Small cardamom genome: development and utilization of microsatellite markers from a draft genome sequence of Elettaria cardamomum Maton.

Small cardamom (Elettaria cardamomum Maton), the queen of spices, is the third most expensive spice in the world after saffron and vanilla, valued highly for its aroma and taste. This perennial herbaceous plant is a native of coastal parts of Southern India and displays a significant amount of morphological diversity. Its genetic potential has not been exploited due to lack of genomic resources limiting our understanding of the genome and important metabolic pathways which give it the economic advantage in the spice industry. Here, we report upon the de novo assembled, draft whole genome sequence of cardamom variety, Njallani Green Gold. We used a hybrid assembly strategy using the reads from the Oxford Nanopore, Illumina and 10x Genomics GemCode sequencing chemistries. The assembled genome length was 1.06 Gb (gigabases) which is close to the estimated genome size of cardamom. More than 75% of the genome was captured in 8000 scaffolds with a N50 of 0.15 Mb. The genome appears to have a high repeat content and 68055 gene models were predicted. The genome is close to Musa species and displays an expansion and contraction in different gene families. The draft assembly was used for in silico mining of simple sequence repeats (SSRs). A total of 2,50,571 SSRs were identified of which 2,18,270 were perfect SSRs and 32,301 were compound SSRs. Among the perfect SSRs, trinucleotides were most abundant (1,25,329) and hexanucleotide repeats appear least (2,380). From the 2,50,571 SSRs mined, 2,27,808 primer pairs were designed based on flanking sequence information. Wet lab validation was performed for 246 SSR loci and based on their amplification profiles, 60 SSR markers were used for diversity analysis of a set of 60 diverse cardamom accessions. The average number of alleles detected per locus were 14.57 with a minimum of 4 and maximum of 30 alleles. Population structure analysis revealed the presence of high degree of admixtures which could primarily be due to cross-pollination prevalent in this species. The SSR markers identified would help in the development of gene or trait-linked markers which can be subsequently used for marker-assisted breeding for crop improvement in cardamom. The information on utilization of the SSR loci for generation of markers has been developed into a public database, 'cardamomSSRdb' that is freely available for use by the cardamom community.

Evolution and co-evolution: insights into the divergence of plant heat shock factor genes.

The Heat Shock Factor (Hsf) genes are widely distributed across the plant kingdom regulating the plant response to various abiotic stresses. In addition to natural selection, breeding and accelerated selection changed the structure and function of Hsf genes. 1076 Hsf genes from 30 genera from primitive algae to the most advanced plant species and major crop plants were used for phylogenetic analysis. The interspecific divergence was studied with 11 members of genus Oryza while intraspecific divergence was studied with sesame pan-genome adapted to diverse ecological niches. B2 genes in eudicots and monocots originated separately while A1 gave rise to the recently evolved Class-C genes and land colonization happened with evolution of A1 genes. An increase in the number of lineages in the Oryza clade with the evolution of AA genome indicated independent domestication and positive selection was observed in > 53% of loci whereas the highly conserved homologues were under purifying selection. The paralogous genes under positive selection exhibited more domain changes for diversified function and increased fitness. A significant co-evolving cluster involving amino acids Phenylalanine, Lysine and Valine played crucial role in maintaining hydrophobic core along with highly conserved Tryptophan residues. A mutation of Glutamic acid to Glutamine was observed in A8 genes of Lamiales affecting protein solvency. Breeding resulted in accumulation of mutations reducing the hydrophobicity of proteins and a further reduction in protein aggregation. This study identify genome duplications, non-neutral selection and co-evolving residues as causing drastic changes in the conserved domain of Hsf proteins. Supplementary information: The online version contains supplementary material available at 10.1007/s12298-022-01183-7.

The chloroplast genome of black pepper (Piper nigrum L.) and its comparative analysis with related Piper species.

