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1. Journal of Cell and Tissue Research 15(1) 4691-4702 (2015)
An over view of intra-cellular membrane trafficking proteins in animals and plants
Saini, K. M.
Department of Biotechnology, Directorate of Groundnut Research, Junagadh 362 001, Gujarat.
E. mail: kunalmsaini@gmail.com
Abstract: Intra-cellular membrane trafficking in the eukaryotic cells is vital for the cellular functioning and homeostasis. Protein transport is one of the most intricate and essential cellular process requiring high fidelity and specificity. Proteins once synthesized at the endoplasmic reticulum need to be transported to the different compartments in the cell via highly specialized and efficient routes of the so called intra-cellular membrane trafficking pathways. The steps involved in trafficking starts with the formation of vesicles at the donor membrane, transport to the correct membrane and finally its docking and fusion to the acceptor membrane. All these processes require an array of highly specialized proteins. Ras super-family of GTPases, especially Rab GTPases play an important role in signalling and vesicle trafficking, whereas, SNARE proteins play a vital and distinct role in docking and fusion at the organelle’s membrane. The present review highlights the ever increasing importance of the these proteins in identification and treatment of several diseases and their importance in the target recognition, inhibitions, and their importance in the fields of synaptic vesicle recycling in neurobiology and in oncology dealing with various carcinomas in case of animals as well as their role and application in the field of plant sciences.
Key words: Intra-cellular membrane-trafficking, vesicle-trafficking, GTPases
2. Journal of Cell and Tissue Research 15(1) 4703-4710 (2015)
In vitro cytotoxicity assessment of Graphene quantum dots in caprine Wharton’s jelly derived mesenchymal stem cells
Dar, R.M., Gade, N.E.,Mishra, O.P., Khan, J.R., Kumar, V. and Patyal, A.
Department of Veterinary Physiology & Biochemistry, College of Veterinary Science & A.H., Anjora, Durg 491 001, Chhattisgarh. E. mail: nitingadeivri@gmail.com,
Abstract: Present study was designed to investigate the cytotoxicity of graphene quantum dots (GQD) in caprine Wharton’s jelly derived mesenchymal stem cells (WJ-MSCs). In vitro caprine WJ-MSCs were isolated, cultured up to 3rd passage and characterized by alkaline phosphatase staining. Cytotoxicity of GQD nanoparticle was evaluated by observing alterations in cell morphology, cell viability, growth kinetics, population doubling time, colony forming unit (CFU) assay and MTT assay in different treatment groups (low dose: 10 µg/ml, moderate dose: 50 µg/ml and high dose: 100 µg/ml of GQD concentrations) in comparison with control group (without GQD). High and moderate dose of GQD exposed cells showed morphological changes and cell viability assay demonstrated a highly significant (P<0.01) decrease in the number of viable cells. Declined growth rate and highly significant (P<0.01) increase in population doubling time (PDT) of high and moderate dose treated cells were observed as compared to control group and low dose treatment group. The results of the CFU assay revealed that the cells exposed to high and moderate dose of GQD showed declined number of clones than control group and the low dose treated cells. The absorbance values of MTT assay showed non-significant (P>0.05) decrease in high and moderate dose treatment groups. In conclusion, results indicates that the dose of the nanoparticle is a potential contributing factor in exhibiting toxicity, as higher and moderate dose (100 μg/ml & 50 μg/ml) are more toxic than lower dose (10 μg/ml) in caprine WJ-MSCs.
Key words: Graphene, Mesenchymal stem cells, Caprine
3. Journal of Cell and Tissue Research 15(1) 4711-4714 (2015)
Motility and functional membrane integrity of buck spermatozoa with soyabean lecithin based extender
Lekshmi Bhai, K., Joseph, M., Behera, S.,Harshan, H. M., Ghosh Aravinda, K.N. and Raghavan, K.C.
Department of animal reproduction, gynaecology and obstetrics, college of veterinary and animal sciences, kerala veterinary and animal sciences university mannuthy, thrissur, kerala 680651.
E. mail: bhai04vet@gmail.com,
Abstract: The study was conducted using 48 ejaculates collected following the double ejaculate regime from two adult malabari bucks maintained at the ai centre in the department of animal reproduction gynaecology and obstetrics. the collected and pooled semen ejaculates were divided into two groups. group i was extended with egg yolk based extender and group ii with soyabean lecithin based extender. pre freeze assessment of semen quality was done just prior to conventional method of freezing. after 24 hours of freezing, post-thaw motility and functional membrane integrity were assessed. soyabean lecithin based extender resulted in significantly higher pre freeze and post-thaw seminal parameters of buck spermatozoa.
Key words: spermatozoa, malabari buck, soyabean lecithin
4.Journal of Cell and Tissue Research 15(1) 4715-4720 (2015
Identification of chicken and dog meat by mitochondrial 12S rRNA gene PCR-RFLP
Patil, M.R., Bannalikar, A.S. and Arora, A.V.
Department of Animal Biotechnology, Bombay Veterinary College, Mumbai,400 012, Maharashtra; 1Mumbai University Mumbai 400 012, Maharashtra. E. mail: dr.mayurapatil@gmail.com,
Abstract: The fraudulent practice of cheap meat in costly meat is common nowadays. Therefore, there is need of precise identification of the meat species. We included in our study dog meat, as it is the most easily available cheap meat for adulteration. For specific identification of origin of meat animal species, we evaluated the common meat animal species including dog meat by PCR –RFLP technique. All the samples from all species resulted amplicons of 456 bp on PCR except in chicken we obtained 444 bp product. These amplicons further subjected to restriction digestion with five enzymes including AluI, HhaI, ApoI, BspTI and MboII, which resulted in speces specific restriction profiles which enable the identification. We were able to differenciate chicken and dog meat with AluI and MboII alone, whereas AluI, HhaI, ApoI, BspTI were specific to beef, buffalo meat, mutton and chevon respectively.
Key words: Meat identification, Mitochodial 12S rRNA gene
5.Journal of Cell and Tissue Research 15(1) 4721-4726 (2015)
K+- Induced vasodilation in middle uterine artery of pregnant Capra hircus is mediatedbyaugmentation NA+-K+ ATPase / K+ channel activity
Rout, S.R. and Parija, S.C.
Department of Pharmacology and Toxicology, College of Veterinary Science and Animal Husbandry, Orissa University of Agriculture and Technology, Bhubaneswar 751003, Orissa.
E mail: profscparijaouat4691@gmail.com,
Abstract: The present study is designed to understand the relative role of Na+-K+-ATPase and K+ channels in mediating vasorelaxation in middle uterine artery of non-pregnant and pregnant goat (Capra hircus). Branches of middle uterine artery nearby uterus was cut into 1-1.5 mm circular rings and mounted in a thermostatically controlled (37 ± 0.5°C) organ bath containing 20 ml of MKHS (pH 7.4), with continuous aeration under 1.5 g tension for 90 min. KCl (1 μM-10mM) was added cumulatively to K+ free MKHS-precontracted rings in absence or presence of ouabain (0.1μM) or Barium (30μM) or 4-aminopyridine (1ìM). K+-induced vasorelaxation was significantly (p<0.001) greater in pregnant goat (EBmax 74.66± 1.30 % ,EC50 10.5 ìM ) than in non-pregnant one (EBmax: 94.45± 2.1% ,EC50: 3.91ìM). Ouabain didn’t alter the basal tone of MUA ring of both goats. In pregnant goat the vasorelaxation of K+ was significantly attenuated with increase in EBmax and EC50 in presence of ouabain (Emax 90.51± 2.13%, EC50 76.0 ìM), 4-AP (Emax 85.48 ± 1.14 %, EC50 20.2 ìM), Ouabain and Ba2+ (Emax 96.33±2.53%, EC50 106ìM) but not in presence of Ba2+. In non-pregnant goat, vasorelaxation of K+ was not altered by these ligands. In conclusion, increased K+-induced vasodilation of middle uterine artery of pregnant goat is due to the augmentation of functional activity or expression of ouabain sensitive Na+-K+-ATPase and Kv channels.
