Muscle Contraction : (Outline Studies in Biology) by C.R. Bagshaw Chapman & Hall; London,New York, 1982 79 pages. £2.75

On incubation of B. subtilis RM125(arg15 leuA8 r_M^? m_M^?) with DNA from alkalophilic Bacillus, the transformants (Arg⁺Leu^? or Leu^?Arg⁺) appeared at pH 10. The transformants were able to grow even at pH 7. Alkalophilic Bacillus was resistant to bacteriophages π105D1C2·1012 grown on B. subtilis 1012(r^-m_M⁺) and π105D1C2·ISMR4 grown on B. subtilis ISMR4r_M⁺r_R⁺m_M⁺m_R⁺), but the recipient B. subtilis and the transformant(Arg⁺Leu^?) were susceptible to both the of the bacteriophages. The results indicate that the transformant is a B. subtilis derivative and that alkalophilicity of alkalophilic Bacillus was transferred to B. subtilis.     

Functional characterization of a UDP-glucose:flavonoid 3-O-glucosyltransferase from the seed coat of black soybean (Glycine max (L.) Merr.)

The seed coats of black soybean (Glycine max (L.) Merr.) accumulate red (cyanidin-), blue (delphinidin-), purple (petunidin-), and orange (pelargonidin-based) anthocyanins almost exclusively as 3-O-glucosides; however, the responsible enzyme has not been identified. In this study, the full-length cDNA which encodes the enzyme that catalyzes the final step in anthocyanin biosynthesis, namely UDP-glucose:flavonoid 3-O-glucosyltransferase (UGT78K1), was isolated from the seed coat tissue of black soybean using rapid amplification of cDNA ends (RACE). Of the 28 flavonoid substrates tested, the purified recombinant protein glucosylated only anthocyanidins and flavonols, and demonstrated strict 3-OH regiospecificity. Galactose could also be transferred with relatively low activity to the 3-position of cyanidin or delphinidin in vitro. These findings are consistent with previous reports of mainly 3-O-glucosylated and minor amounts of 3-O-galactosylated anthocyanins in the seed coat of black soybean. The recombinant enzyme exhibited pronounced substrate inhibition by cyanidin at 100 μM acceptor concentration. Transfer of UGT78K1 into the Arabidopsis T-DNA mutant (ugt78d2) deficient in anthocyanidin and flavonol 3-O-glucosyltransferase activity, restored the accumulation of anthocyanins and flavonols, suggesting the in vivo function of the enzyme as a flavonoid 3-O-glucosyltransferase. Genomic and phylogenetic analyses suggest the existence of three additional soybean sequences with high similarity to UGT78K1. RT-PCR confirmed the co-expression of one of these genes (Glyma08g07130) with UGT78K1 in the seed coat of black soybean, suggesting possible functional redundancies in anthocyanin biosynthesis in this tissue.     

Unstable expression of a soybean gene during seed coat development

Summary The R gene of soybean is involved in anthocyanin synthesis in the seed coat, and its r-m allele conditions a variegated distribution of black spots and/or concentric rings of pigment superimposed on an otherwise brown seed coat. We describe an unusual feature of r-m that causes expression at the R locus to switch between active and inactive phases both somatically and germinally. Non-heritable somatic changes of the allele produce single plants containing mixtures of seed with different coat colors (black+striped or brown+striped). Heritable changes of the r-m allele are manifested in progeny plants which produce all black seed or all brown seed. Surprisingly, subsequent generations from revertant sublines show continued instability of the allele such that brown revertants (r*/r*) or homozygous black seed revertants (R*/R*) can give rise to striped or striped+black-seeded plants. Thus, the revertants produced by the r-m allele are not stable but interconvert between all three forms (R*, r*, and r-m) at detectable frequencies. Mutability of the r-m allele in a different genetic background has also been found after inter-crossing various soybean genotypes.     