Piper nigrum, also known as black pepper, is an economically and ecologically important crop of the genus Piper. It has been titled as the king of spices due to its wide consumption throughout the world. In the present investigation, the chloroplast genome of P. nigrum has been assembled from a whole genome sequence by integrating the short and long reads generated through Illumina and PacBio platforms, respectively. The chloroplast genome was observed to be 161,522 bp in size, having a quadripartite structure with a large single copy (LSC) region of 89,153 bp and a small single copy (SSC) region of 18,255 bp separated by a copy of inverted repeats (IRs), each 27,057 bp in length. Taking into consideration all the duplicated genes, a total of 131 genes were observed, which included 81 protein-coding genes, 37 tRNAs, 4 rRNAs, and 1 pseudogene. Individually, the LSC region consisted of 83 genes, the SSC region had 13 genes, and 18 genes were present in each IR region. Additionally, 216 SSRs were detected and 11 of these were validated through amplification in 12 species of Piper. The features of the chloroplast genome have been compared with those of the genus Piper. Our results provide useful insights into evolutionary and molecular studies of black pepper which will contribute to its further genetic improvement and breeding.

Gene fusions, micro-exons and splice variants define stress signaling by AP2/ERF and WRKY transcription factors in the sesame pan-genome.

Evolutionary dynamics of AP2/ERF and WRKY genes, the major components of defense response were studied extensively in the sesame pan-genome. Massive variation was observed for gene copy numbers, genome location, domain structure, exon-intron structure and protein parameters. In the pan-genome, 63% of AP2/ERF members were devoid of introns whereas >99% of WRKY genes contained multiple introns. AP2 subfamily was found to be micro-exon rich with the adjoining intronic sequences sharing sequence similarity to many stress-responsive and fatty acid metabolism genes. WRKY family included extensive multi-domain gene fusions where the additional domains significantly enhanced gene and exonic sizes as well as gene copy numbers. The fusion genes were found to have roles in acquired immunity, stress response, cell and membrane integrity as well as ROS signaling. The individual genomes shared extensive synteny and collinearity although ecological adaptation was evident among the Chinese and Indian accessions. Significant positive selection effects were noticed for both micro-exon and multi-domain genes. Splice variants with changes in acceptor, donor and branch sites were common and 6-7 splice variants were detected per gene. The study ascertained vital roles of lipid metabolism and chlorophyll biosynthesis in the defense response and stress signaling pathways. 60% of the studied genes localized in the nucleus while 20% preferred chloroplast. Unique cis-element distribution was noticed in the upstream promoter region with MYB and STRE in WRKY genes while MYC was present in the AP2/ERF genes. Intron-less genes exhibited great diversity in the promoter sequences wherein the predominance of dosage effect indicated variable gene expression levels. Mimicking the NBS-LRR genes, a chloroplast localized WRKY gene, Swetha_24868, with additional domains of chorismate mutase, cAMP and voltage-dependent potassium channel was found to act as a master regulator of defense signaling, triggering immunity and reducing ROS levels.

Distinct morpho-physiological and biochemical features of arid and hyper-arid ecotypes of Ziziphus nummularia under drought suggest its higher tolerance compared with semi-arid ecotype.

Tree species in the arid and semi-arid regions use various strategies to combat drought stress. Ziziphus nummularia (Burm. f.) Wight et Arn., native to the Thar Desert in India, is highly drought-tolerant. To identify the most drought-tolerant ecotype of Z. nummularia, one ecotype each from semi-arid (Godhra, annual rainfall >750 mm), arid (Bikaner, 250-350 mm) and hyper-arid (Jaisalmer, <150 mm) regions was selected along with two other Ziziphus species, Ziziphus mauritiana Lamk. and Ziziphus rotundifolia Lamk., and screened for parameters contributing to drought tolerance. Among these, Z. nummularia (Jaisalmer) (CIAHZN-J) was the most drought - tolerant. The tolerance nature of CIAHZN-J was associated with increased membrane stability, root length and number, length of hairs and thorns, root dry/fresh weight ratio, seed germination (at -0.5 MPa), proline content (31-fold), catalase and sugar content (two- to three-fold). Apart from these characteristics, it also exhibited the longest duration to reach highest cumulative drought stress rating, maintained higher relative water content for a longer period of time with reduced leaf size, leaf rolling and falling of older leaves, and displayed sustained shoot growth during drought stress. To determine drought tolerance in Ziziphus, we developed a morphological symptom-based screening technique in this study. Additionally, transcriptome profiling of CIAHZN-J in response to drought revealed the up-regulation of genes involved in sugar metabolism and transport, abscisic acid biosynthesis, osmoregulation, reactive oxygen species homeostasis and maintaining water potential. Expression profiles and semi-quantitative reverse transcription PCR results further correlated with the physiological and biochemical mechanisms. In conclusion, CIAHZN-J is an excellent genetic stock for the identification of drought-responsive genes and can also be deployed in crop improvement programs for drought tolerance.