Key words: Na+-K+ATPase, K+ channels, Middle uterine artery, Goat
6. Journal of Cell and Tissue Research 15(1) 4727-4736 (2015)
The effect of telmisartan versus garlic on renal cortex of experimentally induced hypertension in rats: A biochemical, histological and immunohistochemical study
Faruk, E. M., Salem, M. Y. and El-Eraky El-Azab, N.
Department of Histology and Cell Biology, Benha Faculty of Medicine, Egypt.
E. mail: faruk_eman@yahoo.com
Abstract: Different metabolic disorders including hypertension cause renal damage and increase the risk of cardiovascular events.Telmisartan,is an angiotensin II receptor blocker. Garlic in different forms has antioxidant properties. This study was designed to determine the possible protective effect of telmisartan and garlic on renal cortex of experimentally induced hypertension in rats. Forty adult male albino rats were divided into two groups: Group I; control group (n=10), Group II (n=30); hypertension group. Group II divided into three equal subgroups. Subgroup Ιιa (hypertension only group), Subgroup IIb (hypertension with telmisartan group); rats were given telmisartan daily oral dose 10 mg/kg/day for 4 weeks from the start of experiment with the induction of hypertension, and subgroup IIc (hypertension with garlic group); rats were given garlic extract i.p. daily dose 500 mg/kg/day for 4 weeks from the start of experiment with the induction of hypertension. Blood and kidney samples and measuring of systolic blood pressure were performed after 4 weeks from the start of experiment. Paraffin sections of kidneys were prepared for histological; H&E and PAS staining and immunohistochemical study for kappa B (kB). The results revealed that the hypertension only group (subgroup IIa) showed high systolic blood pressure (mmHg), elevated blood urea level and serum creatinine with alterations in morphology of renal glomeruli and tubules and strong NF-kBexpression. Hypertension with telmisartan group (subgroup IIb) showed significant decreased (P<0.05) systolic blood pressure, blood urea and serum creatinine, NF-kB expression compared with subgroup IIa and showed near normal in morphology of renal glomeruli and tubules. Hypertension with garlic group (subgroup IIc) showed significant decreased (P<0.05) blood urea, serum creatinine, NF-kB expression, insignificant decreased systolic blood pressure with improvement in morphology of renal glomeruli and tubules compared with subgroup IIa. Telmisartan was more effective in hypertension, preservation of normal kidney function and histology than garlic.
Key words: Hypertension, Renal cortex, Telmisartan, Garlic
7.Journal of Cell and Tissue Research 15(1) 4737-4746 (2015)
Effects of thyme extract on hepatotoxicity induced by fenitrothion in adult male mice: A histological, histochemical and immmunohistochemical study
El-Naggar, S.M., Selim, A. A. and El-Mahalaway, A.M.
Department of Basic Sciences, IbnSina National College for Medical Studies –Jeddah 21323, Kingdom of Saudi Arabia. E. mail: abeerelmahalaway@yahoo.com,
Abstract: Fenitrothion (Sumithion) is an organophosphorus pesticide, used worldwide in agricultural practice. It is toxic to animals and humans. Thyme commonly used as food flavor and germ killer. The current work was undertaken with the objective to evaluate the protective effects of Thyme extract on hepatotoxicity induced by fenitrothion (sumithion) in adult male mice. Forty male mice were included in study and divided into 4 groups (each group included10 mice). Group I: the animals received tape water for 4 weeks. Group II: The animals received daily oral dose of fenitrothion” 1/30 LD50 ( 20 mg/kg body weight) by gastric tube for 4 week. Group III: The animals received daily oral dose of Thyme extract (500mg/kg body weight/day) by gastric tube for 4 weeks dnd group IV) animals received both fenitrothion and Thyme extract for 4 weeks. Liver specimens were prepared for histological examination (H&E), histochemical examination of glycogen and protein and immuohistochemical detection of caspase-3. Finally quantitative analysis of DNA fragmentation was done. Microscopically group II showed many histological changes in liver sections in the form of loss of the hepatic architecture and apoptosis of hepatocytes with pre-cancerous changes that was supported by highly expressed caspase-3 reaction in the cytoplasm and nuclei of hepatocytes. Marked decrease PAS positive reaction and protein were detected in most hepatocytes. DNA fragmentation were significantly increased (32%) in comparison with all groups. Group IV showed improvement of histological and histochemical changes in the tissue. Thyme extract possess a hepato-protective against fenitrothion as it had antioxidant, anti-inflammatory and anti-genotoxic activities.
Key words: Fenitrothion, Thyme, Liver toxicity
8. Journal of Cell and Tissue Research 15(1) 4747-4750 (2015)
Uterine histomorphological changes in canine pyometra
Jena, B., Sadasiva Rao, K., Das, D. and Reddy, K.C.S.
Department of Animal Reproduction, Gynaecology and Obstetrics, College of Veterinary Science,
Sri Venkateswara Veterinary University, Rajendranagar, Hyderabad 500 030, Andhra Pradesh.
E. mail: drbasantijena@gmail.com,
Abstract: Pyometra is a common reproductive disorder which affects nearly one fourth of all female dogs. The study aimed at observing the morphological and histopathological changes occurring in uterus in canine pyometra. The bitches affected with pyometra (n = 18) were subjected to ovario-hysterectomy. The ovarian and uterine samples were collected for further studies. Grossly, the ovaries consistently contained multiple corpora lutea and occasionally follicular cysts. Uterus was dilated and distended with purulent to sanguinopurulent materials. Histopathological studies showed endometrium was irregularly thickened, ulcerated and haemorrhagic. The endometrium showed inflammatory changes, vacuolar degeneration and sqamous metaplasia. There was massive neutrophilic and mononuclear cell infiltration. The glandular elements showed hyperplasia, fibrosis and cystic distension.
Key words: Pyometra, Canine Uterus
9.Journal of Cell and Tissue Research 15(1) 4751-4754 (2015)
Histomorphological studies of sebaceous glands in relation to age in different regions of Bakerwali goats
Razvi, R., Suri, S., Tagger, R. K., Sarma, K. and Sharma, R.
Division of Veterinary Anatomy Faculty of Veterinary Science and Animal Hunsbandry, SKUAST-J, R.S. Pura, Jammu 181102, J & K. E. mail: doctorrahika356@gmail.com,
Abstract: For the histomorphological studies, the skin samples from eighteen apparently healthy Bakerwali goats of three different age groups viz. below 1 year (neonatal), between 2-3 years (young) and above 3 years (adult) were collected. The sample collection was done irrespective of the sex of animal. The skin samples of 1mm x 1mm thickness were collected from eight different body regions viz. neck dorsal, neck lateral, neck ventral, thorax dorsal, thorax lateral, thorax ventral, loin dorsal and loin lateral. The samples were then processed as per standard histological procedures to study their histomorphology. The sebaceous glands observed were simple, branched and tubular and the periphery of the secretory unit contained a single layer of low cubiodal cells. The ducts of sebaceous glands were lined by stratified cuboidal epithelium, which emptied into associated hair follicle or into associated ducts. The number and depth of sebaceous glands increased from neonatal to adult age groups of Bakerwali goat. The density of sebaceous glands per field was maximum in thorax dorsal regions and minimum in loin lateral region while the depth of these glands were maximum in neck dorsal and minimum in thorax ventral in all age groups. The sebaceous glands showed intense PAS positive reaction and strong intensity for bound lipids..
Key words: Skin histomorphology, Sebaceous glands, Bakerwal goat.
