PROCEEDINGS - Ferdowsi University of Mashhadprofdoc.um.ac.ir/articles/a/1038728.pdfGlobal Multidisciplinary eConference 10-12 November 2013 UNESCO`s World Science Day Celebration PROCEEDINGS

Global Multidisciplinary eConference 10-12 November 2013 UNESCO`s World Science Day Celebration

PROCEEDINGS

1st Global Multidisciplinary eConference

10-12 November 2013

(UNESCO`s World Science Day Celebration)

European Scientific Institute, ESI (publishing)


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PROCEEDINGS : 1st Global Multidisciplinary eConference, 10-12November 2013 : (UNESCO`s World Science Day Celebration)European Scientific Institute, ESI, 2013. - 3 vol. (465, 544, 149 p.) : ilust. ; 28 cm Kocani, Republic of Macedonia Email: [email protected] Printed in Republic of Macedonia

ISBN 978-608-4642-14-5 (V. 1)

Copyright © 2013 by the authors, ESI and licensors All rights reserved. 2013


PROCEEDINGS

1st Global Multidisciplinary eConference

10-12 November 2013

(UNESCO`s World Science Day Celebration)


Table of ContentsTARGET COSTING BASED ON THE ACTIVITY-BASED COSTING METHOD AND A MODEL PROPOSAL………………………………………………………………1Selim Yuksel PazarcevirenDuygu Celayir

DIFFERENCES BETWEEN B2B AND B2C CUSTOMER RELATIONSHIP MANAGEMENT. FINDINGS FROM THE CZECH REPUBLIC……………………...22Karel KolisKaterina Jirinova

CARBON DYNAMICS: EVER GOING TO STOP INCREASING?...............................28Katalin Hartunga

EVALUATION OF FOREIGN INVESTMENT USE IN OIL AND GAS COMPLEX OF KAZAKHSTAN………………………………………………………………………...37Gulsara DzholdasbayevaGulimai Amaniyazova

STRATEGIC OPPORTUNITIES TO INCREASE THE IMPACT OF SCIENCE AND TECHNOLOGY IN REGIONAL DEVELOPMENT: OPEN INNOVATION AND THE STRATEGIC VALUE OF HORIZONTAL SOCIAL NETWORKS…………………...43Linda Russell

EVALUATION OF PROBABILITY MODELS ON INSURANCE CLAIMS IN GHANA...................................................................................................................................52E. J. DadeyS. Ankrah

METHODOLOGICAL PROBLEMS OF STATISTICAL CALCULATIONS OF MACROECONOMIC INDICATORS…………………………………………………….61E.P. Vasilyev

CHARACTERISTICS OF FAMILY BUSINESSES IN THE CZECH REPUBLIC………………………………………………………………………………….66Ondrej MachekJiri HnilicaMartin Brabec

NATURAL GAS MARKET DEVELOPMENT - IMPLICATIONS FOR DIVERSIFICATION OPTIONS OF EU: CASE STUDY OF SLOVAKIA…………….73Saleh Obadi MothanaMatej Korcek


LEADERSHIP SKILLS IN ORDER TO INCREASE EMPLOYEE EFFEICIENCY......................................................................................................................85Mirjana NedovicZeljko SudarcDarija Ivankovic

APPORT A L’ETUDE DES RISQUES DE L’INFRASTRUCTURE FERROVIAIRE,QUELLES IMPLICATIONS SUR LA DISTRIBUTION PHYSIQUE DES VEHICULES DU SITE DE PRODUCTION AUTOMOBILE DE MELLOUSA (TANGER - MAROC)?.................................................................................91Jihad JAMÏHajar AMRANIA

FRENCH “FIDUCIE” AND RUSSIAN “(TERMINOLOGICAL PECULIARITIES)………………………112

Irina Gvelesiani

STUDYING THE “LEGAL FLOWS” AS A MULTIDISCIPLINARY METHOD TO PROMOTE CONSTITUTIONALISM AS A COMMON PROPERTY OF MANKIND…………………………………………………………………………………118Michele CarducciAnna Silvia Bruno

SUBJECT OF A PROCEDURAL SETTLEMENT SUBJECT (ITS TERMS) IN CIVIL PROCEDURE LAW OF THE REPUBLIC OF LATVIA……………………………...125Sintija Finka

