Azeem Iqbal Khan

Biodiversity in the sorghum (Sorghum bicolor L. Moench) germplasm of Pakistan.

Sorghum ranks fifth in worldwide economic importance among cereal crops and is one of the most important summer annual grasses of Pakistan. As it is a very diverse crop, sorghum genetic fingerprinting requires an efficient marker system. We estimated genetic divergence among 29 sorghum (Sorghum bicolor) genotypes, including approved varieties and local and exotic lines collected from different ecological regions of Pakistan, using random amplified polymorphic DNA (RAPD) markers. A total of 125 RAPD loci, with an average of 66 loci per genotype, were used to calculate genetic divergence among these genotypes, of which 119 were polymorphic, showing 95% overall polymorphism. Genetic similarity ranged from 0.36 to 0.92, indicating a relatively broad genetic base. RAPD analysis revealed maximum similarity between the Indian III and K-A-113 sorghum genotypes (both exotic lines), while the F-601 and F-606 were observed to be the most diverse genotypes. Mean band frequency revealed by these RAPD primers ranged from 0.17 to 0.56, with an average of 0.36. The data presented here support the findings that RAPDs can be effectively used for studying genetic diversity in sorghum.

Thermal Characterization of Purified Glucose Oxidase from A Newly Isolated Aspergillus Niger UAF-1

An intracellular glucose oxidase was isolated from the mycelium extract of a locally isolated strain of Aspergillus niger UAF-1. The enzyme was purified to a yield of 28.43% and specific activity of 135 U mg−1 through ammonium sulfate precipitation, anion exchange and gel filtration chromatography. The enzyme showed high affinity for D-glucose with a Km value of 2.56 mM. The enzyme exhibited optimum catalytic activity at pH 5.5. Temperature optimum for glucose oxidase, catalyzed D-glucose oxidation was 40°C. The enzyme showed a high thermostability having a half-life 30 min, enthalpy of denaturation 99.66 kJ mol−1 and free energy of denaturation 103.63 kJ mol−1. These characteristics suggest the use of glucose oxidase from Aspergillus niger UAF-1 as an analytical reagent and in the design of biosensors for clinical, biochemical and diagnostic assays.

Technology transfer for cucumber ( Cucumis sativus L.) production under protected agriculture in uplands Balochistan, Pakistan

2 . Farmer-managed trials also confirmed better effectiveness and efficiency of protected agriculture tunnel against insect pest of cucumber grown under tunnel. The cucumbers so harvested were of higher quality (no insect damage) and were sold at premium prices during the whole production cycle. Proper crop sequencing by considering the market situations as well as physiological circumstances are important for successful production of high value crops including cucumbers round the year. These decisions should be based on the market demand and consumer preference analyses. Higher technical knowledge is also required to manage plant nutrient requirements, choose right kind of hybrids and irrigation scheduling, maintain temperature and humidity and take proper plant protection measures.

Identification and authentication of Rosa species through development of species-specific SCAR marker(s).

Roses (Rosa indica) belong to one of the most crucial groups of plants in the floriculture industry. Rosa species have special fragrances of interest to the perfume and pharmaceutical industries. The genetic diversity of plants based on morphological characteristics is difficult to measure under natural conditions due to the influence of environmental factors, which is why a reliable fingerprinting method was developed to overcome this problem. The development of molecular markers will enable the identification of Rosa species. In the present study, randomly amplified polymorphic DNA (RAPD) analysis was done on four Rosa species, Rosa gruss-an-teplitz (Surkha), Rosa bourboniana, Rosa centifolia, and Rosa damascena. A polymorphic RAPD fragment of 391 bp was detected in R. bourboniana, which was cloned, purified, sequenced, and used to design a pair of species-specific sequence-characterized amplified region (SCAR) primers (forward and reverse). These SCAR primers were used to amplify the specific regions of the rose genome. These PCR amplifications with specific primers are less sensitive to reaction conditions, and due to their high reproducibility, these species-specific SCAR primers can be used for marker-assisted selection and identification of Rosa species.

Genetic diversity in Indian sub-continental landrace cultivars of the genus Triticum L.

Narrowing genetic diversity is a limiting factor in wheat breeding. Popularity of semi dwarf cultivars, developed after green revolution, has resulted in genetic erosion as they replaced indigenous cultivars derived from landraces. These old cultivars have a wealth of useful genes that can be incorporated in the modern cultivars to improve their tolerance level against biotic and abiotic stresses. Genetic analysis of these indigenous and advanced cultivars by the SSR markers has shown greater diversity in this valuable Indian sub-continental germplasm and grouped them into seven units. The Triticum durum lines T2 and T3 were placed in group A, whereas their counterpart T1 was quite distinct. Of the selections, T4 to T9 that was clustered in Group B, T4 to T7 were of Triticum sphaerococcum , whereas the other two were of Triticum aestivum type. T. aestivum cultivar C-248 also appeared to be distinct and could not be grouped with any other cultivar. Based on genetic divergence, therefore, T1 can be used for enhancing diversity in T. durum and C-248 in T. aestivum . Keywords: Triticum , germplasm, simple sequence repeats, genetic diversity