10. Journal of Cell and Tissue Research 15(1) 4755-4759 (2015)
Relative transcript abundance of telomerase reverse transcriptase (TERT) in cell lines
Palai, T.K., Saini, M. and Gupta, P.K.
Division of Animal Biochemistry, Indian Veterinary Research Institute, Izatnagar, Bareilly - 243122 (UP)
E. mail: tapan.palai@gmail.com,
Abstract: In the present study, telomerase reverse transcriptase (TERT) expression was checked for the first time in cell lines like HEK-293T, BHK-21 and Vero following Real-time PCR using primers specific for human TERT amplification. The expression of TERT transcript was detected in all the studied cell lines. Among cell lines studied, expression was found to be much higher in HEK-293T cells as compared to Vero cells and negligible in BHK-21. The present finding will help further studies on exploring the mechanism of immortality in various cell lines i.e whether the phenomenon is due to TERT expression or due to alternate mechanism.
Key words: Telomerase, HEK-293T, BHK-21
11. Journal of Cell and Tissue Research 15(1) 4761-4764 (2015)
Avian cytokines and disease prevention: An overview
Gupta, A.K.,Lather, A. and Kumar, T.
Department of Veterinary Microbiology, Lala Lajpat Rai University of Veterinary and Animal Sciences, Hisar 125004, Haryana. E. mail: drakki1984@gmail.com,
Abstract: Cytokines are soluble low molecular weight (< 30 kDa) regulatory proteins that serve many functions in the body such as mediating and regulating immunity, inflammation and hematopoiesis. Cytokines are secreted by various cell types that play important role in the regulation of immune response, but the largest group of cytokines is involved in immune cell proliferation and differentiation. Cytokines have enormous potential to control infectious diseases in poultry. Recent advances in recombinant DNA technology and avian genetics have explored the application of avian cytokines in poultry medicine. The use of cytokines in poultry has become more feasible with the discovery of number of avian cytokine genes. This monograph will review the recent advances made in avian cytokines and their therapeutic potential.
Key words: Cytokine, Avian disease
12. Journal of Cell and Tissue Research 15(1) 4765-4770 (2015)
Protective effects of Tinospora cordifolia against methotrexate induced oxidative damage in rat liver
Deepak, J.N., Rao, S., Byregowda, S.M., Vetrivel, M., Purushotham, K.M., Satyanarayana, M.L., Narayanaswamy, H.D. and Renukaprasad, C.
Department of Veterinary Pathology, Veterinary College, Hebbal, Bangalore 560 024.
E. mail: drdeepakjn@gmail.com
Abstract: The role of aqueous extract of Tinospora cordifolia [TC], in preventing methotrexate [MTX] induced oxidative damage in rat liver was studied. Sixty Wistar albino rats were divided into five groups with twelve rats in each group. Group I served as normal control group. To group II rats, MTX was administered at 5mg/kg body weight intraperitoneally for three consecutive days. Rats in group III were administered with MTX and treated with TC extract at 200mg/kg by oral gavaging for 45 days. Group IV rats were pre-treated with TC ten days prior to MTX administration and followed by TC treatment for 45 days. Rats in group V served as TC control and were administered TC at the dose of 200mg/kg body weight for 45 days. The liver samples collected at 7th, 14th, 28th, and 45th day of the study were subjected for estimation of catalase, superoxide dismutase, glutathione peroxidase and malondialdehyde [MDA]. Significant decrease [P<0.01] in the levels of antioxidant enzymes and increase in MDA levels were recorded in MTX treatment group. The TC treatment groups revealed a significant recovery [P<0.01] all enzymes and decrease in MDA levels which suggested that TC has a good antioxidant effect and could be effectively used for prevention of toxic side effects of MTX. The study also indicated that pre-treatment of TC prior to MTX administration has good prophylactic effect.
Key words: Tinospora cordifolia, Methotrexate, Oxidative damage
13. Journal of Cell and Tissue Research 15(1) 4771- 4774 (2015)
Detection of residual concentration of monensin in chicken meat
Singh, R.P., Sahni, Y.P., Shrman, K.and Chandra, N.
Department of Veterinary Pharmacology & Toxicology, NDVSU, Jabalpur, M.P.
E. mail: drncvet@gmail.com,
Abstract: In the present study, surveillance was conducted regarding use of anticoccidials at poultry farms located in and around district Jabalpur. The data on use of various anticoccidials were further shortlisted and found that monensin is one of the most commonly used anticoccidials and therefore aimed to detect monensin residues in chicken meat .Total number of 180 chicken meat samples were analyzed including 36 samples each from five area of Jabalpur. Out of the 180 samples, 13 samples were found positive for residual concentration of monensin in chicken meat and all were detected below maximum residue limits recommended by Commission Regulation of European Union for monensin in chicken muscle, liver and kidney samples. The study was further extended to determine the effect of boiling on residual concentration of monensin in chicken meat samples positive for drug residue. Mean residual concentration of positive samples was detected on pre and post boiling.
Key words: Monensin, Chicken meat
14. Journal of Cell and Tissue Research 15(1) 4775- 4782 (2015)
Angiogenic and antiangiogenic influence of streptozotocin and insulin on chorioallantoic membrane of chick embryo
Surve, Z.R., Patil, S.R., Jha, S.M., Shetty, G.D. and Manekar, A.P.
Molecular Endocrinology Laboratory, Department of Zoology, The Institute of Science, 15- Madam
Cama Road, Mumbai 400 032. E. mail: ashok.manekar@rediffmail.com,
Abstract: Angiogenesis, the development of new blood vessel from pre-existing vessels, is a key process in the formation of the granulation tissue during wound healing. The appropriate development of new blood vessels, along with their subsequent maturation and differentiation establishes the foundation of functional neo-vasculature. Angiogenesis has been evaluated by in vivo and in vitro systems using different types of endothelial cells isolated either from capillaries or large vessels. Therefore, in present study, the induction of angiogenesis by insulin and streptozotocin has been evaluated in chorioallantoic membrane (CAM) of the avian embryo. Streptozotocin and insulin were administered to two respective groups of 8 days old chick embryo by making window on the chorioallantoic membrane. Parallel control was maintained, where embryos received phosphate buffered saline (PBS) of pH 7.4. On 12th day of incubation window was opened to reveal the progressive blood vessels pattern formation in CAM in insulin treated embryos with the help of stereomicroscopic study. In controls, no such changes were observed. Biochemical study by spectrometric analysis and histological study was also carried out to understand the difference between insulin treated, streptozotocin treated and control embryos.
Key words: Chorioallantoic membrane, Streptozotocin, Insulin, Angiogenesis
15. Journal of Cell and Tissue Research 15(1) 4783- 4790 (2015)
Assessment of genetic diversity in Lepidium sativum l. (Chandrasur) genotypes using morphological and molecular markers
Sharma, J., Joshi, A., Dodiya, N.S., Jain, P., Rajoriya, S.K., Rajamani, G., and Jain, D.
Department of Molecular Biology and Biotechnology, Rajasthan College of Agriculture, Maharana Pratap University of Agriculture and Technology, Udaipur 313001.
E. mail: devendrajain@mpuat.ac.in,
Abstract: Lepidium sativum L. (Brassicaceae), also known as Asaliya and Chandrasoor in local languages, is an important medicinal crop in India, is a fast growing annual herb. Despite of its diverse medicinal uses, the genotype characterization and molecular diversity studies till date in Lepidium sativum are very few. Fifteen Chandrasur genotypes isolated from farmer’s field of Udaipur, India were evaluated for genetic diversity using morphological and RAPD markers. Eight morphological traits were studied and subjected to analysis of variance. RAPD analysis was carried out using 20 random primers out of which 10 primers produced reproducible 61 bands, of which 54 were polymorphic. Based on the RAPD marker analysis, a dendrogram was constructed using UPGMA method. The similarity coefficient ranged from 0.31 to 0.92 with an average of 0.68. Cluster analysis based on RAPD data separated the genotypes into four clusters. The results showed that the level of genetic variation was high among the Chandrasur genotypes. The results of the present study can be used for Lepidium sativum crop improvement through molecular breeding and Marker assisted selection of for desired traits in future.