Genetic instability in Drosophila Melanogaster: Mutable miniature (mμ)

A new mutable gene, mutable miniature wing (m^μ), is described. This mutable gene mutates spontaneously at an inordinate rate both germinally and somatically. Two classes of reversions of m^μ have been found in approximately equal frequency: those to an allele with an intermediate phenotype (mⁱ) and those to a subliminal allele (m^s) equivalent to wild type in phenotype. Reversions appear to be mutationally stable. The chronology of genetic events leading to the discovery of m^μ implicates, but does not prove, the insertion of a “foreign” DNA segment as the basis of mutability.     

Detection and fine mapping of two quantitative trait loci for partial resistance to stripe virus in rice (Oryza sativa L.)

Rice stripe virus (RSV) is one of the most destructive pathogens of rice (Oryza sativa L.) in East Asia. Development of resistant varieties offers a more economical and efficient way to control this disease. In the present study, tests using four inoculation methods were used on 85 backcross inbred lines of Sasanishiki (japonica)/Habataki (indica) to map quantitative trait loci (QTL) conferring resistance to RSV. One QTL on chromosome 3 and two on chromosome 11 were detected, jointly explaining 18?C47?% of the trait variance. The QTL (qSTV11 ^HAB -1 and qSTV11 ^HAB -2) on chromosome 11 were closely linked, and mapped in the intervals G257-RM457 and RM457-RM187, respectively. The stabilities of qSTV11 ^HAB -1 and qSTV11 ^HAB -2 were validated using a set of 38 established chromosome segmental substitution lines. The two QTL, when combined, showed higher resistance than either of them alone in both field and mass inoculation tests, indicating additivity. Fine mapping of the two genes was carried out using 147 recombined F2:3 lines selected from 2,750 secondary F2 plants of the cross Sasanishiki/SL437. Four SSR (simple sequence repeat) and eight InDel (insertion?Cdeletion) markers newly developed to fine-map the two loci. According to the Nipponbare genomic sequence, qSTV11 ^HAB -1 was localized to a 333.2-kb interval which was about 230?kb from the well-known Stvb-i. The other locus, qSTV11 ^HAB -2, which appears to be a new QTL for RSV resistance, was delimited to a 203.9-kb region. Four flanking markers (R15, RM209, R69 and R73) can be used in marker-assisted selection. These results provide an opportunity for map-based cloning of qSTV11 ^HAB -1 and qSTV11 ^HAB -2, thereby promoting the breeding program of RSV resistance.     

Density and distribution of seeds in bottom sediments in Zostera marina beds in Ago Bay,central Japan

The density of Zostera marina L. seeds in bottom sediments was examined to study the reproductive patterns of the Z. marina population in Ago Bay, Mie Prefecture, central Japan.Seeds and seed coats were numerous in Tategami, where the annual type of Z. marina grows. In contrast, seeds were scarce in Hamajima, where the perennial type of Z. marina grows. Bottom sediment was sampled with sediment cores at Tategami in November 2004 and March 2005 to examine density and depth distribution of the seeds. Seeds were found as deep as 8 cm, but no deeper. On the other hand, empty seed coats were found as deep as 16 cm in both months. In the upper layers of the sediment to a depth of 8 cm, the average number of seed coats was 7960 ± 2997 m⁻² in November and 16,318 ± 2922 m⁻² in March. Deeper than 8 cm, the number of seed coats gradually decreased owing to decomposition, and none was found below 16 cm. We used the density of reproductive shoots and number of seeds per spadix in Tategami to estimate the fate of seeds and seed coats of the annual type of Z. marina in bottom sediments: out of the 6000 seeds m⁻² produced annually, 72% disappears from the stand and 28% is buried locally. The density and distribution of Z. marina seeds are among the most important factors in maintenance and propagation of the annual population at Tategami.     