-………………………………………………………………………………...136

COSMOPOLITANISM AND DEMOCRACY BEYOND STATES: THE COSMOPOLITAN DEMOCRACY PROJECT………………………………………...144Ricardo Norberto Medina

DIGITAL SCHOOL AS NEW ORGANIZATION BASED ON TECHNOLOGIES OF INFORMATION AND COMMUNICATION…………………………………………...153Loreta MamaniEva Cipi

PLANTING THE SCIENCE SEED: ENGAGING STUDENTS IN PLACE-BASED CIVIC ACTIONS………………………………………………………………………….161Kevin O’ConnorRobert Sharp

ANCIENT AND CONTEMPORARY EDUCATIONAL SYSTEMS (SIGNIFICANCE OF PAST IN THE FORMATION OF FUTURE)……………………………………….168Irina Gvelesiani


NATIONAL UNITY AT THE UNIVERSITY LEVEL: IMPORTANCE OF CIVILISATIONAL DIALOGUE AND WAY FORWARD…………………………….173Lee Wei ChangBaharuddin AzizanMuhammad Amran

IMPLEMENTATION OF DIGITAL SIGNATURE IN WEB TO IMPROVE THE QUALITY OF MANAGEMENT INFORMATION SYSTEMS IN EDUCATION SECTOR…………………………………………………………………………………...187Arbnor HaliliBlerim RexhaAlbert Aliu

ORGANISATIONAL POLITICS AND THEIR EFFECT ON WORKPLACE LEARNING………………………………………………………………………………..198Karen Cacciattolo

CYBER RESEARCH MENTORING IN SCIENCE IN PHILIPPINE PRIVATE UNIVERSITIES: LEVERAGING ONLINE COLLABORATION TECHNOLOGIES AND OFFSHORE-BASED ALUMNI……………………………………………………212Michael Paul A. JallorinaIvan B. CulabaMinella AlarconJames Simpas Erika B. AranasBenjamin B. Dingel

EDUCATIONAL SERVICE MARKET AND THE PROSPECTS FOR THE DEVELOPMENT OF KNOWLEDGE IN GEORGIA…………………………………222Lali Chagelishvili-AgladzeKoba Chagelishvili

THE ACADEMIC PROFESSION AMONG FACULTY AT SAUDI UNIVERSITIES…………………………………………………………………………...227Mohsen A Al Mohsen

HIGHER EDUCATION FUNDING IN THE CZECH REPUBLIC IN THE YEARS 2001-2011…………………………………………………………………………………241Jaromir Tichy

CREATIVITY AT SCHOOL? CONCLUSIONS FROM POLISH STUDY………….256Jewdokimow Marcin

SCIENTIFIC LITERACY IN THE DIGITAL AGE: TOOLS, ENVIRONMENTS AND RESOURCES FOR CO-INQUIRY………………………………………………………263Alexandra Okada


EDUCATION IN A WORKPLACE FROM THE PERSPECTIVE OF A LEARNING ORGANIZATION…………………………………………………………………………275Renata Tomaszewska-Lipiec

BUDDHIST INFLUENCE ON WINDSCREEN LITERATURE: A STUDY ON STICKER PHRASES POSTED ON THE THREE WHEEL TAXIES IN SRI LANKA…………………………………………………………………………………….284Lokeshwari Sandamali Karunarathna (B.A. Peradeniya)

LA CRUZ Y EL WATESINK: SINCRETISMO TRANSCULTURAL DE LA IDIOSINCRASIA MAYA Q´ EQCHI´ EN RITUALES Y COTIDIANIDAD………..306David Caballero Mariscal

INFLUENTIAL INDIVIDUALS ON SOCIAL NETWORKS: AN EXAMPLE OF FACEBOOK IN GEORGIA……………………………………………………………...320Zakaria BabutsidzeTengiz LomitashviliKoba Turmanidze

PLAYAS Y MANEJO APTITUD RECREATIVA EN PLAYAS TURÍSITICAS DE MANZANILLO, COLIMA, MÉXICO…………………………………………………..331Silva-Iñiguez, L.Gutiérrez-Corona, C.G. Pérez-López, R. Sosa-Avalos, R. López-Magaña, S.A.