Key words: Lepidium sativum, Morphological and molecular markers
16. Journal of Cell and Tissue Research 15(1) 4791- 4794 (2015)
Clinicopathological attributes of swine pasteurellosis
Tigga, M., Ghosh, R.C., Choudhary, B. K. and Tigga, P.
Department of Veterinary Pathology, College of Veterinary Science and Animal Husbandry,
Chhattisgarh Kamdhenu University, Anjora, Durg - 491 001, Chattisgarh; E. mail: chiyamum@gmail.com,
Abstract: Swine pasteurellosis was observed in descript as well as non-descript pigs of Chhattisgarh, India. Affected animals exhibited pyrexia, dullness, staggering gait, anorexia, serous nasal discharge and dyspnoea. Case fatality rate was 95% in adult animals and 100% in piglets. Typical lesions of oedematous swellings noted in the pharyngeal region, these swellings spread to the ventral cervical region and brisket of affected pigs. Gross lesions included severe pneumonia and haemorrhages in lungs, petechial haemorrhages on serous membranes and other visceral organs. Lymph nodes were enlarged, oedematous and haemorrhagic. The blood smears from heart blood and tissue impression smears revealed bipolar organisms indicating the presence of Pasteurella spp. The cultures obtained were subjected to Gram’s staining for checking the purity, bipolar morphology and biochemical characterization of the organisms. Molecular characterization revealed Pasteurella multocida, capsular type B. On histopathological examination, lungs showed typical fibrinous bronchopneumonia, multifocal suppuration. Pleura was severely thickened. Heart of some pigs showed presence of thrombi, haemorrhages and necrosis of myocardium.
Key words: Swine Pasteurellosis, Pasteurella multocida, Haemorrhagic septicaemia
17. Journal of Cell and Tissue Research 15(1) 4795- 4800 (2015)
Effect of addition of bovine serum albumin on the liquid storage (5°C) of mithun (Bos frontalis) semen
Perumal, P., Nahak, A.K., Vupru, K., Khate, K., Balamurugan, T.C. and Prakash Krupakaran, R.
ICAR- National Research Centre on Mithun, Jharnapani 797 106, Nagaland.
E. mail: Perumalponraj@gmail.com,
Abstract: The present study was undertaken to assess the effect of bovine serum albumin (BSA) on sperm motility, viability, total sperm abnormality, acrosomal and plasma membrane integrity. A total of 30 ejaculates were collected from mithun bulls and semen was split into five equal aliquots, diluted with the TEYC extender. Group 1: semen without additives (control), group 2 to group 4: semen was diluted with 5 mg/ml, 10 mg/ml and 20 mg/ml of BSA, respectively. These seminal parameters were assessed at 5oC for 0, 6, 12, 24 and 30 h of incubation. Inclusion of BSA into diluent resulted in significant (P <0.05) decrease in percentages of dead spermatozoa, abnormal spermatozoa and acrosomal abnormalities at different hours of storage periods as compared with control group. Additionally, BSA at 5 mg/ml has significant (p< 0.05) improvement in quality of semen than BSA at 10 mg/ml and 20 mg/ml stored in in-vitro for up to 30 h. It was concluded that the possible protective effects of BSA on sperm parameters may be due to the prevention of lipid peroxidation.
Key words: Mithun bull (Bos frontalis), Bovine serum albumin.
18. Journal of Cell and Tissue Research 15(1) 4801- 4804 (2015)
Treatment of cholestrerol loaded cyclodextrin improves goat sperm cryosurvival
Behera, S., Harshan, H. M., Aravinda Ghosh, K. N., Metilda, J., Mini, M. and Lekshmi Bhai, K.
Department of Animal Reproduction Gynaecology and Obstetrics, College of Veterinary and
Animal Sciences, Mannuthy, Trissur 680 651, Kerala Veterinary and Animal Sciences
University. E. mail: sunita5656@gmail.com,
Abstract: The study was carried out using 48 semen sample collected from adult Malabari bucks maintained at the AI centre in the Department of Animal Reproduction Gynaecology and Obstetrics. The collected and pooled semen ejaculates were divided into three groups. Group I was kept as control, group II and III were treated with 1 mg and 2 mg of Cholesterol-loaded-cyclodextrin (CLC)/ 120 × 106 spermatozoa respectively in an extender containing Tris- citric acid-glucose buffer without egg yolk and were incubated at 37°C for 15 min followed by manual freezing. Treatment of semen samples with CLC resulted in improved sperm motility, viability, acrosome integrity at pre-freeze and post-thaw stages of cryopreservation. Treatment of semen with 1 mg of CLC/120 × 106 spermatozoa was observed to be better than treatment with 2 mg of CLC/120 × 106 spermatozoa and the control.
Key words: Malabari bucks, Cryopreservation
19. Journal of Cell and Tissue Research 15(1) 4805- 4810 (2015)
Effect of methyl -β- cyclodextrin and cholesterol- loaded cyclodextrinon malabari buck spermatozoa thermoresistance
Behera, S., Harshan, H.M., Lekshmi Bhai, K., Ghosh, K.N.A., Joseph, M.and Mini, M.
Department of Animal Reproduction, Gynaecology and Obstetrics, College of Veterinary and Animal Sciences, Mannuthy, Thrissur, Kerala – 680651, Kerala Veterinary and Animal Sciences University.
E. mail: sunita5656@gmail.com,
Abstract: The study was carried out using 48 semen sample collected from two adult Malabari bucks maintained at the AI centre in the Department of Animal Reproduction Gynaecology and Obstetrics. The collected and pooled semen ejaculates were divided into three groups. Group I served as control, group II was treated with 1 mg of cholesterol-loaded-cyclodextrin (CLC)/ 120 × 106 spermatozoa and group III with 1 mg of methyl-β-cyclodextrinin an extender containing Tris- citric acid-glucose buffer without egg yolk and were incubated at 37°C for 15 min followed by manual freezing. Treatment of semen samples with CLC resulted in improved sperm motility, viability, functional membrane integrity at post-thaw stages of cryopreservation and up to two hours of incubation periods. Methyl-β-cyclodextrin treated spermatozoa resulted in significantly lower seminal parameter values as compared to other groups.
Key words: Methyl-β-cyclodextrin , Malabari buck, Spermatozoa
20. Journal of Cell and Tissue Research 15(1) 4811- 4814 (2015)
Evaluation of Emblica officinalis and Mentha piperata supplementation on biochemical parameters in
growing beetal kids
Madan, J., Sindhu, S., Gupta, M.andPoonia, J.S
Department of Veterinary Physiology and Biochemistry, Lala Lajpat Rai University of Veterinary and Animal Sciences, Hisar 125 004, Haryana. E. mail: jyoti_mad@yahoo.com,
Abstract: A ninety days trial was conducted on 36 growing beetal kids (3-4 months age) to evaluate the effect of Emblica officinalis and Mentha piperata supplementation on biochemical parameters at Goat farm, Animal Genetics & Breeding, LUVAS, Hisar. The experiment was conducted from June to August. The maximum environmental temperature ranged from 24.6°C to 45.0°C. The overall mean Thermal humidity index values were 81.21 in the morning and 88.80 in the evening. All the experimental animals were under mild heat stress. The kids were randomly divided into three groups each comprising of six male & six female. The groups were assigned to treatment as T 1 –concentrate feed, T 2 -98% concentrate feed + 2% Emblica powder, T 3 -98% concentrate feed + 2% Mentha powder. Blood samples were collected at 0, 30, 60 and 90 days and plasma was separated for analysis of various parameters. A significant decrease was observed in blood glucose, total cholesterol, triglyceride, lactate dehydrogenase, alanine amino transferase and aspartate amino transferase by feeding Emblica officinalis in kids. Supplementation of Mentha has shown no alteration in total protein, triglyceride, urea and enzymatic activity (Alanine amino transferase and aspartate amino transferase), however, a significant decrease was observed in glucose, cholesterol and lactate dehydrogenase. The results revealed that feeding of Emblica officinalis and Mentha piperata has hypoglycemic, hypolipidemic effects in goat kids. It may be concluded that supplementation of Emblica and Mentha have shown beneficial effects, however the effect was more significant in kids fed Emblica officinalis. As these supplements are easily available and they can be used as feed supplements in growing kids to achieve their maximum potential.