Dosimetric parameters of the new design 103Pd brachytherapy source based on Monte Carlo study

In this study version 5 of the MCNP photon transport simulation was used to calculate the dosimetric parameters for new palladium brachytherapy source design following AAPM Task Group No. 43U1 report. The internal source components include four resin beads of 0.6 mm diameters with ¹⁰³Pd uniformly absorbed inside and one cylindrical copper marker with 1.5 mm length. The resin beads and marker are then encapsulated within 0.8 mm in diameter and 4.5 mm long cylindrical capsule of titanium. The dose rate constant, Λ, line and point-source radial dose function, g_L(r) and g_P(r), and the anisotropy function, F(r,θ) of the IR01-¹⁰³Pd seed have been calculated at distances from 0.25 to 5 cm. All the results are in good agreement with previously published thermoluminescence-dosimeter measured values [3] for the source. The dosimetric parameters calculated in this work showed that in dosimetry point of view, the IR01-¹⁰³Pd seed is suitable for use in brachytherapy of prostate cancer.     

Translocation of iron from soybean cotyledons

Soybean seeds, Glycine max L. Merrill, were produced by plants treated from anthesis to seed maturity with ⁵⁹Fe supplied as ferric ethylenediaminedi (o-hydroxyphenylacetate). Seed coats accounted for 7.4% of dry seed weight and had Fe concentrations 5 times greater than the embryos. After germinating 2 days, cotyledons contained 69.6% and radicles 5.0% of original seed Fe. Fractions of seed Fe unavailable to seedlings were: 19.8% in seed coats, 1.7% in germination paper, 0.1% in the water under germinating seeds, and 3.8% unaccounted for. Every 3 days seedlings received nutrient solution without Fe or with 10 μm ferric ethylenediaminedi (o-hydroxyphenylacetate) and developed as deficient Fe or normal Fe plants. The deficient Fe cotyledons on day 18 retained 13% of the labeled Fe originally present. Cotyledons of normal Fe plants retained 50 to 70% of their original Fe. Moreover, cotyledons of the normal Fe plants accumulated externally supplied Fe and finally contained twice the quantity of Fe originally present. Stem exudate collected above cotyledons of deficient Fe plants contained 5.3 μm⁵⁹Fe. Electrophoresis of exudate showed that most of the ⁵⁹Fe migrated anodically as a single band and was in the position of ferric citrate.     

The Arabidopsis MYB5 Transcription Factor Regulates Mucilage Synthesis,Seed Coat Development,and Trichome Morphogenesis

The Arabidopsis thaliana MYB5 gene is expressed in trichomes and seeds, including the seed coat. Constitutive expression of MYB5 resulted in the formation of more small trichomes and ectopic trichomes and a reduction in total leaf trichome numbers and branching. A myb5 mutant displayed minimal changes in trichome morphology, while a myb23 mutant produced increased numbers of small trichomes and two-branched trichomes. A myb5 myb23 double mutant developed more small rosette trichomes and two-branched trichomes than the single mutants. These results indicate that MYB5 and MYB23 regulate trichome extension and branching. The seed coat epidermal cells of myb5 and myb5 myb23 were irregular in shape, developed flattened columellae, and produced less mucilage than those of the wild type. Among the downregulated genes identified in the myb5 seeds using microarray analysis were ABE1 and ABE4 (α/β fold hydrolase/esterase genes), MYBL2, and GLABRA2. The same genes were also downregulated in transparent testa glabra1 (ttg1) seeds, suggesting that MYB5 collaborates with TTG1 in seed coat development. These genes were upregulated in leaves and roots by ectopically expressed MYB5. The MYBL2, ABE1, and ABE4 promoters were active in seeds, including seed coats, and the latter two also in trichomes. Models of the MYB5 regulatory networks involved in seed coat and trichome development are presented.     