THE BENEFITS OF SIGN LANGUAGE FOR DEAF CHILDREN WITH ANDWITHOUT COCHLEAR IMPLANT(S)………………………………………………...341Krammer Klaudia

FEMINIST CRITICISM AS A WAY OF LEARNING THE CATEGORIES OF “PRIVATE” AND “PUBLIC” IN KAZAKHSTAN GENDER LITERATURE………350Zhanna Tolysbayeva

CHALLENGES OF REMOTE SENSING IMAGE FUSION TO OPTIMIZE EARTH OBSERVATION DATA EXPLOITATION……………………………………………..355Christine Pohl

EXPLORING THE POTENTIAL OF ONLINE GROUP COUNSELING : A CASE-STUDY FOR MAURITIAN STUDENTS PRESENTING THE INTERNET ADDICTION DISORDER SYNDROME………………………………………………..366Ponnusawmy HarrySantally Mohammad Issack

«LE POETIQUE HABITE DANS L’HOME » – COMMENTAIRES SUR HEIDEGGER……………………………………………………………………………...380Gorun Manolescu


THE STATE OF SCIENCE IN 2013……………………………………………………..385Peter StallingaIgor Khmelinskii

LAS COMPETENCIAS: UNA PROPUESTA CONCEPTUAL HACIA LA UNIFICACIÓN MULTIDIMESIONAL EN EL CONTEXTO DE LOS RECURSOS HUMANOS………………………………………………………………………………...391Cuahtémoc Guerrero DávalosGermán Aníbal Narváez Vásquez

THE DESTROYED MINARET OF THE UMAYYAD MOSQUE OF ALEPPO, THE SURVEY OF THE ORIGINAL STATE…………………………………………………403Gabriele FangiWissam Wahbeh

THE CASTLE, THE TOWN OF RASHAN (TREPÇA)..................................................410Pajazit Hajzeri

TOPIC: ALBANIAN – GREEK DICTIONARIES, COMPILED IN THE 90’S........................................................................................................................................415Kostando Ksera

MODES OF THINKING IN LANGUAGE STUDY…………………………………….421Jesus Martinez del Castillo

THE HISTORIC BACKGROUND OF “LLAIKO VIMA” NEWSPAPER (1945-2012).......................................................................................................................................432Oleta Polo

DIALECTICAL UNITY OF LANGUAGE AND SPEECH (ON THE MATERIAL OF WORDS ENDING ON THE SUFFIX -LY IN ENGLISH LANGUAGE)……………..437Nino Kemertelidze

THE ANALYSIS OF HAMIT YERGALIYEV POETRY……………………………441Kamarova Nagbdu

SOME PARALLELS IN THE WORKS OF AKAKI TSERETELI AND ADAM MITSKEVICH…………………………………………………………………………….445Manana Kajaia

QUALITY EVALUATION OF NEW VEGETARIAN BEAN SPREADS…………….453Asnate KirseDaina Karklina


RICE TERRACES MACRO-INVERTEBRATES AS INDICATOR OF WATER QUALITY IN INDIGENOUS COMMUNITIES OF THE CORDILLERA REGION NORTH LUZON, PHILIPPINES………………………………………………………..458Banwa,Tules.P.Bawer,Maria.Cyrila.C.Estoque, Henrilyn, M.,Maganon, Zorayda B.

THE POTENTIAL OF TITHONIA DIVERSIFOLIA (WILD SUNFLOWER) AS ORGANIC FOLIAR FERTILIZER..................................................................................465Dela Pena,CennyI. Bartolome, Leah. D.Banwa, Tules.P.

MACROFLORAL BIODIVERSITY CONSERVATION IN IFUGAO……………….469Napoleon K. Taguiling

AVAILABILITY, PREPARATION AND USES OF HERBAL PLANTS IN KALINGA, PHILIPPINES……………………………………………………………………………..483Christina L. AmmakiwMarymina P. Odiem

SCHIZOSTACHYUM LUMAMPAO (BYUYU): ITS DIVERSE ETHNO-BOTANICAL USES BY LUBUAGAN SUB TRIBE OF KALINGA IN NORTH LUZON PHILIPPINES......................................................................................................................490Banwa,T.P. Bawer,M.C.C.

