Regeneration of plants from long-term cultures of Oryza sativa L.

Root and embryo derived callus tissues of rice grown on sucrose alone as carbon source lost their ability to organise shoots by 75 and 100 days in culture respectively. Along with 2% sucrose, either 3% sorbitol or 3% mannitol was found to be necessary in the growth medium for the callus to regenerate whole plants over a period of 1400 days without any decline in the shoot forming ability. Our results indicated that incorporation of sorbitol or mannitol in the callus proliferating medium provides long-term totipotent rice cultures with a high frequency (50–60%) of shoot differentiation.     

The ubiquitin-binding domain of DNA polymerase η directly binds to DNA clamp PCNA and regulates translesion DNA synthesis

DNA polymerase eta (Polη) is a unique translesion DNA synthesis (TLS) enzyme required for the error-free bypass of ultraviolet ray (UV)-induced cyclobutane pyrimidine dimers in DNA. Therefore, its deficiency confers cellular sensitivity to UV radiation and an increased rate of UV-induced mutagenesis. Polη possesses a ubiquitin-binding zinc finger (ubz) domain and a PCNA-interacting-protein (pip) motif in the carboxy-terminal region. The role of the Polη pip motif in PCNA interaction required for DNA polymerase recruitment to the stalled replication fork has been demonstrated in earlier studies; however, the function of the ubz domain remains divisive. As per the current notion, the ubz domain of Polη binds to the ubiquitin moiety of the ubiquitinated PCNA, but such interaction is found to be nonessential for Polη''s function. In this study, through amino acid sequence alignments, we identify three classes of Polη among different species based on the presence or absence of pip motif or ubz domain and using comprehensive mutational analyses, we show that the ubz domain of Polη, which intrinsically lacks the pip motif directly binds to the interdomain connecting loop (IDCL) of PCNA and regulates Polη''s TLS activity. We further propose two distinct modes of PCNA interaction mediated either by pip motif or ubz domain in various Polη homologs. When the pip motif or ubz domain of a given Polη binds to the IDCL of PCNA, such interaction becomes essential, whereas the binding of ubz domain to PCNA through ubiquitin is dispensable for Polη''s function.     

Isolation of novel lipolytic genes from uncultured bacteria of pond water

Metagenomic libraries give access to gene pool of bacteria present in environmental samples avoiding the culture bias. A metagenomic library of pond water microbial assemblage in plasmid vector containing about 532 Mb of community DNA was prepared. Screening of a part of the unamplified library resulted in isolation of 11 unique lipolytic clones with an ability to hydrolyze tributyrin. DNA sequence of the lipolytic genes varied in G+C composition from 57% to 75%. Twelve lipolytic genes encoding proteins with 25-70% amino acid identity with proteins in the databases were identified. Ten of the encoded proteins belonged to seven known lipolytic protein families. One of the proteins was similar to recently identified esterase BioH. A lipolytic protein with high similarity to yet uncharacterized alpha/beta hydrolase protein family abh_upf0017 was identified from one of the clones. Conserved motif for lipolytic enzymes GXSXG, conserved aspartic and histidine residues were identified in this encoded protein.     

A rapid protocol for somatic embryogenesis from immature leaflets of groundnut (Arachis hypogaea L.)

Summary Plant regeneration via somatic embryogenesis was developed in two groundnut varieties. Somatic embryogenesis was induced from immature leaflets on MS medium with different concentrations of the auxins 2,4-dichlorophenoxyacetic acid (2,4-D) or naphthaleneacetic acid (NAA) in combination with 0.5 mg/l of the cytokinin BA. The highest frequency of somatic embryo formation occurred on MS medium fortified with 20 mg 2,4-D per l. Of the two auxins tested individually 2,4-D was more effective for induction of embryogenesis as well as production of embryos. Embryo development and maturation was achieved on MS medium supplemented with N⁶-benzyladenine (BA) (0.5–2.0 mg/l) and 2,4-D (0.5 mg/l). Plant conversion frequency from somatic embryos was highest in presence of 2.0 mg BA per l and 0.5 mg NAA per l. The frequency of embryogenesis and plant regeneration was higher in the VRI-2 cultivar than in the other cultivar tested. Regenerated plants were transferred to soil, grown to maturity, and produced viable seeds.     

Base Pairing between U3 Small Nucleolar RNA and the 5′ End of 18S rRNA Is Required for Pre-rRNA Processing

The loop of a stem structure close to the 5' end of the 18S rRNA is complementary to the box A region of the U3 small nucleolar RNA (snoRNA). Substitution of the 18S loop nucleotides inhibited pre-rRNA cleavage at site A(1), the 5' end of the 18S rRNA, and at site A(2), located 1.9 kb away in internal transcribed spacer 1. This inhibition was largely suppressed by a compensatory mutation in U3, demonstrating functional base pairing. The U3-pre-rRNA base pairing is incompatible with the structure that forms in the mature 18S rRNA and may prevent premature folding of the pre-rRNA. In the Escherichia coli pre-rRNA the homologous region of the 16S rRNA is also sequestered, in that case by base pairing to the 5' external transcribed spacer (5' ETS). Cleavage at site A(0) in the yeast 5' ETS strictly requires base pairing between U3 and a sequence within the 5' ETS. In contrast, the U3-18S interaction is not required for A(0) cleavage. U3 therefore carries out at least two functionally distinct base pair interactions with the pre-rRNA. The nucleotide at the site of A(1) cleavage was shown to be specified by two distinct signals; one of these is the stem-loop structure within the 18S rRNA. However, in contrast to the efficiency of cleavage, the position of A(1) cleavage is not dependent on the U3-loop interaction. We conclude that the 18S stem-loop structure is recognized at least twice during pre-rRNA processing.     

MUC4 and MMP7 in saliva and gingival crevicular fluid (GCF) in adolescents at West Bengal,India

Mucin 4 (MUC4) and matrix metalloproteinase 7 (MMP7) have been reported to be associated with chronic periodontitis as seen in gingival tissue biopsies. Therefore, it is of interest to estimate the levels of MUC4 and MMP7 in saliva and gingival crevicular fluid (GCF) samples of periodontitis in adolescents patients at West Bengal, India. MUC4 levels were significantly lower in saliva and GCF from periodontitis patients compared to healthy controls. However, MMP7 levels were found to be significantly higher in saliva and GCF from periodontitis patients. Thus, MUC4 and MMP7 are biomarkers for periodontitis diagnosis and towards further consideration.     

The effect of hycanthone and maleic hydrazide on the frequency of micronuclei in the bone-marrow erythrocytes of mice.

Male Swiss mice were assigned to 6 groups of either 3 or 4 animals each. 3 groups were given hycanthone methanesulfonate intraperitoneally, at 40, 80 or 120 mg/kg, respectively; the dose was repeated after an interval of about 24 h. At the same time 2 groups received maleic hydrazide at 100 or 200 mg/kg, and the remaining group was given dimethyl sulfoxide which was used as a solvent for both drugs. 6 h after the second injection, the mice were killed and bonemarrow preparations were made. Hycanthone induced a significant increase in the frequency of micronuclei in the polychromatic erythrocytes and suppressed the P/N ratio significantly. However, there was no dose-response relationship. Maleic hydrazide, on the other hand, failed to influence the incidence of micronuclei or the ratio of poly- to normo-chromatic erythrocytes.