Key words: Emblica officinalis, Mentha piperata, Goat kids
21. Journal of Cell and Tissue Research 15(1) 4815- 4818 (2015)
Studies on TLR4 gene polymorphism with eference to production of serum lysozyme and IgG in goats
Kumar, S.B., Singh, S.R., Verma, S.B.,Mandal, K.G. and Ranjan, R
.
Department of Animal Breeding and Genetics, Bihar Veterinary College, Patna, 800 014
E. mail: rkrbiotech@gmail.com,
Abstract: Toll-like receptors (TLRs) are single transmembrane cell-surface receptors, which play a significant role in the innate immune system. Lysozyme is abundant in the body fluid and it is considered as a non-specific bactericidal substance. Serum immunoglobulin levels provide key information on the humoral immune status of the animals. The aim of the present investigation is to find out the polymorphism of TLR4 gene and the level of serum lysozyme and IgG in the three breeds of goats. PCR-RFLP of 494 bp amplicon of TLR4 gene shown AA and AB genotypes in Beetal goats, whereas Jamunapari and Black Bengal goats were found monomorphic and having only AA genotype. The allelic frequencies of A and B alleles in Beetal were 0.67 and 0.33 respectively, while the frequency of A allele in Jamunapari and Black Bengal was 1.0. Serum concentration of IgG in Beetal was significantly (P<0.01) higher by 5.52 mg/ml and 4.22 mg/ml than the Jamunapari and Black Bengal goats respectively. Beetal had higher concentration of serum lysozyme than the Janmunapari and Back Bengal. The study shows that Beetal goat breeds having high serum IgG and Lysozyme level may be selected for relatively better disease resistance against bacteria and virus.
Key words: Toll like receptors, Lysozyme, Goat
22. Journal of Cell and Tissue Research 15(1) 4819- 4822 (2015)
Reverse transcription polymerase chain reaction confirmation of catalytic domain of matrix metalloproteinases-9 gene in semen of Murrah buffaloes
Prakash Krupakaran, R., Arunkumar, S., Balamurugan, T. C., Pandiyan, G.D.V. and Perumal, P.
Department of Veterinary Physiology and Biochemistry,Veterinary College and Research Institute Tamilnadu Veterinary and Animal Sciences University (TANUVAS), Orathanadu, Thanjavur 614 625, Tamil Nadu. E. mail: tcbalamurugan@gmail.com,
Abstract: Matrix metalloproteinases (MMPs) form a family of structurally-related, zinc-dependent proteases which are capable of restructuring tissue components by proteolytic degradation of extracellular matrix and basement membrane compounds. MMP-2 and MMP-9 are the two major enzymes belonging to this family. These Proteases in the reproductive tract are considered to play key roles in fertilization processes. A study on biochemical and molecular characterization of seminal plasma proteins in buffalo bulls was carried out. Total RNA was isolated from the semen samples of buffalo using standard procedure. The concentration and purity of RNA was determination by 260 and 280 nm in a spectrophotometer. The ratio of A260/A280 was 1.82 indicating the isolated RNA was reasonably pure. The integrity of RNA was assessed by agarose gel electrophoresis. Primers targeting catalytic domain of the MMP-9 were designed as the forward primer 5'-CGGCGGATCCTGGCA-CCACAACGACATCACTTA-3 and the reverse primer 5'-CGGCGTCGACTCTAGAGGGAGGACCAGTAGCGCAGA-3'. Restriction enzyme (RE) sites were incorporated viz. BamHI, XbaI and SalI along with appro-priate overhanging sequences on forward and reverse primers respectively for directional cloning of the amplified product. After incorporation of the RE sites the expected size of the amplicons was 1080 bp. RT-PCR was carried out. The first step was to synthesize cDNA and the second step to amplify the desired genes from cDNA by PCR. The amplified PCR product was checked by submarine gel electrophoresis using 1.5% agarose. The presence of catalytic domain of MMP-9 was observed at 1080bp, which was confirmed by the presence of MMP-9 gene in semen of Murrah buffaloes.
Key words: Catalytic domain, MMP-9 Gene, RT-PCR, Murrah buffaloes
23. Journal of Cell and Tissue Research 15(1) 4823- 4826 (2015)
Comparative studies on serum biochemical profiling at various stages of reproduction in Murrah buffaloes
Balamurugan, T.C., Prakash Krupakaran, R., Pandiyan, G.D.V., Arunkumar,S.and Perumal, P.
Department of Veterinary Physiology and Biochemistry; Veterinary College and Research Institute, Tamilnadu Veterinary and Animal Sciences University (TANUVAS), Orathanadu, Thanjavur 614 625, Tamil Nadu. E. mail: tcbalamurugan@gmail.com,
Abstract: A comparative study on serum biochemical constituents was carried out in pregnant, estrus, anestrus and regular cyclic Murrah buffaloes. A total of 24 healthy Murrah buffaloes, aged about 3-6 years, were selected and they were divided into four groups, each comprising of six animals (Group-I: pregnant; Group-II: estrus; Group-III: anestrus and Group-IV: regular cyclic). Animal were properly vaccinated and dewormed. In early morning before feeding the animals, blood samples were collected in heparinised vacutainer and immediately transported to the laboratory. The blood samples were centrifuged at 3000 rpm for 15 min. Serum samples were separated and kept at -20ºC until further use. Biochemical assays were carried out for glucose, total protein, albumin, urea, creatinine, total cholesterol, alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), calcium and phosphorous. These parameters were investigated with Span Diagnostic kits as per the standard biochemical procedures. The serum biochemical values of regular cyclic and the estrus animals were almost similar in all the parameters. Among the four groups, the serum biochemical values were higher in pregnant group and the anestrus group had lowest values.
Key words: Murrah buffaloes, Serum biochemistry
24. Journal of Cell and Tissue Research 15(1) 4827- 4831 (2015)
Study of endocrine profile of responded and nonresponded buffaloes before and after
ovsynch treatment
Kumari, K., Mishra, U.K., Kumar, K,. Praveen, P.K., Dalai, N. and Shekhar, S.
Department of Veterinary Physiology and Biochemistry, College of Veterinary Science and Animal Husbandry, Anjora, Durg (C.G). E. mail: drkanchankumari@gmail.com,
Abstract: In the present study the effect on endocrine profile of 10 Murrah buffaloes (7 responded and 3 nonresponded) was carried out before and after Ov synch treatment. The result showed that the mean level of Progesterone in responded animals were found to be 2.13±0.57 ng/ml on 22 day before treatment (-22 days). Minimum level of progesterone was found 1.8±0.46ng/ml on 4th day before start of the treatment (-4 day) and maximum level was 2.83±0.48ng/ml was found at the time of 1st GnRH injection (0 day). In nonresponded animals progesterone level was significantly & continuously increasing from -22 to 0 days. The mean level of estradiol-17 β in responded animals 9.57±2.43 pg/ml on 22 day before treatment (-22days). Minimum level of estradiol-17β was found 6±0 pg/ml on 14th, 12th day before start of the treatment & at the time of 1st GnRH injection (-14, -12, & 0 days) and maximum level was 10.64±2.83pg/ml was found at 8th day before treatment (-8day). The mean level of estradiol-17 β in non responded animals was found to be 20.33±1.46 pg/ml on 22 day before treatment (-22days). Insulin level observed during the present study in both responded and non responded animals were well within the normal range.