Abscisic Acid and its relationship to seed filling in soybeans

The effect of exogenous abscisic acid (ABA) on the rate of sucrose uptake by soybean (Glycine max L. Merr.) embryos was evaluated in an in vitro system. In addition, the concentrations of endogenous ABA in seeds of three soybean Plant Introduction (PI) lines, differing in seed size, were commpared to their seed growth rates. ABA (10⁻⁷ molar) stimulated in vitro sucrose uptake in soybean (cv `Clay') embryos removed from plants grown in a controlled environment chamber, but not in embryos removed from field-grown plants of the three PI lines. However, the concentration of ABA in seeds of the three field-grown PI lines correlated well with their in situ seed growth rates and in vitro [¹⁴C] sucrose uptake rates.

Across genotypes, the concentration of ABA in seeds peaked at 8.5 micrograms per gram fresh weight, corresponding to the time of most rapid seed growth rate, and declined to 1.2 micrograms per gram at physiological maturity. Seeds of the large-seeded genotype maintained an ABA concentration at least 50% greater than that of the small-seeded genotype throughout the latter half of seed filling. A higher concentration of ABA was found in seed coats and cotyledons than in embryonic axes. Seed coats of the large-seeded genotype always had a higher concentration of ABA than seed coats of the small-seeded line. It is suggested that this higher concentration of ABA in seed coats of the large-seeded genotype stimulates sucrose unloading into the seed coat apoplast and that ABA in cotyledons may enhance sucrose uptake by the cotyledons.

     

Association mapping of grain color,phenolic content,flavonoid content and antioxidant capacity in dehulled rice

Phytochemicals such as phenolics and flavonoids in rice grain are antioxidants that are associated with reduced risk of developing chronic diseases including cardiovascular disease, type-2 diabetes and some cancers. Understanding the genetic basis of these traits is necessary for the improvement of nutritional quality by breeding. Association mapping based on linkage disequilibrium has emerged as a powerful strategy for identifying genes or quantitative trait loci (QTL) underlying complex traits in plants. In this study, genome-wide association mapping using models controlling both population structure (Q) and relative kinship (K) were performed to identify the marker loci/QTLs underlying the naturally occurring variations of grain color and nutritional quality traits in 416 rice germplasm accessions including red and black rice. A total of 41 marker loci were identified for all the traits, and it was confirmed that Ra (i.e., Prp-b for purple pericarp) and Rc (brown pericarp and seed coat) genes were main-effect loci for rice grain color and nutritional quality traits. RM228, RM339, fgr (fragrance gene) and RM316 were important markers associated with most of the traits. Association mapping for the traits of the 361 white or non-pigmented rice accessions (i.e., excluding the red and black rice) revealed a total of 11 markers for four color parameters, and one marker (RM346) for phenolic content. Among them, Wx gene locus was identified for the color parameters of lightness (L*), redness (a*) and hue angle (H ^o). Our study suggested that the markers identified in this study can feasibly be used to improve nutritional quality or health benefit properties of rice by marker-assisted selection if the co-segregations of the marker–trait associations are validated in segregating populations.     

Isolation of a new flavonol glycoside and its effects on the blue color of seed coats ofOphiopogon jaburan

From the blue seed coats ofOphiopogon jaburan, a new flavonol glycoside was isolated as needles and determined to be kaempferol 3-O-β-d-galactoside-4′-O-β-d-glucoside (OK-2) by UV and NMR spectral analyses. OK-2 and kaempfrol 3, 4′-di-O-β-d-glucoside (OK-1), which was detected previously, in the blue seed coat were present in a molar ratio of about 13:7. OK-2 was newly found as a factor causing the blueing effects on ophionin which is a main anthocyanin in the blue seed coats. The mixture of 4.8×10⁻³ M OK-2 and 2.5×10⁻³ M ophionin in Mcllvaine's buffer solution (pH 5.6) showed stable blue color, and the absorption spectrum of the mixture showed two absorption peaks and a shoulder in visible reasion, coinciding with that of the fresh blue seed coat. The effect of ophionin and OK-2 co-pigmentation on the blue color of seed coat ofO. jaburan was discussed.     

