In food industry such as the production of cheese, bacteria takes a significant role on the wide variety of cheese. Lactic acid bacteria(LAB), a bacteria that can be found in the production of cheese, its stress gene was investigated in the experiment by using various biochemical and genetic techniques to identify and extract.
The characterisation of the strain illustrates how identification of strains differ using different methods, such as gram stain and 16s rRNA screening. After the characterisation, the stress gene isolation assist the further understanding of the gene on LAB be giving different stress in future work. Aims
Whole experiment can be separated into two parts
(I) The characterisation of the isolate(lactococcal species) given by * Using simple biochemical tests
* Using genetic tests by extraction of total genomic DNA
* Amplifying the gene encoding for the 16s rRNA by PCR
* Understanding the fermentation growth kinetics in controlled liquid media to know the capability of organisms * The investigation of the digestion of the 16s rDNA amplicon with Hinf1 on 16s rDNA strains
(II) The amplification and cloning of a specific stress gene given by * Designing primers for cloning
* cloning stress gene into cloning vector (pGEM-T)
* cloning stress gene into expression plasmid(pET23b) – not done
Lactic acid bacteria (LAB)
LAB are defined as bacteria that produce lactic acid as their major fermentation production. The examples of LAB include Streptococcus, Enterococcus, Leuconostoc and Lactococcus. They are gram-positive rods or cocci bacteria with high AT content. (Prescott et. al, 2008). There are few applications of LAB in food, medical and biotechnological industries. The major use of LAB is on the food industries. The non-pathogenic species, which helps the process of fermentation of different food such as dairy products, meat and vegetables. They mainly convert the hexose sugar exist in food to lactic acid( Makarova et al., 2007). Recent research shows that they can produce antimicrobial compounds that have probiotic properties, they also helps different medical aspects. It includes “antitumoral activity, reduction of serum colesterol, alleviation of lactose intolerance, stimulation of the immune system, stabilization of gut microflora”(Faid et. al, 2009). Another application in medical field is the production of bacteriocins by LAB. A more well known one is Nisin, which is a lantibiotic produced by Lactococus lactis.(Anastasiadou et.al, 2009) Different examples are further illustrated in following paragraphs indicating the industrial importance. Food Production
Feta Cheese, which is a Greek cheese traditionally made from raw milk without starter cultures. The contemporary production is in well-structured dairies. Ewe milk is thermized(treated at 63 to 65°C for 16 s)( GRIFFITHS, 1998) with yogurt as starter. From Feta Cheese, different species of LAB can be identified, they are responsible for the unique flavour of Feta cheese.(Arvanitis et. al, 2009). LAB can also be used to improve the preservation of ready-to-eat(RTE) seafood such as cooked and peeled tropical shrimp and cold-smoked salmon. France is the second largest shrimp exporter in Europe and largest salmon exporter in the world. Technology for preservation is significant for them. Chemicals such as nitrates, sulfates and sorbate can be added to preserve shrimp in France. However, these chemicals are forbidden for smoked salmon. Under these circumstance, biopreservation plays a significant role for smoke salmon industries. (Cardinal et al., 2009) According to a review from Rodgers, “Biopreservation is a preservative method that uses the growth of selected microorganisms on a food product to control the undesirable (spoiling and pathogenic) microflora”(Rodgers, et al. 2003) There are few pathogens such as Vibrio spp., Salmonella spp. and Staphylococcus spp. can be found in RTE...
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