Genotype dependent callus induction and shoot regeneration in sunflower (Helianthus annuus L.)

This study aims to observe the effect of genotype, hormone and culture conditions on sunflower (Helianthus annuus L.) callus induction and indirect plant regeneration. Calli were obtained from hypocotyl and cotyledon explants of five different sunflower genotypes; Trakya 80, Trakya 129, Trakya 259, Trakya 2098 and Viniimk 8931, which are commercially important for Turkey. Seeds germinated on Murashige and Skoog (MS) media contained no hormones. Hypocotyl and cotyledon explants were cultured on MS media supplemented with 1 mg/l 2,4-D (2,4-dichlorophenoxy acetic acid) and different percentage of callus inductions were obtained. Calli were cultured on MS + 1 mg/l BA (6-benzylaminopurine) and 0.5 mg/l NAA (-naphthalene acetic acid). Some genotypes showed high regeneration response while others showed lower on the same media with hypocotyl and cotyledon derived calli. This study showed that genotypic differences affect callus induction and plant regeneration in sunflower tissue culture studies

BAGY2 Retrotransposon Variations in Barley Calli Cultures and Regenerated Plantlets

Retrotransposons are ubiquitous components of genomes especially in cereals. Because of its high retrotransposon content barley is an excellent model plant for retrotransposon studies. BAGY2 is one of the active retrotransposon in barley and it encodes five (gag, protease, reverse transcriptase, ribonuclease H and integrase) proteins needed for its own transposition. It is known that some retrotransposons are activated by tissue culture. To find out whether BAGY2 have a copy number alteration during callus and shoot development we analyzed copy number of BAGY2 internal domains by qPCR in 45- and 90-day-old calli and regenerants deriving from the same embryo. qPCR results were evaluated by One-Way ANOVA. It was found that the copy number of all the domains increased in different tissue culture samples. Further, BAGY2 internal domains were higher in calli samples then shoots. This result shows that the cells which have more stable genome than other cells have more chance for regeneration

Optimization of gene transfer into barley (Hordeum vulgare L.) mature embryos by tissue electroporation

This study was conducted to detect the optimum conditions for DNA transfer into mature embryos of barley via electroporation. Cultured mature embryos of barley were directly electroporated in the presence of the pBI 121 vector carrying both the beta-glucuronidase (GUS) and neomycin phosphotransferase II (npt II) genes. It was found that 500 v/cm and 500 mu Fd capacitance was the optimum combination for healthy germination of the transformed plants from mature electroporated embryos. Effects of culture duration before electroporation and selection antibiotic concentrations on germination were also examined. Gene transfer performed on 3-day-old cultures resulted in the highest germination frequencies. GUS expression was observed on transversal sections of embryos and mature leaves from 3 month-old regenerants. PCR and Southern blot analyses show the presence of the npt II transgene in the genome of a plant

Direct somatic embryogenesis and synthetic seed production from Paulownia elongata

We have developed a reproducible system for efficient direct somatic embryogenesis from leaf and internodal explants of Paulownia elongata. The somatic embryos obtained were subsequently encapsulated as single embryos to produce synthetic seeds. Several plant growth regulators [6-benzylaminopurine, indole-3-acetic acid, alpha-naphthaleneacetic acid, kinetin and thidiazuron (TDZ)] alone or in combination were tested for their capacity to induce somatic embryogenesis. The highest induction frequencies of somatic embryos were obtained on Murashige and Skoog (MS) medium supplemented with 3% sucrose, 0.6% Phytagel, 500 mg l(-1) casein hydrolysate and 10 mg l(-1) TDZ (medium MS10). Somatic embryos were induced from leaf (69.8%) and internode (58.5%) explants on MS10 medium after 7 days. Subsequent withdrawal of TDZ from the induction medium resulted in the maturation and growth of the embryos into plantlets on MS basal media. The maturation frequency of somatic embryos from leaf and internodal explants was 50.8% and 45.8%, respectively. Subculturing of mature embryos led to their germination on the same medium with a germination frequency of 50.1% and 29.8% from leaf and internode explants, respectively. Somatic embryos obtained directly on leaf explants were used for encapsulation in liquid MS medium containing different concentrations of sodium alginate with a 30-min exposure to 50 mM CaCl2. A 3% sodium alginate concentration provided a uniform encapsulation of the embryos with survival and germination frequencies of 73.7% and 53.3%, respectively. Storage at 4degreesC for 30 days or 60 days significantly reduced the survival and complete germination frequencies of both encapsulated and non-encapsulated embryos relative to those of non-stored somatic embryos. However, the survival and germination rates of encapsulated embryos increased following storage at 4degreesC. After 30 days or 60 days of storage, the survival rates of encapsulated embryos were 67.8% and 53.5% and the germination frequencies were 43.2% and 32.4%, respectively. These systems could be useful for the rapid clonal propagation and dissemination of synthetic seed material of Paulownia elongata

Indirect somatic embryogenesis and plant regeneration from leaf and internode explants of Paulownia elongata

