈ８７０６－ ５７ 及其亲本濮农３６６５、百农３０３９ 为材料，用生化标记（醇溶蛋白）及分子标记（ＲＡＰＤ）分析了各材料间的差异，发现突变体与亲本相比，不仅发生了蛋白质水平的变异，而且也在ＤＮＡ 水平上证明了突变的发生，从而为耐盐突变体的真实性提供了有力的证据，排除了盐适应的可能性； 经用２１８ 个引物对５ 个突变体之间的多态性进行ＲＡＰＤ分析，结果表明，它们之间的差异很小，其遗传背景相似。

应用细胞工程获得受主效基因控制的水稻耐盐突变系

实验以水稻花药为材料，经ＥＭＳ处理或未处理，在ＮａＣｌ胁迫下筛选耐盐突变体。所获变异系耐盐性已稳定遗传到第１３代。回交测定Ｆ２耐盐性呈３∶１的分离，表明耐盐性受一个主效基因控制。实验结果还表明，通过合适的筛选方法，无论诱变处理与否，都能得到稳定遗传的耐盐突变体。