CAN KIDS LEARN BY THEMSELVES USING TECHNOLOGY AND WHAT DO THEY LEARN?....................................................................................................................500Shabnam HusnooGoonesh K Bahadur Mohammad Santally

GAUSSIAN DISTRIBUTION OF BREAKDOWN SHOTS IN NON-UNIFORM FIELDS…………………………………………………………………………………….511John Tarilanyo Afa

AN AUTONOMOUS SIMULATION BASED SYSTEM FOR ROBOTIC SERVICES IN PARTIALLY KNOWN ENVIRONMENTS…………………………………………518Eva Cipi

DESIGN AND DEVELOPMENT OF NOISE SUPPRESSION SYSTEM FOR DOMESTIC GENERATORS.............................................................................................526Umar HammadAhmad AizazAbid Ali KhanTaimur Qurehi


CONTEXT AWARE MOBILE COMPUTING AS A CHALLENGE FOR DEVELOPERS AND SOFTWARE ENGINEERS:A REVIEW.....................................534Rabail Tahir

IMPACT OF EMPIRICAL PATH LOSS MODELS ON SPATIAL TV WHITE SPACE UTILIZATION…………………………………………………………………………….543Nasir FarukAdeseko A. AyeniYinusa. A. Adediran

MICROMEGAS PERFORMANCE BASED IN ARGON-ISOBUTANE AND ARGON-DEMETHYL-ETHER.........................................................................................................548Hamid Mounir Mustapha Haddad Seddik Bri

L’IMPACT DU SOUTIEN SOCIAL SUR LE SUJET MALADE DU CANCER ET SURSON PARCOURS DE MALADIE ETUDE QUALITATIVE………………………….557Laura Petrescu Perlier

HEALTH CARE WASTE MANAGEMENT PRACTICES IN THE HOSPITALS OF TABUK CITY……………………………………………………………………………...584Christina L. AmmakiwJudith S.BalicagMarymina P. Odiem

DEVELOPMENT & OPTIMIZATION OF ALLYL AMINE ANTIFUNGAL NANOEMULGEL USING 23 FACTORIAL DESIGN: FOR THE TREATMENT OF TINEA PEDIS……………………………………………………………………………...597UrmilaSri Syamala

DESIGN, DEVELOPMENT AND EVALUATION OF FENOFIBRATE AND ROSUVASTATIN NANODROPLETS…………………………………………………..606Raman Suresh Kumar

REDUCING STIGMATISATION AND DISCRIMINATION THROUGH PROVISION OF CARE AND SUPPORT TO FELLOW PERSONS LIVING WITH THE HUMAN IMMUNODEFICIENCY VIRUS (PLHIV) IN GHANA: THE CASE OF THE ‘MODELS OF HOPE’.........................................................................................................618Raphael Avornyo

BACTERIAL MUTATION; TYPES, MECHANISMS AND MUTANT DETECTION METHODS: A REVIEW………………………………………………………………….628Mohammad B. Habibi NajafiParnian Pezeshki

BACTERIAL MUTATION; TYPES, MECHANISMS AND MUTANT DETECTION ; ,METHODS: A REVIEW………………………………………………………………….628Mohammad B. Habibi NajafiParnian Pezeshki


FROM OBSERVED VISIBLE HUMAN BEHAVIOR TO ANIMATED HUMANOID 3D COMPUTER MODELS - BIVARIATE NOTATIONS AS A PSYCHOLOGICAL METHOD FOR SYSTEMATIC OBSERVATION……………………………………..639Zysk, Wolfgang

SELF-PACED LEARNING WITH 3D HUMANOID VIRTUAL TUTORS…………651Zysk, WolfgangFilkov, Rumen

SOLUCIÓN AL PROBLEMA COMBINATORIO USANDO UNA FUNCIÓN DE CLASE HÖLDER……………………………………………………………………….658Anna TarasenkoOleksandr KarelinManuel González-Hernández