Microsatellite based DNA fingerprinting and assessment of genetic diversity in bougainvillea cultivars.

Novel microsatellite markers were developed to investigate the genetic diversity and DNA fingerprinting of bougainvillea cultivars. Total of 175 SSRs were designed from over 50,000 SSRs identified in the whole genome sequence data, 33 highly polymorphic markers were identified. These selected SSRs produced a total of 165 alleles with 2 (BOUG-3 and BOUG-50) to 9 (BOUG-69) alleles per loci with an average of 5 alleles per locus. The overall size of the amplified products ranged from 90 bp (BOUG-51 and BOUG-81) to 320 bp (BOUG-162). The gene diversity per locus ranged from 0.13 to 0.91 with a mean of 0.71. Primer BOUG-73 and BOUG-124 exhibited highest gene diversity with greater number of alleles. The mean Nei's genetic diversity index was 0.678 with range of 0.134 (BOUG-77) to 0.958 (BOUG-69). The UPGMA based dendrogram divided the cultivars into seven major clusters. Clustering pattern was more distinct for bract types and variegated cultivars which were also confirmed by PCA scatter plot diagram. The pair-wise genetic distance estimates ranged from 0.089 to 0.86 with an average of 0.56. Each of the 125 cultivar profiled had unique marker profile indicating that the SSR markers identified are useful for identification and differentiation of bougainvillea cultivars. These informative markers identified from the study will be of great utility to assess the genetic diversity, understanding the population structure and in marker assisted breeding for improvement of bougainvillea.

Investigations on diverse sesame (S. indicum L.) germplasm and its wild allies reveal wide variation in antioxidant potential.

Free radicals, the key mediators of a range of oxidative reactions involved in lipid oxidation are responsible for food quality deterioration leading to several health hazards. Antioxidants synthesized naturally or synthetically are capable of preventing oxidation of lipids and other related compounds. However, natural antioxidants have many benefits over synthetic ones. Sesame seeds contain large amount of natural bioactive components with high antioxidant potential. In the present study, 14 accessions of sesame containing wild species and cultivars were investigated. The antioxidant potential of sesame seed meal extract was evaluated by total phenolic content (TPC) method using Folin-Ciocalteu reagent, linoleic acid peroxidation by Ferric thiocyanate method, and free radical scavenging assay with 2,2-diphenyl-1-picryl hydrazyl radical. S. laciniatum showed highest mean values for total polyphenol content with maximum % inhibition of linoleic acid peroxidation on 10th day of course of the reaction span and highest antioxidant scavenging power. S. indicum subsp. malabaricum and S. radiatum also showed high total phenol content and radical scavenging capacity. Among the Sesamum indicum cultivars, Gujarat til 2 showed high TPC and high radical scavenging activity. Higher antioxidant property of Sesamum species in comparison to sesame cultivars highlights the need to utilize the wild genepool for the improvement of cultigens for enhanced nutraceutical value.

Advances in understanding salt tolerance in rice.

KEY MESSAGE: This review presents a comprehensive overview of the recent research on rice salt tolerance in the areas of genomics, proteomics, metabolomics and chemical genomics. Salinity is one of the major constraints in rice cultivation globally. Traditionally, rice is a glycophyte except for a few genotypes that have been widely used in salinity tolerance breeding of rice. Both seedling and reproductive stages of rice are considered to be the salt-susceptible stages; however, research efforts have been biased towards improving the understanding of seedling-stage salt tolerance. An extensive literature survey indicated that there have been very few attempts to develop reproductive stage-specific salt tolerance in rice probably due to the lack of salt-tolerant phenotypes at the reproductive stage. Recently, the role of DNA methylation, genome duplication and codon usage bias in salinity tolerance of rice have been studied. Furthermore, the study of exogenous salt stress alleviants in rice has opened up another potential avenue for understanding and improving its salt tolerance. There is a need to not only generate additional genomic resources in the form of salt-responsive QTLs and molecular markers and to characterize the genes and their upstream regulatory regions, but also to use them to gain deep insights into the mechanisms useful for developing tolerant varieties. We analysed the genomic locations of diverse salt-responsive genomic resources and found that rice chromosomes 1-6 possess the majority of these salinity-responsive genomic resources. The review presents a comprehensive overview of the recent research on rice salt tolerance in the areas of genomics, proteomics, metabolomics and chemical genomics, which should help in understanding the molecular basis of salinity tolerance and its more effective improvement in rice.