Key words: Progesterone, Estradiol-17β, Insulin, Ovsynch treatment
25. Journal of Cell and Tissue Research 15(1) 4833- 4837 (2015)
Changes in serum biochemical parameters before and after ovsynch treatment in Murrah buffaloes
Kumari, K., Mishra, U.K., Kumar, K., Praveen, P.K., Dalai, N. and Shekhar, S.
Department of Veterinary Physiology and Biochemistry, College of Veterinary Science and Animal Husbandry, Anjora, Durg (C.G). E. mail: drkanchankumari@gmail.com,
Abstract: In the present study the effect on endocrine profile of 10 Murrah buffaloes (7 responded and 3 nonresponded) was carried out before and after Ov synch treatment. The result showed a range of 9.5±0.2mg/dl to 11.48±1.05mg/dl serum calcium level in responded group and that of 7.7±1.78mg/dl to 11.26±0.38mg/dl serum calcium level in non responded group was observed. A non significant difference has been observed between the value of phosphorous level before and after treatment in both the group. It was between 6.6±0.35mg/100ml to 9.6±1.03mg/100ml in both the groups. Total serum protein, albumin and A/G ratio were well within the normal rage. The blood serum level of total cholesterol was minimum on -14 day (69.87±4.08 mg/dl) and it was maximum on +22 day (110.8±7.38 mg/dl) during the study in responded animals. But in non responded animals, blood serum level of total cholesterol was lowest on +12 day (60±12.61mg/dl) and it was highest on +18 day (99.9±0.25mg/dl). Difference was non significant before and after treatment.
Key words: Serum biochemistry, Ov synch treatment, Murrah buffaloes
26. Journal of Cell and Tissue Research 15(1) 4839- 4842 (2015)
Biochemical profile of buffalo under rural managemental condition of india
Maurya, S.K., Kumar, B., Kumawat, B.L., Saxena, A., Sagar, R., Jan, M.H., Jai Singh, Vandana, Das, G. K. and Perumal, P.
Department of ARGO, U.P. Pt. Deen Dayal Upadhyaya Veterinary and Animal Sciences University Mathura 281001, Uttar Pradesh. E. mail: drbrijeshvet02@gmail.com,
Abstract: A study was undertaken to assess the effect of mineral supplementation on conception rate, endocrinological profile and biochemical composition of estrus buffaloes. Twenty parous estrus buffaloes from different villages in and around DUVASU, Mathura were selected and divided randomly into two group viz. group I: animals were fed with commercially mineral mixture and group II: control (without mineral supplementation). Mineral mixture supplementation was continued for twenty five days and blood sample were collected on day 0 and day10 of subsequent estrus and preserved at -20°C in deep freezer until use. The result revealed that there was an increase in conception rate in treated than in control group. The progesterone concentration has a significantly (P<0.05) higher, insulin has non- significantly higher and estradiol has non- significantly lower value on day 10 in the treatment than in control group. Mineral elements such as calcium, magnesium, copper and zinc were non- significantly differed between the experimental groups day 0 whereas on day 10, there was a significant (P<0.01) difference between groups. But phosphorus and cobalt were shown non- significantly higher concentration in treated group than in control group in both day 0 and 10. Biochemical profile such as total protein and glucose were differed significantly between the treated and control group on day 10 and for glucose on day 0 whereas other biochemical profiles were non- significantly higher in the treatment group than in control group at both day 0 and 10 of estrus. These results indicated that mineral supplementation has improvement in the conception rate and significantly increased in the concentration of some important macro and micro elements.
Key words: Buffalo, Mineral supplementation
27. Journal of Cell and Tissue Research 15(1) 4843- 4848 (2015)
Improved protoplast culture conditions in barley (Hordeum vulgare l.)
Tiwari, V.K.and Cocking, E.C.
Plant Genetic Manipulation Group, Department of Life Science, University of Nottingham, University Park, Nottingham, NG 7 2 RD, England. E. mail: vkt786@rediffmail.com,
Abstract: The plasmolyses of cells before enzymatic incubation of cell aggregates significantly produced massive totipotent protoplasts from immature embryo-derived cell suspensions of cv. Dissa. Heat shock treatment (45 °C for 5 min followed by 10 s cooling on ice) of protoplasts before culture were significantly increased plating efficiency and viability of protoplasts than without-heat-shock treated protoplasts. Maximum numbers of protoplasts were obtained 4 days after subculture for all cell lines. Highest protoplast plating efficiency was observed for the 8, 12, 16 months old cell lines. Maximum plating efficiency were observed when protoplasts were cultured in KMP8 medium at the density of 5x105 protoplasts/ml in all cell lines and no division occurred at the density of 2x105 protoplasts/ml. Cell divisions in protoplasts-derived cells were enhanced by the addition of low osmoticum (Osmolarity = 440 ± 20 mOsm/kg) KM8 medium after 21 days of culture. Protoplast division frequency and colony formation varied significantly for all the lines using agarose-layer method and droplet method, except for line LDR1.2
Key words: Protoplast culture, Barley
28. Journal of Cell and Tissue Research 15(1) 4849- 4854 (2015)
Refinement of in-vitro regeneration system in Elite safflower (Carthamus tinctorius l.) genotypes
Dhumale Dipti, R., Dudhare, M.S., Mohite, N.R., Shingote, P.R., Jadhav, P.V. and Moharil, M.P.
Biotechnology centre, Department of Agricultural Botany, Dr. Panjabrao Deshmukh Agricultural University, Akola- 444104, Maharashtra, India.E. mail: diptidhumale@gmail.com,
Abstract: In-vitro regeneration system for two safflower genotypes, AKS-207 and PKV-Pink using cotyledonary leaf explant was established. Highest percentage of callusing response was observed on MS medium with equal concentration of 2,4-D and KIN (1 mg/l each) in both the genotypes. Shoot induction as well as multiple shoot formation recorded maximally on medium supplemented with 3 mg/l BAP in AKS-207 and 4 mg/l BAP in PKV-Pink. Highest percentage of shoot elongation was achieved by using GA3 at 1 mg/l in both the genotypes. Rooting was achieved on half strength MS medium fortified with NAA (2 mg/l) in both the genotypes. In-vitro rooted plantlets were transferred to pre-hardening conditions but failed to respond ex-vitro. To overcome the major constrain of verification in the present regeneration system, repeated sub-culturing of explants on fresh shooting medium was practiced. Overall regeneration system indicated that, safflower genotype AKS-207 is more responsive than PKV-Pink for in-vitro study.
Key words: Carthamus tinctorius L., In-vitro regeneration
29. Journal of Cell and Tissue Research 15(1) 4855- 4860 (2015)
Improved somatic embryogenesis in indian bread wheat (Triticum aestivm l.) cultivar HD 2967 through media manipulations
Gill, A.K. and Gosal, S.S.
