Polymerase chain reaction text

One of the most significant technological advances in molecular biology is the development of th e polymerase ch ain reaction (PCR). This procedu re can be used to detect single copy gen es or diagnose microbia l infec tions in minute samples, due lo Lh e in vi tro exponentia l a mplification of s pecific nucleic acid sequen ces . The technique has been res tricted to amplifying extracted target DNA or RNA sequen ces. precluding correla tions between cytological or his tological features and molecular findings. However . by lh e publica tion of PCR in situ Hybridization , lhis limit.alion is overcome. This teArt des cribes in detail a technique lhal marries surgical pathology and molecular b iology. wi th markedly improved sensitivi ty over classic in s itu hybridization. The infections cau sed by human papillomavirus and human immunodefi c ien cy virus l ar e used effec tively throughout the book lo illus trate lhe application of a variety of m olecular techniqu es, a lthough the author em ph asizes that th e procedures are broadly applica ble to gen etically based, as well as other infectious, human diseases. The a uthor s ets out to descr ibe lhe PCR in s itu technique in comprehen s ible terms for both lhe su rgical path ologist and the molecula r biologist.. Hence, th e first two chapters are extremely basic, focu s ing on lhe s tructure of DNA and defining the 'ja rgon · associa ted with molecular biology. In Chapter 3, lhe reader is finally introduced to the conventional PCR technique. The procedure, with associated limita tions and pilfalls. is outlined very clearly in the text. The description of the effect of various tissue fixa tives on the amplifiabilily of DNA is particularly useful. However , the accompanying figure misses the mark, while filling three pages .

One of the most significant technological a d vances in molecular biology is the development of th e polymerase ch ain reaction (PCR).This procedu re can be used to d etect single copy gen es or diagnose m icrobia l infec tions in minute samples, du e lo Lh e in vi tro expon entia l a mplification of s pecifi c nu cleic acid sequen ces .The techniqu e h as b een res tricted to amplifying extracted target DNA or RNA sequ en ces.precluding correla tions b etween cytological or his tological features and molecular findings.However .by lh e pub lica tion of PCR in situ Hybridization , lhis limit.alion is overcome.This teA rt des cribes in detail a technique l h al marries surgical pathology and m olecular b iology.wi th markedly improved s ensitivity over classic in s itu hybridization .The infections cau sed by h uman pa pillomavirus and human immunodefi cien cy viru s l ar e used effec tively throughout the book lo illu s trate lh e application of a variety of m olecular techniqu es, although the a uthor em ph asizes that th e procedures are broadly a pplica ble to gen etically based, as well as other infectious, human diseases.
The a uthor s ets out to descr ibe lhe PCR in s itu technique in compreh en s ible term s fo r both lhe su rgical path ologist and the molecula r biologist.. Hence, th e first two chapters are extrem ely basic, focu s ing on lhe s tructure of DNA and defining th e 'j a rgon • associa ted with molecular biology.In Chapter 3, lhe reader is finally introduced to the conventional PCR technique.The procedure, with associated limita tions and pilfalls.is outlined very clearly in the text.The description of the effect of various tissue fixa tives on the amplifia bilily of DNA is particularly useful.However , the accompanying figure misses the mark, while filling three pages .Th e fu ndamental p roblem of nonspecific amplification of DNA is dealt with in Ch apter 4. with lhe support of excellent illustration s .Th e k ey concept of 'hot start' is introduced as a means of improving bolh lhe specificity a n d sen s itivity of conven tional PCR and is cited in later ch apters as essential lo lhe success of in situ PCR.
Ch apter 5 describes the classic in situ hyb i-idization procedu re .A detailed description of lhe technique is provided, augmented by photographs depicting lhe effects of protease treatment and tissue fiXation on hybiidization signals.Unfortunately.lhe use of black and whi le photography lo illu strate lhe appearance of differen t cou nlerslains and chr omagens is counlerproduclive.While a few colou r ph otos are provided, these are sequ estered in a later section of lhe book.The reader m u st.fli p back and forth to make comparisons.
The remainder of lhe text deals with PCR in situ hybridization or in situ PCR.In lhe later case, labelled nucleotides are incorporated d ireclly into lhe amplified product.precluding lhe hybridization step.The author emphasizes lh al lhe technique is still under development.and as s u ch th e methods he describes "are most.definitely not.lhe final story".Thus, wh il e the procedure provides insight.inlo the cellular and histological distribution of viral DNA.RNA and human mRNA, lhe utility of the technique in lhe routine diagnostic laboratory is questionable .
In summary, the strength of lhe book lies in lhe emphasis placed on lhe use of appropriate controls and in providing excellent instructions and problemsolving algorillm1s.Th e a u thor has gone lo great.length s lo build up grad ually lhe e)qJerlise ofllie reader.s u ch lha l lhe novice.as well as U1e eA'J)erl molecu lar b iologist.. will be u nda u nted by lhe technology.For a nyo n e co ntemplating using PCR in s itu hybridization in a research se lli ng.th is book is invaluable.

P McNicol PlllJ Cadham Provincial Laboratory and
The Uniuersily of Manitoba Winnipeg.Manitoba CAN j INFECT D IS VOL 5 N o 6 N OVEMBER/ D ECEMBER 1994