THE PROVISION ECOLOGICAL FAVOURABLE ENERGY, NOT REQUIRING RAW MATERIAL………………………………………………………………………...666Mashkov V.V


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BACTERIAL MUTATION; TYPES, MECHANISMS ANDMUTANT DETECTION METHODS: A REVIEW

Mohammad B. Habibi NajafiParnian Pezeshki

Department of Food Science & Technology, Ferdowsi University of Mashhad, Mashhad, Iran

AbstractMutation is a very important concept in biology today that leads to variations in

genes. A mutation is a permanent alteration in the sequence of nitrogenous bases of a DNA molecule. The result of a mutation is generally a change in the end-product specified by that gene. In some cases, a mutation can be beneficial if a new metabolic activity arises in amicroorganism, or it can be detrimental if a metabolic activity is lost. Mutations can be spontaneous, or induced by a mutagen in the environment. Mispairing is probably mostly due to cellular processes such as Tautomeric shift of bases , oxidative damage to DNA , Depurination and Deamination or caused by “environment”, i.e, chemicals, radiation, viruses,diet and lifestyle (Mutagens). Substitution of a nucleotide and Deletion or addition of them is two mechanisms of mutation. Mutation in bacteria has some results such as missense, nonsense, silent, frameshift, lethal, suppressor and conditional lethal mutation. Identifying these mutations requires detection methods. Classic methods i,e, Replica plating, Penicillin enrichment, Ames test, Use of chromogenic substrate and novel tests such as Polymerase chain reaction (PCR) and Gel electrophoresis, Gene probes and Southern blotting, DNA sequencing and DNA microarray are some of these methods that are highlighted in this review.

Keywords: Mutation, Spontaneous and induced mutation, mechanisms of mutation and their results, detection methods

IntroductionToday bacteria are an important tool in the study of genetics and biotechnology, but

for 40 years after the rediscovery of Mendel's work and the rebirth of genetics, they were considered too simple to have genes, undergo mutation, or reproduce sexually. This is not surprising - bacteria are so small that it's very difficult to study individuals. Scientists had long observed differences between bacterial colonies, but had never realized that these differences were the results of mutations. Mutations are a very important concept in biology today. Mutations lead to variations in genes. These genes can have a good or bad influence in the characteristics of an organism. Variations are also very important in evolution. Without variations evolution would not be possible, and changes in any part of the environment affecting the organism could make the organism become extinct [1,2,3].

A mutation is any heritable change in DNA sequence. This may, or may not, affect the phenotype of the organism [4,5,6,7,8]. The term “mutation” was coined by Hugo de Vries, which is derived from Latin word meaning “to change”[5].

The process of mutation is called mutagenesis and the agent inducing mutations is called mutagen. (Organisms selected as reference strains are called wild type, and their progeny with mutations are called mutants). Changes in the sequence of template DNA (mutations) can drastically affect the type of protein end product produced [4,7].


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Types of mutationsMutations can be classified by the kinds of alterations in the DNA, or by whether the

mutation was spontaneous, or induced by a mutagen in the environment. Mispairing is probably mostly due to cellular processes such as Tautomeric shift of bases , oxidative damage to DNA , Depurination and Deamination or caused by “environment”, i.e, chemicals,radiation, viruses, diet and lifestyle (Mutagens)[4,5,6,8,9].

Mechanisms of mutationa. Substitution of a nucleotide: Base substitution, also called point mutation, involves

the changing of single base in the DNA sequence. This mistake is copied during replication to produce a permanent change. If one purine [A or G] or pyrimidine [C or T] is replaced by the other, the substitution is called a transition. If a purine is replaced by a pyrimidine or vice-versa, the substitution is called a transversion. This is the most common mechanism of mutation.

b. Deletion or addition of a nucleotide: deletion or addition of a nucleotide during DNA replication. When a transposon (jumping gene) inserts itself into a gene, it leads to disruption of gene and is called insertional mutation [2, 4,5,6,7].

Mutation SourcesMutations arise from a variety of sources, including:

Tautomeric shift of basesEach of the bases in DNA can appear in one of several forms, called tautomers, which

are isomers that in the positions of their atoms and in the bonds between the atoms. The keto(C-O) and amino(C-NH2) form of each base is normally present in DNA, whereas the imino(C=NH) and enol(C-OH) forms of the bases are rare (Figure 1).Conversion of keto group in thymine & guanine to enol form and changing amino group in adenine & cytosine to imino form, are examples of tautomerization. Naturally, A in amino form pairs with T in keto form, whereas A in its imino form pairs with C and T in enol form match with G [1,6,8] (Figure 2).