GinMicrosatDb: a genome-wide microsatellite markers database for sesame (Sesamum indicum L.).

Molecular breeding in sesame is still at infancy due to limited number of microsatellite markers available and the low level of polymorphism exhibited by them. Therefore, whole genome sequencing was used for development of microsatellite markers so as to ensure availability of substantial number of polymorphic markers for use in marker assisted breeding programs. Whole genome sequencing of sesame variety 'Swetha' was done using Illumina paired-end sequencing and Roche 454 shotgun sequencing technologies (GCA_000975565.1 in GenBank). 'GinMicrosatDb', a genome-wide microsatellite marker database has been developed using the whole genome sequence data of sesame variety 'Swetha'. The database consists of microsatellites localized on both linkage groups and scaffolds with their genomic co-ordinates. It provides five sets of forward and reverse primers for each of the microsatellite loci along with the flanking sequences, primer GC content, product size and melting temperature etc. The distribution of microsatellites can be viewed and selected through a genome browser as well as through a physical map. The newly identified microsatellite markers are expected to help sesame breeders in developing marker tags for traits of economic importance thereby bringing about greater efficiency in marker-assisted selection programs.

Identification of genes associated with stress tolerance in moth bean [Vigna aconitifolia (Jacq.) Marechal], a stress hardy crop.

Moth bean is the most drought and heat tolerant cultigens among Asian Vigna. We performed comparative transcriptome analysis of moth bean cultivar "Marumoth" under control and stress condition. De novo transcriptome assembly was carried out by using Velvet followed by Oases softwares. Differential expression analyses, SSR identification and validation and mapping of pathways and transcription factors were conducted. A total of 179,979 and 201,888 reads were generated on Roche 454 platform and 48,617,205 and 45,449,053 reads were generated on ABI Solid platform for the control and stressed samples. Combined assembly from Roche and ABI Solid platforms generated 16,090 and 15,096 transcripts for control and stressed samples. We found 1287 SSRs and 5606 transcripts involved in 179 pathways. The 55 transcription factor families represented 19.42% of total mothbean transcripts. In expression profiling, ten transcripts were found to be up-regulated and 41 down-regulated while 490 showed no major change under moisture stress condition. Stress inducible genes like Catalase, Cyt P450 monooxygenase, heat shock proteins (HSP 90 and HSP 70), oxidoreductase, protein kinases, dehydration responsive protein (DRP), universal stress protein and ferridoxin NADH oxidoreductase genes were up-regulated in stressed sample. Genes which might be involved in moisture stress tolerance in moth bean were identified and these might be useful for stress tolerance breeding in moth bean and other related crops.

Analysis of spatial distribution of genetic diversity and validation of Indian foxtail millet core collection.

Foxtail millet [Setaria italica (L.) P. Beauv.] is an important small millet, grown as a short duration, drought tolerant crop across the world. This crop can be grown on wide ranges of soil conditions and has an immense potential for food and fodder in rainfed and arid regions of the India. In the present study, 31 primer pairs (27 SSR and 4 EST-SSR) were used to analyse the genetic diversity in 223 core collection accessions. Analysis resulted in detection of a total of 136 alleles with an average of 4.38 alleles per locus. Among these 136 alleles, 22 were rare, 70 were common and 44 were frequent. The PIC value ranged from 0.01 to 0.86 with an average of 0.31. The average number of observed alleles ranged from 2.0 (northern hills of India accessions) to 4.06 (exotic) with an average of 2.72. The mean Shannon's Information Index ranged from 0.44 (northern hills of India) to 0.69 (exotic) with an average of 0.52. Pair-wise Fst values indicated little to moderate genetic differentiation among the group of accessions. UPGMA clustering grouped the accessions into two major groups while analysis for population substructure indicated presence of four subpopulations. However there was no statistically well supported grouping of the accessions based on eco-geographic specificities. The core collection designated here represented substantial genetic diversity at molecular level, hence may be a good source of diversity for use in foxtail improvement programs in the region.

Tracking sesamin synthase gene expression through seed maturity in wild and cultivated sesame species--a domestication footprint.