The role of the seed coat in adaptation of dimorphic seeds of the euhalophyte Suaeda salsa to salinity

The role of the seed coat in adaptation of dimorphic seeds of the euhalophyte Suaeda salsa to salinity was investigated during germination and early seedling growth. Black and brown seeds were treated with chloroform for 1 min before the extract was used to analyze waxes and the seeds to investigate the protective role of the seed coat under saline conditions. Waxes in black seed coats were more abundant than those in brown seed coats. Salinity (500 mM NaCl) increased the concentration of Na⁺ and decreased the concentration of K⁺ in both black and brown seeds regardless of chloroform treatment. Chloroform treatment alone (in the absence of NaCl) had no effect on the concentration of Na⁺ or K⁺ in black or brown seeds and in the presence of 500 mM NaCl had no effect on the concentration of Na⁺ or K⁺ in brown seeds. However, chloroform treatment increased Na⁺ and decreased K⁺ in black seeds with 500 mM NaCl. A change of MDA (malondialdehyde) concentration in black and brown seeds treated with or without chloroform was similar to the change of Na⁺ concentration. High salinity (1500 mM NaCl) pretreatment for 40 days had a less adverse effect on germination of black seeds compared with brown seeds after they were transferred to fresh water regardless of chloroform treatment. Similar results were found for seedling emergence. In conclusion, a black seed coat may be more protective than a brown seed coat, probably by shielding the embryo from ion toxicity, because of its higher content of waxes. Thus black seeds can better maintain seed viability than brown seeds for extended periods under hypersaline conditions.     

In vitro characterization of the splicing efficiency and fidelity of the RmInt1 group II intron as a means of controlling the dispersion of its host mobile element

Group II introns are catalytic RNAs that are excised from their precursors in a protein-dependent manner in vivo. Certain group II introns can also react in a protein-independent manner under nonphysiological conditions in vitro. The efficiency and fidelity of the splicing reaction is crucial, to guarantee the correct formation and expression of the protein-coding mRNA. RmInt1 is an efficient mobile intron found within the ISRm2011-2 insertion sequence in the symbiotic bacterium Sinorhizobium meliloti. The RmInt1 intron self-splices in vitro, but this reaction generates side products due to a predicted cryptic IBS1* sequence within the 3′ exon. We engineered an RmInt1 intron lacking the cryptic IBS1* sequence, which improved the fidelity of the splicing reaction. However, atypical circular forms of similar electrophoretic mobility to the lariat intron were nevertheless observed. We analyzed a run of four cytidine residues at the 3′ splice site potentially responsible for a lack of fidelity at this site leading to the formation of circular intron forms. We showed that mutations of residues base-pairing in the tertiary EBS3–IBS3 interaction increased the efficiency and fidelity of the splicing reaction. Our results indicate that RmInt1 has developed strategies for decreasing its splicing efficiency and fidelity. RmInt1 makes use of unproductive splicing reactions to limit the transposition of the insertion sequence into which it inserts itself in its natural context, thereby preventing potentially harmful dispersion of ISRm2011-2 throughout the genome of its host.     

Protein chemical shift assignments of the unbound and RNA-bound forms of the alternative splicing factor SUP-12 from C. elegans

The splicing factor SUP-12 from Caenorhabditis elegans binds to regulatory RNA elements in pre-mRNA in order to generate tissue-specific alternative splicing for genes such as the fibroblast growth factor receptor egl-15. In nematode muscle cells, SUP-12 promotes the use of a mutually exclusive exon to impart variant binding specificity to the EGL-15 extracellular protein domain. Here we report the side chain and backbone ¹H, ¹³C and ¹⁵N chemical shift assignments for the bacterially expressed RNA recognition motif domain from SUP-12, both in isolation as well as bound to a short RNA derived from the intron sequence between exon 4 and exon 5B of egl-15. Comparison of protein chemical shift values for both the backbone and side chain nuclei, coupled with secondary chemical shift analysis, reveal initial details of the RNA recognition.