School of Agricultural Biotechnology, Punjab Agricultural University, Ludhiana-141004, Punjab
E. mail: amangill1189@gmail.com,
Abstract: Different culture medium formulations were used to achieve higher somatic embryogenesis rates in bread wheat cultivar HD 2967 and were compared with embryogenesis rates in Bobwhite. Immature embryo derived primary calli were cultured on different media but with constant 2,4-D concentration (2.0 mgL-1). Effects of kind and concentration of carbohydrate source were studied at 3%, 6% and 9% levels of sucrose and maltose. Maltose (3%) was found to be superior over all the other concentrations of sucrose and maltose. Profound effect of increased agar concentration was observed on embryogenesis in both the varieties. Among the different agar concentrations studied (0.8%, 1.2% and 1.6%), agar (1.2%) showed maximum embryogenesis. Effect of phytagel (0.2% and 0.5%) was also studied but the per cent calli showing embryogenesis were found to be more on medium solidified with agar (1.2%). Use of proline and cefotaxime at a certain concentration in the culture medium showed positive effect on embryogenesis but no effect of proline was observed in increasing embryogenesis in Bobwhite.
Key words: Triticum aestivum L., Somatic embryogenesis
30. Journal of Cell and Tissue Research 15(1) 4861- 4866 (2015)
Effect of heat stress on enzymes and isoenzyms of wheat genotypes at tillering and grain
filling stages
Ramani, H.R. and Mandavia, M.K.
Department of Biochemistry, Junagadh Agricultural University, Junagadh 362 001, Gujarat.
E. mail: hiteshramani2007@gmail.com,
Abstract: Heat tolerant (GW-190) and heat susceptible (J-2010-11) wheat genotypes were selected for enzymatic studies. One set was grown up to Tillering stage (35 days) and divided into two groups, control and heat treatments. Other set was grown up to grain filling stage (around 100 days) and also divided into two sets, control and heat treatments. Heat treatments (40°C and 45°C for 2h and 4h) were given using heating house at both the stages. Results of enzyme activities of peroxidase, superoxide dismutase and glutathione reductase and isoenzymic pattern of peroxidase and superoxide dismutase show difference between heat tolerant and heat susceptible genotype as well as difference between development stages with temperature.
Key word: Wheat genotypes, Heat stress enzymes
31. Journal of Cell and Tissue Research 15(1) 4867- 4875 (2015)
Genetic diversity amongs Withania somenifera (L.) dunal genotypes using morphological and molecular markers
Sahu, V., Dodiya, N.S.,Joshi, A., Rajoriya, S.K., Jain, P. and Jain, D.
Department of Plant Breeding and Genetics, Rajasthan College of Agriculture, Maharana Pratap University of Agriculture and Technology, Udaipur 313 001 (Rajasthan). E. mail: devendrajain@mpuat.ac.in,
Abstract: Withania somnifera (L.) Dunal (solanaceae), also known as Asandh, Asagandh and Ashwagandha in local languages is an important medicinal crop in India. In Ayurveda the plant of W. somnifera is considered to cure swelling, poultices lesions, tubercular ulcers, scabies, ophthalmia, muscular pain, dropsy, rheumatism, diabetes and even cancer. Despite of its diverse medicinal uses, the genotype characterization and molecular diversity studies till date in W. somnifera are very few. Twenty Ashwagandha genotypes were evaluated for genetic diversity using morphological and RAPD markers. Nine morphological traits were studied and subjected to analysis of variance, estimation of different variability parameters, correlation and path coefficient analysis. RAPD analysis was carried out and 9 random primers produced 46 bands, of which 41 were polymorphic and reproducible with 89.13% polymorphism. Based on the RAPD analysis, dendrogram was constructed using UPGMA method and the similarity coefficient ranged from 0.37 to 0.86 with an average of 0.65. The results showed that the level of genetic variation was high among the Ashwagandha genotypes. The groups/clusters obtained by molecular marker could also be distinguished by dissimilarity/similarity for the morphological characteristics within each group. The results of the present study can be used for W. somnifera crop improvement through molecular breeding and Marker assisted selection of for desired traits in future.
Key words: Withania somnifera, Molecular marker, Genetic diversity
32. Journal of Cell and Tissue Research 15(1) 4877- 4882 (2015)
In vitro mass multiplication of noni (Morinda citrifolia L.) through nodal segment explants
Saini, M.K. and Patel, R.M.
Department of Plant Molecular Biology and Biotechnology, ASPEE College of Horticulture
and Forestry, Navsari Agricultural University, Navsari 396450, Gujarat.
E. mail: mksainihorti@yahoo.com,
Abstract: Noni (Morinda citrifolia L.) fruits are widely used for blood pressure, respiratory problems, immune deficiencies, antibacterial, anti-inflammatory, analgesic and anti-congestive therapies. To produce large scale true to type plant a tissue culture protocol was developed. Maximum establishment of nodal segment explants observed in Murashige and Skoog (MS) medium containing 2.0 mg/l 6- benzylaminopurine (BAP). However, MS medium fortified with 2.0 mg/l BAP + 1.0 mg/l kinetin exhibited maximum multiplication rate in second and third sub-cultures. The maximum frequency of multiple shoots in nodal segment explants (84.33 %) was observed on treatment MS + 2.0 mg/l BAP + 1.0 mg/l kinetin. In vitro rooting of regenerated shoot developed in half strength MS medium supplemented with 1.0 mg/l IBA, which produced maximum number of roots per shoot (10) and length of root (3.10). In vitro grown plantlets having 6.23 cm length of shoot were transferred to coco peat media under green house, which showed better survival of plantlets (95 %).
Key words: Morinda citrifolia, Tissue culture
33. Journal of Cell and Tissue Research 15(1) 4883- 4889 (2015)
Molecular and biochemical characterization of soybean genotypes for the presence of kunitz
trypsin inhibitor
Kaur, H., Dhaliwal, H. S. and Chugh, V.
Department of Biotechnology, Akal College of Agriculture, Eternal University, Baru Sahib 173101,
Himachal Pradesh. E. mail: vishalchugh3@gmail.com,
Abstract: Present study was conducted to characterize soybean genotypes for the kunitz trypsin inhibitor at molecular and biochemical level. Genotypes used in the present study included one null KTI soybean line PI542044 (the source of null allele of kunitz trypsin inhibitor being used in soybean breeding programs in India), DSRtiti (homozygous recessive genotype for KTi gene) and seven other elite soybean cultivars. Genomic DNA of all the soybean lines was amplified using gene specific primer for null locus of KTi3 gene and a SSR marker, Satt228 which is tightly linked to Ti locus. Both the markers evidently distinguished between the kunitz trypsin inhibitor free genotypes (PI542044, and DSRtiti) and cultivars with kunitz trypsin inhibitor (Bragg, VL Bhatt, SL-688, SL-525, PS-1042, SL-744 and Dadacha respectively). The results presented show the utility of gene specific as well as SSR maker for identification of kunitz trypsin inhibitor free genotypes developed from PI542044. Kunitz trypsin inhibitor protein appeared as 21.5 kDa protein band as revealed by SDS-PAGE. A perfect correspondence was observed at DNA and protein level in terms of kunitz trypsin inhibitor which was further validated by assaying the trypsin inhibitor activity in which kunitz trypsin inhibitor free genotypes showed very low trypsin inhibitor activity.Therefore, the results clearly indicate that the either the gene specific marker used in the present study or linked SSR marker Satt228 can be efficiently used in the selection of target plants carrying null allele of kunitz trypsin inhibitor in the marker assisted breeding for development of kunitz trypsin inhibitor free soybean in our country.
Key words: Gene Specific marker, Kunitz trypsin inhibitor
34. Journal of Cell and Tissue Research 15(1) 4891- 4896 (2015)
Thidiazuron induced shoot organogenesis in wild guar (Cyamopsis serrata Shinz.)
Singh, V., Yadav Neelam, R., Arora, A., Batra, P. and Yadav, R.C.