Sesamin and sesamolin are the major oil-soluble lignans present in sesame seed, having a wide range of biological functions beneficial to human health. Understanding sesame domestication history using sesamin synthase gene expression could enable delineation of the sesame putative progenitor. This report examined the functional expression of sesamin synthase (CYP81Q1) during capsule maturation (0-40 days after flowering) in three wild Sesamum species and four sesame cultivars. Among the cultivated accessions, only S. indicum (CO-1) exhibited transcript abundance of sesamin synthase along with high sesamin content similar to S. malabaricum, while the other cultivated sesame showed low expression. The sesamin synthase expression analysis, coupled with quantification of sesamin level, indicates that sesamin synthase was not positively favoured during domestication. The sesamin synthase expression pattern and lignan content, along with phylogenetic analysis suggested a close relationship of cultivated sesame and the wild species S. malabaricum. The high genetic identity between the two species S. indicum and S. malabaricum points towards the role of the putative progenitor S. malabaricum in sesame breeding programmes to broaden the genetic base of sesame cultivars. This study emphasises the need to investigate intraspecific and interspecific variation in the primary, secondary and tertiary gene pools to develop superior sesame genotypes.

CYTOGENETICAL TREATISE OF INDIAN REPRESENTATIVE SPECIES OF CUCUMIS. A KARYOTYPIC APPROACH.

Karyomorphological studies have been carried out in nine species and five varieties of the genus Cucumis representing Indian gene pool. The present investigations reveal the occurrence of two somatic chromosome numbers 2n = 14, 24 in the genus. C. ritchiei and C. indicus the two new species, were found to be having somatic chromosome numbers of 2n = 24 and 2n = 20 respectively. The wild species viz. C. hystrix, C. setosus, C. prophetarum, C. dipsaceus, C. indicus have very less number of median-centromeric chromosomes, high asymmetry indices, while melon groups have intermediate number of median -centromeric chromosomes. C. sativus, C. callosus, C. ritchiei show lesser number median-cen-tromeric chromosomes and very less asymmetry indices. The importance of karyotypic variation with respect to speciation within the genus Cucumis have been discussed.

Value addition in sesame: A perspective on bioactive components for enhancing utility and profitability.

Sesame seed is a reservoir of nutritional components with numerous beneficial effects along with health promotion in humans. The bioactive components present in the seed include vital minerals, vitamins, phytosterols, polyunsaturated fatty acids, tocopherols and unique class of lignans such as sesamin and sesamolin. The presence of phenylpropanoid compounds namely lignans along with tocopherols and phytosterols provide defense mechanism against reactive oxygen species and increases keeping quality of oil by preventing oxidative rancidity. In this article, we have reviewed the nutraceutical, pharmacological, traditional and industrial value of sesame seeds with respect to bioactive components that hold high antioxidant value. Valuable information on superior functional components of sesame will strongly promote the use of sesame seeds in the daily diet world-wide. In spite of huge repertoire of sesame germplasm collection, limited research efforts on the use of conventional and biotechnological methodologies have resulted in minimal success in developing nutritionally superior cultivars. In consequence, value addition efforts in sesame would enable development of genotypes with high antioxidant activity and subsequently prevention of free radical related diseases. Modification of bioactive components in sesame would enable production of stabilized sesame oil with enhanced shelf life and better market value.

Analysis of molecular genetic diversity in a representative collection of foxtail millet [Setaria italica (L.) P. Beauv.] from different agro-ecological regions of India.