Assessment of biodiversity based on morphological characteristics and RAPD markers among genotypes of wild rose species

Conservation and utilization of the native plant resources is essential for long term sustainability of biodiversity. Wild native resources are adapted to specific and diverse environmental conditions and therefore, these adaptive features can be introduced into modern cultivars either through conventional breeding or advanced molecular genetic techniques. Understanding the genetic make up of the wildly growing plant species and of target desirable genes is a prerequisite for this purpose. Five wild rose ( Rosa L.) genotypes were collected from different locations in northern hilly areas of Pakistan for this study. Different morphological characteristics and PCR based random amplified polymorphic DNA (RAPD) technique was used to find out the diversity and relationship among the genotypes. On morphological basis, Rosa webbiana collected from Muree and Nathia gali showed maximum (83%) similarity, whereas on DNA pattern basis, Rosa brunonii collected from Bansra gali and Sunny bank showed maximum (72%) similarity, while R. webbiana showed maximum diversity among all the species.

Elucidation of thermotolerance diversity in cotton (Gossypium hirsutum L.) using physio-molecular approaches.

Cotton (Gossypium hirsutum) is an important cash crop, but high temperature during its growing season is one of the major factors that limit its productivity. This problem compels plant breeders to breed for heat tolerance, which can help to overcome this challenge. It is very important to make a comprehensive screening of heat-tolerant genotypes so that only the best are chosen. Here we report the combined use of several techniques that can help breeders to screen their germplasm. Twelve cultivated cotton genotypes were evaluated for thermotolerance, using assays that included electrolyte leakage, chlorophyll accumulation and protein profiling, as well as RAPDs to assess genetic diversity. Two genotypes (B-557 and NIAB-78) showed tolerant behavior in three thermotolerance assays. RAPD analysis results showed maximum similarity in a range of 86.7-66.7% between the genotypes MNH-554 and CIM-443. We conclude that combined use should be made of relative electrolyte leakage, chlorophyll stability and differential display with SDS-PAGE to aid in screening for stress tolerance. RAPD-based diversity analysis will further help to improve the efficiency of breeding programs.

Identification and characterization of the Bcl-2- associated athanogene (BAG) protein family in rice

The Bcl-2-associated athanogene (BAG) proteins are involved in the regulation of Hsp70/HSC70 in animals. There are six BAG genes in human that encode nine isoforms with different subcellular locations. Arabidopsis thaliana is reported to contain seven BAG proteins. We searched BAG proteins in Oryza sativa using profile-sequence (Pfam) and profile-profile (FFAS) algorithms and found six homologs. The BAG protein family in O. sativa can be grouped into two classes based on the presence of other conserved domains. Class I consists of four OsBAG genes (1 to 4) containing an additional ubiquitin-like domain, structurally similar to the human BAG1 proteins and might be BAG1 orthologs in plants. Class II consists of two OsBAG genes (5 and 6) containing calmodulin-binding domain. Multiple sequence alignment and structural models of O. sativa BAG proteins showed conservation of surface charge (except OsBAG5) and critical residues for the binding of BAG domain to Hsp70 nucleotide binding domain (NB). Meta analysis of microarray data showed that OsBAG genes are up or down regulated under different stresses (biotic and abiotic). Data obtained from real-time PCR of OsBAG genes under heat stress showed that maximum induction in the expression of all the genes occurred after one hour exposure to heat stress, while reduction in the expression was observed in the following time course and ultimately returned to the basal level at 24 h treatment. These results suggest that OsBAG genes might play important role at the onset of heat stress. A further detailed study may explore the exact function of the members of this gene family and help to make understanding of programmed cell death (PCD) mechanism in plants. Key words : Rice, ubiquitin-like domain, nucleotide-binding domain, real-time PCR.

Estimation of genetic distance between 10 maize accessions with varying response to different levels of soil moisture

Ten maize accessions (NC-9, A50-2, M-14, B-42, NC-3, T-7, N-48-1, B-34, USSR, and WFTMS) were studied to estimate the genetic distance on molecular level by random amplified polymorphic DNA. These accessions were selected on the basis of their variable responses against different levels of moisture. Twenty-five primers were used to test genetic diversity, of which 14 were observed to be polymorphic. Ninety-three loci were amplified; among these, 77 showed polymorphism and the other 16 were monomorphic. Primers A-13 and C-02 gave the most polymorphic bands, while primers A-01 and C-06 gave the fewest polymorphic bands. The genetic similarities of the 10 maize accessions ranged from 82.8 to 54.8%. Accessions USSR and WFTMS showed greatest similarity, and accessions M-14 and B-42 were found more dissimilar than the other accessions. On the basis of cluster analysis, these 10 accessions were classified in two major groups, A and B, and than further divided into sub-groups. The cluster analysis showed that accessions in the same group as well as in the sub-groups were similar in their physical and morphological characters, since the characters are controlled genetically.