Department of Molecular Biology and Biotechnology, CCS Haryana Agricultural University,
Hisar 125004. E. mail: virenderssheoran@gmail.com,
Abstract: Cyamopsis serrata shinz. is a drought tolerant and early maturing wild species of guar. In the present study the effect of various growth regulators and their combinations on hypocotyl and cotyledonary node explants was studied. Direct shoot regeneration was observed in cotyledonary node explants on Murashinge and Skoog basal medium containing B5 vitamins and BAP, TDZ, IAA alone or in combination. In cotyledonary node explants highest shoot regeneration response of 89.60% was observed on MSB medium supplemented with BAP (1.5 mg/l) giving an average number of 1.5±0.2 shoots per explants but highest numbers of shoots per explant (3.6±0.3) with shoot induction response of 87.26% was observed on MSB medium supplemented with thiadizuron (1 mg/l). In hypocotyl explants, direct shoot regeneration was observed on MSB medium fortified with BAP (0.5-2 mg/l). Maximum shoot regeneration response (38.06%) was observed on medium having BAP (2 mg/l) with 4.7±0.3 numbers of shoots per explants however maximum number of shoots per explants (6.3±0.4) was observed on medium supplemented with BAP 1.5 mg/l with 30.90% frequency of shoot regeneration. Indirect organogenesis in hypocotyl explants was observed on medium containing TDZ (0.5- 2 mg/l). In addition, maximum percentage shoot regeneration (89.06%) and maximum number of shoots per explants (12.5±1.1) was observed on MSB medium supplemented with TDZ (0.5 mg/l). This study provides invaluable information regarding regeneration of recalcitrant legume plant species.
Key words: Thidiazuron, Cyamopsis serrata shinz., Shoot regeneration
35. Journal of Cell and Tissue Research 15(1) 4897- 4902 (2015)
Efficiency of method and media composition on transformation of andrographis paniculata for hairy root production
Mahobia, A. and Jha,Z.
Department of Plant Molecular Biology and Biotechnology, IGKV, Raipur, 492012
E. mail: zenujha@yahoo.co.in,
Abstract: Agrobacterium rhizogenes mediated transformation produces hairy roots in the plant parts, which is used for the production of secondary metabolites because of its stability and high productivity in hormone-free culture conditions, to ensure economic benefits. Andrographis paniculata produces pharmaceutically important chemical andrographolide which is a diterpene lactone and has been reported for multiple clinical applications in ayurveda, sidha and unani system of medicine. For establishment of hairy roots for andrographolide production from A. paniculata, two types of explant (leaf and apical meristem) were selected and co-cultured with A. rhizogenes (MTCC 532 strain) by two different methods (dip and drop) and inoculated on medium supplemented with different concentrations of acetosyringone (300 µM and 400 µM) with/without 1 mg/L NAA. Among the two methods of infection drop method was found to be superior in inducing roots. Among the different concentrations tested, HMS (Half strength) medium supplemented with 30g/L sucrose and acetosyringone 400µM/L without NAA was found to have highest transformation frequency (58.76 + 0.79 %). The highest transformation frequency was observed when average co-cultivation period was 3.00 + 0.57 days. Further increasing the co-cultivation period has reduced the number of root induction.PCR with rol C gene specific primer of A. rhizogenes has amplified DNA product of size 1000 bp in transformed A. paniculata hairy roots, which confirms the transformation.
Key words: Agrobacterium rhizogenes, Andrographis paniculate, Hairy root.
36. Journal of Cell and Tissue Research 15(1) 4903- 4911 (2015)
Isolation, identification and metabolic characterization of cellulolytic enzyme producing bacteria
from banana pseudostem waste using phenotypic microarray technology
Patel, H. and Mahatma, L.
Department of Plant Molecular Biology and Biotechnology, N.M.C.A., Navsari Agricultural University, Navsari 396 450, Gujarat. E. mail: mahatmalalit@yahoo.co.in ,
Abstract: The purpose of this study was to isolate cellulolytic enzyme producing bacteria which may able to utilize banana pseudostem as sole carbon source for production of cellulolytic enzymes. This was attempted by isolating 10 bacteria from banana pseudostem waste. Out of the screened, isolate NAULH-5 showed high cellulolytic activity. The isolate NAULH-5 produced highest 5.97 U/mg endoglucanase, 2.27 U/mg exoglucanase and 3.55 U/mg β-glucosidase enzymes at neutral pH and 5.76 U/mg endoglucanase, 2.18 U/mg exoglucanase & 3.19 U/mg β-glucosidase enzymes at 300C. It also showed maximum 5.36 U/mg endoglucanase, 2.23 U/mg exoglucanase and 3.38 U/mg β-glucosidase enzyme production at 28 days. A cellulolytic bacterial strain NAULH-5 exhibited morphological, biochemical and cultural characteristics of the Serratia genus. The antibiotic profiling data indicated that isolate NAULH-5 showed resistant against many complex antibiotics. The taxonomic characterization was done using phenotypic microarray technology, which assigned the isolate NAULH-5 to Serratia marcescens with 0.71 SIM index. The metabolic profiling revealed that isolate NAULH-5 may able to degrade most of the complex molecule. The carbon source metabolic network indicated that it operates all general metabolic pathways for utilization of complex molecules.
Key words: Cellulolytic enzymes, Serratia marcescens
37. Journal of Cell and Tissue Research 15(1) 4913- 4919 (2015)
Isolation and characterization of metallo-alkaline serine protease from Bacillus subtilis strain CC2
Chudasama, C.J., Parmar, S.R. and Patel, H.N.
Shri A. N. Patel Postgraduate Institute, Sardar Patel University, Anand. E. mail: cjcbiochem@gmail.com
Abstract: Twenty nine bacterial cultures were isolated from the garden soil and screened for protease production. Typical growth curve of the selected isolate cc-2 was obtained and media parameter was optimized for maximum production of protease. Optimum pH for protease activity was found to be 9.0 (alkaline protease) and optimum temperature was found at 50°C. Protease derived from cc-2 was 100% inhibited by 1 mM PMSF and 93.5% with 5 mM EDTA respectively showed its nature of serine group of metallo-protease. Protease enzyme was entrapped in calcium alginate beads. Immobilized enzyme retained its activity for longer time and upon repeated use, the enzyme showed 99% activity during the second and third reuse and 80% activity on its fourth reuse in the sample.
Key words: Metallo-proteases, Bacillus
38. Journal of Cell and Tissue Research 15(1) 4921- 4928 (2015)
In vitro antagonism between biocontrol agent Trichoderma and pathogen Sclerotium rolfsii causing stem rot in Arachis hypogaea L.
Parmar Heena. J., Bodar, N.P., Bhadja, N.V., Patel, S.V., Kandoliya, U.K. and Golakiya, B.A.
Department of Biotechnology, College of Agriculture, Junagadh Agriculture University,
Junagadh -362001. E. mail: heena_odedra@rediffmail
Abstract: Six Trichoderma strains (collected from IARI, New Delhi and MTCC, Chandigarh) were tested for their ability to inhibit soil born pathogen of groundnut mainly Sclerotium rolfsii (causing stem rot on groundnut). Morphological observations of Trichoderma strains as well as phytopathogenic fungi S. rolfsii were made from culture grown at 28 0C for about one week on PDA media. The radial growth, fastest growth and coiling of test fungus of various Trichoderma strains on plant pathogenic fungi S. rolfsii were examined under LEICA phase contrast binocular light microscope. In vitro percent growth inhibition of S.rolfsii by various Trichoderma strain was recorded at 5 days after inoculation at 28 0C in the 90 cm petriplates. Results obtained from the antagonism study indicated that Trichoderma viride (NBAII Tv 23) inhibited 61% growth of phytopathogenic fungi S.rolfsii followed by T. harzianum (NBAII Th1) (55% growth inhibition of pathogen). This suggested that among different Trichoderma strains, T. viride was the best bio-control agent to inhibit in vitro growth of phytopathogen S.rolfsii which otherwise cause stem rot disease in groundnut.
Key words: Trichoderma, Sclerotium rolfsii, Arachis hypogaea
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