Figure 1 : Pairing between the normal (keto) forms of the bases.

Figure 2 : Mismatched bases. (a) Mispairs resulting from rare tautomeric forms of the pyrimidines; (b) mispairs resulting from rare tautomeric forms of the purines.


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DepurinationDepurination consists of the interruption of the glycosidic bond between the base and

deoxyribose and the subsequent loss of a guanine or an adenine residue from the DNA. in replication, the resulting apurinic sites cannot specify a base complementary to the original purine. So, efficient repair systems remove apurinic sites. Under this certain conditions, a base can be inserted across from an apurinic site; this insertion will frequently result in a mutation [6] (Figure 3) .

Figure 3 : The loss of a purine residue (guanine) from a single strand of DNA. The sugar-phosphate backbone is left intact.

DeaminationThe deamination of cytosine yields uracil. Unrepaired uracil residues will pair with

adenine in replication, resulting in the conversion of a G–C pair into an A–AT transition). Another example for deamination, is conversion of 5-methylcytosine to thymine [1,2,6] (Figure 4) .

Figure 4 : Deamination of (a) cytosine and (b) 5-methylcytosine.

Oxidatively damaged basesActive oxygen species, such as superoxide radicals (O2·), hydrogen peroxide (H2O2),

and hydroxyl radicals (OH·), are produced as by-products of normal aerobic metabolism. They can cause oxidative damage to DNA. For example, The 8-oxo-7-hydrodeoxyguanosine (8-oxodG, or GO) product frequently mispairs with A, resulting in a high level oftransversions. Thymidine glycol which induced of hydroxyl radical attack to thymine , blocks


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DNA replication if unrepaired but has not yet been implicated in mutagenesis [6,10] (Figure 5) .

Figure 5 : DNA damage products formed after attack by oxygen radicals.

Ultraviolet irradiationUltraviolet light (opt 250 nm) causes bonds to form between adjacent pyrimidine

residues (commonly referred to as thymine dimers, although the effect can also occur with cytosine) in the same polynucleotide strand. These are called pyrimidine dimers. UV irradiation can result in the formation of covalent links between pyrimidine dimers (Figure 6) .These bonds distort the DNA conformation and inhibit DNA replication and transcription [1,2,6,7,11].

Figure 6 : Structure of thymine dimers.

Chemical mutagensVariety of chemical mutagens have been identified that are classified into three

groups based on their function [1,2,6,8].First group are those agents that act by chemically modifying a base on the DNA so

that it resembles a different base. For example, nitrous acid causes an oxidative deamination in which amino groups are converted to keto groups and thus cytosine residues for example will be converted to uracil. Uracil will be capable of pairing with adenine, thus causing a change from a C–G pair to T–A. Similarly deamination of adenine creates the base hypoxanthines, which will base-pair with cytosine. Some types of chemical agent are not incorporated into the DNA but instead alter a base, causing specific mispairing. They act against the DNA within cells, rather than against isolated DNA. Certain alkylating agents, such as ethylmethanesulfonate (EMS) and nitrosoguanidine (NG), add alkyl groups (an ethyl group in EMS and a methyl group in NG) to many positions on all four bases, mutagenicity is best correlated with an addition to the oxygen at the 6 position of guanine to create an O-6-


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alkylguanine. This addition leads to direct mispairing with thymine, and would result in GC

Figure 7 : Alkylation-induced specific mispairing. The alkylation (in this case, EMS-generated ethylation) of the O-6position of guanine and the O-4 position of thymine can lead to direct mispairing with thymine and guanine, respectively.

Second group are mutagens that induct frameshift formation. The interchalating agents that include proflavin, acridine orange and ethidium bromide are planar molecules, which mimic base pairs and are able to slip themselves in (interchalate) between the stacked nitrogen bases at the core of the DNA double helix. In this intercalated position, the agent can cause single-nucleotide-pair insertions or deletions. Intercalating agents may also stack between bases in single-stranded DNA; in so doing, they may stabilize bases that are looped out during frameshift formation.