Foxtail millet [Setaria italica (L.) P. Beauv.], an important crop of East Asia is known for its drought tolerance and was once an indispensible crop of vast rainfed areas in semi-arid regions in India. In India it is cultivated in Andhra Pradesh, Karnataka, Maharashtra, Tamil Nadu, Rajasthan, Madhya Pradesh, Uttar Pradesh and north eastern states. The grain finds use in several local recipes such as roti (bread), jaula, singal, sirol. Foxtail millet grain contains 12.3 % protein, 4.7 % fat, 60.6 % carbohydrates, and 3.2 % ash. The present study was conducted to analyse the genetic diversity among foxtail accessions from different states of India and a few exotic accessions using RAPD and ISSR techniques and identify diverse accessions for use in variety improvement programmes. A set of 125 foxtail millet accessions selected from 11 different agro-ecological regions of India were analyzed using random amplified polymorphic DNA (RAPD) and inter simple sequence repeat (ISSR) marker techniques. A total of 146 (115 RAPD and 31 ISSR) scoreable markers were generated with 16 RAPD and four ISSR primers. The dendrogram generated using Nei's genetic distances and principal component analyses revealed presence of two clusters and two subclusters in group I. The accessions from Andhra Pradesh, Karnataka, Maharashtra and Uttarakhand were more diverse since they were distributed in both the clusters. There was no clear geographical differentiation observable. The bootstrap support for the major groups identified was strong (above 80 %) indicating good statistical support. The average value of Nei and Li's genetic distance was lowest (0.081) for accessions from West Bengal while the collections from Karnataka showed highest dissimilarity (average genetic distance = 0.239). The average genetic distance for all 125 accessions together was 0.177 indicating presence of only moderate genetic diversity in the collections. The analysis of molecular variance indicated that only 2.76 % variation was explained by variations among the groups and 11.55 % among populations within groups. However the percentage of variation observed within populations was high (85.68). The value of Fst was observed to be very low (0.028) indicating low differentiation of the accessions analysed. The population genetic analysis carried out indicates that highest number of alleles per locus (1.745 ± 0.438) was observed for Andhra Pradesh with 35 accessions. When four eco-geographic regions were considered, the southern region comprising AP, Karnataka and TN showed the highest number of alleles per locus (1.787 ± 0.411). The value of Gst was lowest for south (0.123) and highest for central west (0.455). This indicated that all the landraces from south share common alleles. The gene flow between the accessions from different regions was also observed to be high with the highest migration (3.557) recorded for south.

Evaluation of computed tomography virtual bronchoscopy in paediatric tracheobronchial foreign body aspiration.

OBJECTIVE: Virtual bronchoscopy is a noninvasive technique which provides an intraluminal view of the tracheobronchial tree. This study aimed to evaluate this technique in comparison with rigid bronchoscopy, in paediatric patients with tracheobronchial foreign bodies undetected by plain chest radiography. METHODS: Plain chest radiography was initially performed in 40 children with suspected foreign body aspiration. Computed tomography virtual bronchoscopy was performed in the 20 in whom chest radiography appeared normal. Virtual bronchoscopic images were obtained. All patients underwent rigid bronchoscopy performed by an otolaryngologist blinded to the computed tomography virtual bronchoscopy findings, within 24 hours. Virtual bronchoscopic findings were then compared with the results of rigid bronchoscopy. RESULTS: In 12 patients, foreign bodies detected by virtual bronchoscopy were confirmed by rigid bronchoscopy. In one case, a mucous plug was perceived as a foreign body on virtual bronchoscopy. In another case, a minute foreign body was missed on virtual bronchoscopy. The following parameters were calculated: sensitivity, 92.3 per cent; specificity, 85.7 per cent; validity, 90 per cent; positive likelihood ratio, 6.45; and negative likelihood ratio, 0.089. CONCLUSION: In the presence of a positive clinical diagnosis and negative chest radiography, computed tomography virtual bronchoscopy must be considered in all cases of tracheobronchial foreign body aspiration, in order to avoid needless rigid bronchoscopy. Computed tomography virtual bronchoscopy is particularly useful in screening cases of occult foreign body aspiration, as it has high sensitivity, specificity and validity.

SSR and AFLP based genetic diversity of soybean germplasm differing in photoperiod sensitivity.

Forty-four soybean genotypes with different photoperiod response were selected after screening of 1000 soybean accessions under artificial condition and were profiled using 40 SSR and 5 AFLP primer pairs. The average polymorphism information content (PIC) for SSR and AFLP marker systems was 0.507 and 0.120, respectively. Clustering of genotypes was done using UPGMA method for SSR and AFLP and correlation was 0.337 and 0.504, respectively. Mantel's correlation coefficients between Jaccard's similarity coefficient and the cophenetic values were fairly high in both the marker systems (SSR = 0.924; AFLP = 0.958) indicating very good fit for the clustering pattern. UPGMA based cluster analysis classified soybean genotypes into four major groups with fairly moderate bootstrap support. These major clusters corresponded with the photoperiod response and place of origin. The results indicate that the photoperiod insensitive genotypes, 11/2/1939 (EC 325097) and MACS 330 would be better choice for broadening the genetic base of soybean for this trait.

