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Explain the different variations on genetic testing and how they can be used in identification and diagnosis.Learning OutcomeProcedures for Identifying Pathogens and Diagnosing Infections (2)11Genotypic MethodsThe exact order and arrangement of the DNA code is unique to each organism. With a technique called hybridization, it is possible to identify a bacterial species by analyzing segments of its DNA. This requires small fragments of single-stranded DNA (or RNA) called probes that are known to be complementary to the specific sequences of DNA from a particular microbe. The test is conducted by extracting unknown test DNA from cells in specimens or cultures and binding it to special blotter paper. After several different probes have been added to the blotter, it is observed for visible signs that the probes have become fixed (hybridized) to the test DNA. The binding of probes onto several areas of the test DNA indicates close correspondence and makes positive identification possible (see figures 10.5 and 10.6).CP 40: Explain the different variations on genetic testing and how they can be used in identification and diagnosisGenotypic MethodsDNA Analysis Using Genetic Probes2Genotypic MethodsDNA Analysis Using Genetic Probes

CP 40: Explain the different variations on genetic testing and how they can be used in identification and diagnosisRONI AFRIANSYA3Genotypic MethodsDNA Analysis Using Genetic Probes

CP 40: Explain the different variations on genetic testing and how they can be used in identification and diagnosisTIARA DINI HARLITA4Another genetic-based technique often brought in to analyze outbreaks or epidemics is pulse-field gel electrophoresis (PFGE). This rapid method of DNA typing makes a restriction fragment fingerprint of the suspected pathogens on a single gel that identifies and compares the isolates alongside each other. It is especially useful to trace the causative agents to a particular source or reservoir (see Insight 27.1).CP 40: Explain the different variations on genetic testing and how they can be used in identification and diagnosisGenotypic MethodsDNA Analysis Using Genetic ProbesOCTAVIA PERMATA SARI5Genotypic MethodsDNA Analysis Using Genetic ProbesCP 40: Explain the different variations on genetic testing and how they can be used in identification and diagnosis

OCTAVIA PERMATA SARI6The polymerase chain reaction (PCR) is one of the most valuable high technology tools for DNA and RNA analysis. A review of chapter 10 (figure 10.7) will remind you how it can rapidly synthesize hundreds of thousands and even millions of copies of a particular segment of DNA. It is both highly sensitive and specific, amplifying minute quantities from unknown samples that would otherwise be lost. It is one of the most powerful techniques in forensics and molecular biology, but it can also be applied to the identification of microbes. With this method, laboratories can often completely dispense with traditional culturing techniques and go directly to diagnosis in a few hours. CP 40: Explain the different variations on genetic testing and how they can be used in identification and diagnosisGenotypic MethodsRoles of the Polymerase Chain Reaction and Ribosomal RNA in Identification7

WIDODOPCR-based tests are used routinely for detecting HIV, Lyme disease, the human papilloma virus, tuberculosis, hepatitis, and a number of other bacterial and viral infections (figure 17.6). In many laboratories, the PCR or a similar rapid test is the method of choice for identifying gonorrhea and chlamydiosis. One version of this technique called realtime PCR is able to pull DNA from a sample register the amount that is formed, and identify it.CP 40: Explain the different variations on genetic testing and how they can be used in identification and diagnosisGenotypic MethodsRoles of the Polymerase Chain Reaction and Ribosomal RNA in Identification

INDAH SULISTIYAWATI9One particularly versatile aspect to PCR is that it can be combined with other technologies for additional applications. For example, if the sample being analyzed contains mainly RNA (from an RNA virus or ribosome), the initial introduction of reverse transcriptase will convert the RNA in a sample to DNA, and this DNA can then be amplified by PCR. This techniqueRT-PCRis valuable for verifying RNA-based viruses such as rabies virus (see figure 17.17 e ). DNA of PCR origin can be analyzed with the other tools of technology: it can be probed, hybridized, fingerprinted, and sequenced.CP 40: Explain the different variations on genetic testing and how they can be used in identification and diagnosisGenotypic MethodsRoles of the Polymerase Chain Reaction and Ribosomal RNA in Identification

Figure 17.17 Summary of methods used to diagnose viral infections.TISNA SENDY PRATAMA10One of the most viable indicators of evolutionary relatedness and affiliation is comparison of the sequence of nitrogen bases in ribosomal RNA, a major component of ribosomes. Ribosomes have the same function (protein synthesis) in all cells, and they tend to remain more or less stable in their nucleic acid content over long periods. Thus, any major differences in the sequence, or signature, of the rRNA is likely to indicate some distance in ancestry.This technique is powerful at two levels: It is effective for differentiating general group differences (it was used to separate the three superkingdoms of life discussed in chapter 1), and it can be finetuned to identify at the species level (for example, in Mycobacterium and Legionella ).CP 40: Explain the different variations on genetic testing and how they can be used in identification and diagnosisGenotypic MethodsRoles of the Polymerase Chain Reaction and Ribosomal RNA in IdentificationTISNA SENDY PRATAMA11Procedures for Identifying Pathogens and Diagnosing Infections (3)12Immunologic Methods