Third group are type of agent that acts only against growing cells consists of the base analogues such as 5-bromouracil. Despite its name, this is an analogue of thymine and adenine. 5-BU causes mutations when it is incorporated in one form and then shifts to another form (Figure 8). In its normal keto state, 5-BU pairs like thymine (5-BUT). Thus, 5-BU is

GC transitions, but in its enol form, 5-BU pairs like cytosine (5-BUC), so 5-BU is

AT transitions (Figure 9).

Figure 8 : Enhanced tautomerism by the base analogue 5-bromouracil.


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Figure 9 : The mechanism of 5-BU mutagenesis. (a) In its normal keto state, 5-BU pairs like thymine (5-BUT).

Thus, 5-transitions. (b) In its ionized form, 5-BU pairs like cytosine (5-BU

C). Thus, 5-BU is misincorporated across

Results of mutationMissense mutation: Missense mutations are DNA mutations which lead to changes

in the amino acid sequence (one wrong codon and one wrong amino acid) of the protein product [1,4,5].

Nonsense mutation: A mutation that leads to the formation of a stop codon is called a nonsense mutation. Since these codon cause the termination of protein synthesis, a nonsense mutation leads to incomplete protein products [1,4,5].

Silent mutation: Sometimes a single substitution mutation change in the DNA base sequence results in a new codon still coding for the same amino acid. Since there is no change in the product, such mutations are called silent [1,4,5] (Figur 10).


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Figure 10 : Mechanism of silent mutation.

Frameshift mutation: Frameshift mutations involve the addition or deletion of base pairs causing a shift in the “reading frame” of the gene. This causes a reading frame shift and all of the codons and all of the amino acids after that mutation are usually wrong. Since the addition of amino acids to the protein chain is determined by the three base codons, when the overall sequence of the gene is altered, the amino acid sequence may be altered as well [1,4,5,6].

Lethal mutation: Sometimes some mutations affect vital functions and the bacterial cell become nonviable. Hence those mutations that can kill the cell are called lethal mutation [5,7].

Suppressor mutation: It is a reversal of a mutant phenotype by another mutation at a position on the DNA distinct from that of original mutation. True reversion or back mutation results in reversion of a mutant to original form, which occurs as a result of mutation occurring at the same spot once again [1,2,4,5,12] (Figur 11).

Figure 11 : Mechanism of suppressor mutation.

Conditional lethal mutation: Sometimes a mutation may affect an organism in such a way that the mutant can survive only in certain environmental condition. Example; a temperature sensitive mutant can survive at permissive temperature of 35oC but not at restrictive temperature of 39oC [1,5,7].

Isolation and identification of mutantsIn general, changes at the amino acid level in a protein will have a consequential

effect on the protein’s activity, usually detrimental. Thus, an important metabolic pathway


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may be blocked by the lack of functional enzyme resulting in identifiable phenotypes [8].Thisphenotypes can be classified in to three main groups:1- mutants that are resistant to antibiotics, or to specific bacteriophages, toxic chemicals or any other agents that are usually lethal or inhibitory to the parent cell;2- auxotrophs, i.e. mutants that require some additional growth factor, such as an amino acid;3- mutants that are unable to use a particular growth substrate (usually a sugar)[13].

Methods for mutant selectionReplica plating

In this procedure, the mutagenized culture is plated out to obtain single colonies on a nutrient medium on which mutants and parents will grow. After incubation, the colonies are replicated, using a sterile velvet pad, onto a minimal agar plate and then a similar plate to which the appropriate supplement (in this case, histidine, since we are looking specifically for histidine auxotrophs) has been added. Histidine requiring auxotrophs will be unable to grow on the first plate, but will grow on the second one. Thus, mutant colonies can be identified with localization of colonies that have failed to grow in second plate in comparison with first one [1,5,14] (Figur 12).

Figure 12 : Replica plating to isolate auxotrophic mutants.