Several plant growth regulators BA, TDZ, 2,4-D and Kn were tested alone or in combination for their capacity to induce indirect somatic embryogenesis from leaf and internode explants of Paulownia elongata. Calli were produced when leaf explants were cultured on Murashige and Skoog (MS) medium containing 3% sucrose, 0.4% phytagel, 4 mg l(-1) TDZ and 0.1 mg l(-1) Kn after 3 weeks and the initiation rate was 54.1%. After subculturing on the same medium, embryos at various developmental stages ( globular, heart and torpedo shaped) were transferred for maturation onto MS medium supplemented with 3% sucrose, 0.4% phytagel, 0.1 mg l(-1) TDZ, 1 mg l(-1) Kn and 2 mM glutamine. An average of 50.7 somatic embryos were obtained from 100 mg of embryogenic callus after 4 weeks at high frequency (64.7%). Afterward, mature somatic embryos were isolated and cultured on hormone-free MS medium for germination (80%) and development into plantlets. Plantlets were transferred to pots with a mixture of peat and perlite in a 3: 1 ratio and showed a survival rate of 70 - 80%. Plantlets regenerated by this procedure were morphologically identical to the donor material and developed normally in the greenhouse

Evaluation of Barley lncRNAs Expression Analysis in Salinity Stress

Long noncoding RNAs (lncRNAs) act important roles in a wide range of biological processes. The regulatory roles of lncRNAs are still poorly understood. One of the major problems of limiting plant productivity is the salinity in the worldwide that barley (Hordeum vulgare L.) seems to be relatively well adapted to salinity environments. The aim of this study is the investigation of lncRNAs' expression levels on four barley genotypes (Hasat, Beysehir 99, Konevi 98 and Tarm 92) to 150 mM salt stress application during 3 days germination. Grains were placed randomly in petri dishes containing filter paper soaked in (a) only H2O (control), (b) 150 mM NaCl for 72 h. RNA extraction were carried out using TriPureA (R) reagent from root and shoot samples obtained after 150 mM salt treatment. Expression levels of CNT0018772 and CNT0031477 were determined by qPCR. Expression analysis demonstrated salinity effected expression levels of CNT0018772 and CNT0031477 on roots and shoots during germination. The expression levels of CNT0018772 for 150 mM salt applied groups were down-regulated raged between (log2-0.52 and-35.65) compared controls on roots and shoot. The expression levels of CNT0031477 in 150 mM salt applied groups were also down-regulated ranged between (log(2)-10.40 and 33.59) compared controls on roots and shoot except for Tarm 92 variety. On the contrary, expression levels of CNT0031477 were up-regulated on root and shoot of Tarm 92. Comparison of CNT0018772 and CNT0031477 expression levels on roots, there was no significant difference between barley varieties compared to controls (p > 0.05). However, it was found there was statistically significant difference between 150 mM salt treatment and control groups for CNT0031477 expression levels (p < 0.05). It was determined Konevi 98 shoot control expression level was statistically higher than Tarm 92 shoot control. This is the first report about the lncRNAs expression levels of barley under salinity

Analyses of abiotic stress and brassinosteroid-related some genes in barley roots grown under salinity stress and HBR treatments: Expression profiles and phylogeny

We aimed to investigate abiotic stress (WAK, HvPIP1.1, HvPIP1.2, HvPIP1.3, HvPIP1.5, CYCD3, DREB2) and brassinosteroid-related gene (DWARF4) expressions in barley (Hordeum vulgare L. cv. Hilal) roots grown under different salt (150 and 250mM), HBR (0.5 and 1M), and salt+HBR applications during 48 and 72h at dark with their controls. Phylogenetic trees were also constructed to observe relationships among genes found in other plants. The expression of HvPIP1.2 and WAK reduced after salt treatment while HvPIP1.3, DREB2 and DWARF4 expressions increased. HvPIP1.1, HvPIP1.2, HvPIP1.3, HvPIP1.5 and DWARF4 expressions were upregulated under only HBR applications. Salt+HBR treatments increased HvPIP1.1, DREB2 and DWARF4 but decreased HvPIP1.2. Phylogenetic analyses indicated that Oryza sativa L. shared similar sequences with HvPIP1.5. CYCD3 could diverge relatively earlier from cyclin genes during evolution as it segregates in a distinct clade. Sorghum bicolor showed sequence homology with DREB2. Oryza australiensisL. and DWARF4 were found in the same clade. To our knowledge, this is the first detailed report related to salt stress and HBR applications in terms of the expression of different genes in barley, providing a valuable information for molecular breeding improvement of stress-related traits

Agricultural Biotechnology İn Turkey

1st Balkan Countries Workshop on Crop Improvement by Means of Biotechnology Including Genetic Engineering -- JUL 03-05, 1995 -- VARNA, BULGARIAWOS: A1995TP94400020Balkan Plant Biotechnol Networ

Retrotransposition Events in Barley Callus Cultures

Gene transfer techniques offer an alternative to improve existing cultivars and to obtain new ones. Establishment of tissue culture procedures is a prerequisite in order to obtain stable transformants. However, plants regenerated from tissue culture may exhibit somaclonal variation. These variations, generally undesired, include nucleotide mutations, gene activation/silencing, chromosome abnormalities. Gene silencing and activation occur by DNA methylation and activation of transposable elements. Movement of transposable elements also causes insertional polymorphisms in genome. DNA polymorphisms in 15-day-old, 30-day-old calli and 4-day-old barley (Hordeum vulgare 'Zafer-160') seedling were investigated with Inter-Retrotransposon Amplified Polymorphism (IRAP), a retrotransposon-based marker system. PCR primers designed from BARE-1 and Sukkula elements were combined and amplification products were resolved on non-denaturing polyacrylamide gels. Different polymorphic bands were detected, although banding patterns were highly monomorphic among different tissues. However, control (4-day-old seedling) tissue and 30-day-old callus gave more similar fingerprints