SSR and AFLP based genetic diversity of soybean germplasm differing in photoperiod sensitivity

Forty-four soybean genotypes with different photoperiod response were selected after screening of 1000 soybean accessions under artificial condition and were profiled using 40 SSR and 5 AFLP primer pairs. The average polymorphism information content (PIC) for SSR and AFLP marker systems was 0.507 and 0.120, respectively. Clustering of genotypes was done using UPGMA method for SSR and AFLP and correlation was 0.337 and 0.504, respectively. Mantel's correlation coefficients between Jaccard's similarity coefficient and the cophenetic values were fairly high in both the marker systems (SSR = 0.924; AFLP = 0.958) indicating very good fit for the clustering pattern. UPGMA based cluster analysis classified soybean genotypes into four major groups with fairly moderate bootstrap support. These major clusters corresponded with the photoperiod response and place of origin. The results indicate that the photoperiod insensitive genotypes, 11/2/1939 (EC 325097) and MACS 330 would be better choice for broadening the genetic base of soybean for this trait.

Cryopreservation of asian Dioscorea bulbifera l. and D. alata l. by vitrification: importance of plant growth regulators.

The aim of this study was to develop cryopreservation protocols for Asian races of Dioscorea bulbifera and D. alata with high survival and plant regeneration after cryopreservation. Using a vitrification procedure, survival of shoot tips postcryopreservation of up to 89% in D. bulbifera and up to 82% in D. alata were recorded when excised shoot tips were pretreated overnight with 0.3 M sucrose in MS medium, followed by loading with 2 M glycerol plus 0.4 M sucrose for 20 min at 25 degrees C, exposure to PVS2 solution for 90 min at 0 degrees C, immersion in liquid nitrogen for 1 h, rewarming at 40 degrees C for 2 min, unloading in medium with 1.2 M sucrose for 20 min and culturing on growth recovery medium. During growth recovery, 58% shoot regeneration was obtained in D. bulbifera when cryopreserved shoot tips were initially cultured for 40 days on MS medium with 1.5 mg/L BAP, 0.15 mg/L NAA and 0.2 mg/L GA3 followed by culturing on a medium with 0.05 mg/L BAP and 0.15 mg/ L NAA. However, a maximum of 39% shoot regeneration was recorded in D. alata when cryopreserved shoot tips were initially cultured for 40 days on medium M2 (MS containing 1/5 NH4NO3 and 40 g/L sucrose) supplemented with 1.0 mg/L BAP, 1.0 mg/L zeatin, 0.15 mg/L IAA and 0.2 mg/L GA3. Subsequently, the regenerating shoots were cultured for 30 days on medium M2 with 1.0 mg/L BAP, 0.3 mg/L zeatin, 0.02 mg/L NAA and 0.2 mg/L GA3 followed by culturing for another 30 days on medium with 0.5 mg/L BAP, 0.02 mg/L NAA and 0.2 mg/L GA3. Finally, transfer onto medium with 0.05 mg/L BAP and 0.15 mg/L NAA stimulated production of fully grown plantlets. Alteration of post-thaw culture media with plant growth regulators and their application at various stages of growth recovery was crucial for regeneration of shoot tips and formation of plantlets in D. alata.

Cortical mastoidectomy in quiescent, tubotympanic, chronic otitis media: is it routinely necessary?

OBJECTIVE: This study aimed to compare outcomes for mastoidotympanoplasty and for tympanoplasty alone in cases of quiescent, tubotympanic, chronic, suppurative otitis media. STUDY DESIGN: Single-blinded, randomised, controlled study within a tertiary referral hospital. METHODS: Sixty-eight cases were randomly allocated into two groups. In group one, 35 ears underwent type one tympanoplasty along with cortical mastoidectomy. In group two, 33 ears underwent type one tympanoplasty alone. Outcome measures were as follows: perforation closure and graft uptake, hearing improvement, disease eradication, and post-operative complications. RESULTS: There were no statistically significant differences in hearing improvement, tympanic perforation closure, graft uptake or disease eradication, comparing the two groups at three and six months post-operatively. CONCLUSION: Mastoidotympanoplasty was not found to be superior to tympanoplasty alone over a short term follow-up period. Hence, it may not be necessary to undertake routine mastoid exploration at this stage of disease.