Penicillin enrichmentSome antibiotics (i.e penicillin) acts only against growing bacteria. Cells that are

stopped their growth for any reason, relatively not sensitive. Resuspending the bacteria in a minimal medium and adding penicillin will kill the parental cells, which are able to grow, while allowing the non-growing mutants to survive [1,5,14,15,16] (Figur 13).

Figure 13 : Mutant screening using penicillin enrichment.


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Ames testThe Ames test uses several strains of the bacterium Salmonella typhimurium that

carry mutations in genes involved in histidine synthesis i.e. it is an auxotrophic mutant, so that they require histidine for growth. The method tests the capability of mutagen in creating mutations that can result in a reversion back to a non-auxotrophic state so that the cells can grow on a histidine-free medium. The bacteria are spread on an agar plate with a small amount of histidine. This small amount of histidine in the growth medium allows the bacteria to grow for an initial time and have the opportunity to mutate. When the histidine is depleted,only bacteria that have mutated to gain the ability to produce their own histidine will survive. The plate is incubated for 48 hours. The mutagenicity of a substance is proportional to the number of colonies observed [7,14 ,17,18] (Figur 14).

Figure 14 : Ames test procedure.

Use of chromogenic substrateMutants that are not able to utilize a particular carbon source (lactose, for example)

can be isolated with use a chromogenic substrate that shows an easily detectable color change when acted on by the enzyme concerned. In this case the enzyme is -galactosidase, which catalyses the hydrolysis of lactose into its constituent sugars glucose and galactose. A commonly used chromogenic substrate for -galactosidase is 5-bromo-4-chloro-3-indolyl- -D-galactoside, more popularly known as X-gal. This is a synthetic analogue of the natural substrate, containing a dye linked to galactose. X-gal itself is colorless; the color of the dye is only manifest when it is released by hydrolysis of the linkage by -galactosidase. Lac+colonies will be blue on a medium containing X-gal and colonies that do not produce -galactosidase will be white[1,19].Polymerase chain reaction (PCR) and Gel electrophoresis

In this method, target gen (mutant sequence in this case) can be increased exponentially. If the mutation has caused a significant change in the size of a specific gene (such as an insertion or a deletion) it will be detect by a change in the size of the PCR product, using gel electrophoresis [1,20,21,22,23].

Gene probes and Southern blottingThis method involves separating fragments of DNA by electrophoresis in an agarose

gel and transferring them to a filter which can then be hybridized with the labeled probe. By using highly specific probes, detection of difference in the sequence is possible [1,14,21,23].


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DNA sequencingDNA sequencing template is amplified using PCR and oligonucleotide primers

flanking the region of interest. The amplified fragment is directly cycle sequenced using fluorescent sequencing primers, Sanger dideoxy sequencing chemistry and an enzyme mixture of a mutant Taq DNA polymerase and thermostable pyrophosphatase. The sequence ladders produced are analyzed on a real-time, automated four-color sequencing system. The method produces sequence ladders from unpurified PCR fragments of sufficiently high quality such that heterozygotes can be reproducibly detected and identified by software that recognizes signal-strength patterns indicative of mixed-base positions [1,24,25,26].

DNA microarray Base-pairing or complementarity is the principle behind this emerging technology.

The potential applications of microarray technology are gene expression profiling and identification of gene sequences (including sequences that bear mutations). In this method, a large number of DNA fragments are placed on a glass slide. The fragments are allowed to complement or bind with the labeled DNA (probes), which hybridize with the DNA on the glass slide. The amount of hybridization is then analyzed in each spot on the slide. The genes are given a color where the hybridized ones are colored red and the genes that are hybridized least are colored green. It has been reported that microarray technology could be used in monitoring chromosome gains and losses, tumor classification, drug discovery and development [27,28,29].

ConclusionMutations lead to variations in genes that are very important in evolution. A mutation

is any heritable change in DNA sequence. Mutation can be beneficial or it can be detrimental. Mutations can be spontaneous due to cellular processes, or induced by a mutagen in the environment. Substitution of a nucleotide and Deletion or addition of them is two mechanisms of mutation. Mutation in bacteria has some results that may, or may not; affect the phenotype of the organism. Several methods have been developed for the mutant detection, of these methods molecular methods Due to the high sensitivity and specificity,and also more time saving than the classical methods is recommended.

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