Stephen Bustin

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Stephen Andrew Bustin (born 1954) [1] is a British scientist, former professor of molecular sciences at Queen Mary University of London from 2004 to 2012, as well as visiting professor at Middlesex University, beginning in 2006. [2] [3] In 2012 he was appointed Professor of Allied Health and Medicine at Anglia Ruskin University. [4] He is known for his research into polymerase chain reaction, and has written a book on the topic, entitled A-Z of Quantitative PCR. This book has been called "the bible of qPCR." [5]

Contents

Education

Bustin obtained his B.A. and PhD from Trinity College, Dublin in molecular genetics. [6]

Career

In 2023, he was elected a member of the Academia Europaea. [7]

Research

His research group’s general areas of interest are the small and large bowel, as well as colorectal cancer with particular emphasis on investigating the process of invasion and metastasis. An important aim is to translate molecular techniques into clinical practice by including molecular parameters into clinical tumor staging. [8] To this end, Bustin has published many papers on PCR techniques, [9] [10] in particular reverse transcription polymerase chain reaction, the subject of his most cited paper, published in 2000. [11]

He also developed the MIQE guidelines in a 2009 paper published in Clinical Chemistry , [12] the goal of which is to create guidelines for how PCR should be performed to ensure that PCR results are being reliably conducted and interpreted, as well as to make replication of experiments easier. This paper is the fifth most cited one ever to be published in Clinical Chemistry, [13] [14] with over 1700 citations on Google Scholar as of September 2013. [15]

Testimony

Autism omnibus trial

Bustin testified on behalf of the Department of Justice in the autism omnibus trial about what he stated was the unreliability of the O'Leary lab's results with regard to testing for contamination. The lab had claimed to find measles virus in the intestines of children with developmental disorders. [16] Bustin describes his conclusions with regard to the lab's alleged detection of measles virus RNA as follows: "My clear conclusion then was that O'Leary's results were caused by defective experimental technique and inappropriate interpretation of results, since he was detecting DNA, and measles virus does not exist as DNA." [17] [18] Bustin was described as "one of the most highly qualified and credible expert witnesses I [the Special Master] have ever encountered." [4] In addition to his testimony, Bustin published an analysis of Andrew Wakefield's 2002 study, which had been published in the journal Molecular Pathology . This analysis, like Bustin's testimony, concluded that "The only conclusion possible is that the assays were detecting contaminating DNA. Since MeV is an RNA-only virus and never exists in DNA form, these data must be ignored and it is my opinion that the authors should withdraw this publication from the peer-reviewed literature." [19]

Lundy murders

Bustin testified in the trials pertaining to the Lundy murders in 2015, criticizing tests that had claimed to detect human brain cells on Mark Lundy's shirt. [20] [21]

Related Research Articles

<span class="mw-page-title-main">Complementary DNA</span> DNA reverse transcribed from RNA

In genetics, complementary DNA (cDNA) is DNA that was reverse transcribed from an RNA. cDNA exists in both single-stranded and double-stranded forms and in both natural and engineered forms.

<span class="mw-page-title-main">Polymerase chain reaction</span> Laboratory technique to multiply a DNA sample for study

The polymerase chain reaction (PCR) is a method widely used to make millions to billions of copies of a specific DNA sample rapidly, allowing scientists to amplify a very small sample of DNA sufficiently to enable detailed study. PCR was invented in 1983 by American biochemist Kary Mullis at Cetus Corporation. Mullis and biochemist Michael Smith, who had developed other essential ways of manipulating DNA, were jointly awarded the Nobel Prize in Chemistry in 1993.

<span class="mw-page-title-main">Reverse transcriptase</span> Enzyme which generates DNA

A reverse transcriptase (RT) is an enzyme used to convert RNA genome to DNA, a process termed reverse transcription. Reverse transcriptases are used by viruses such as HIV and hepatitis B to replicate their genomes, by retrotransposon mobile genetic elements to proliferate within the host genome, and by eukaryotic cells to extend the telomeres at the ends of their linear chromosomes. Contrary to a widely held belief, the process does not violate the flows of genetic information as described by the classical central dogma, as transfers of information from RNA to DNA are explicitly held possible.

<span class="mw-page-title-main">Gene expression</span> Conversion of a genes sequence into a mature gene product or products

Gene expression is the process by which information from a gene is used in the synthesis of a functional gene product that enables it to produce end products, proteins or non-coding RNA, and ultimately affect a phenotype. These products are often proteins, but in non-protein-coding genes such as transfer RNA (tRNA) and small nuclear RNA (snRNA), the product is a functional non-coding RNA. The process of gene expression is used by all known life—eukaryotes, prokaryotes, and utilized by viruses—to generate the macromolecular machinery for life.

<span class="mw-page-title-main">Transcription (biology)</span> Process of copying a segment of DNA into RNA

Transcription is the process of copying a segment of DNA into RNA. Some segments of DNA are transcribed into RNA molecules that can encode proteins, called messenger RNA (mRNA). Other segments of DNA are transcribed into RNA molecules called non-coding RNAs (ncRNAs).

<span class="mw-page-title-main">RNA polymerase</span> Enzyme that synthesizes RNA from DNA

In molecular biology, RNA polymerase, or more specifically DNA-directed/dependent RNA polymerase (DdRP), is an enzyme that catalyzes the chemical reactions that synthesize RNA from a DNA template.

Viral load, also known as viral burden, is a numerical expression of the quantity of virus in a given volume of fluid, including biological and environmental specimens. It is not to be confused with viral titre or viral titer, which depends on the assay. When an assay for measuring the infective virus particle is done, viral titre often refers to the concentration of infectious viral particles, which is different from the total viral particles. Viral load is measured using body fluids sputum and blood plasma. As an example of environmental specimens, the viral load of norovirus can be determined from run-off water on garden produce. Norovirus has not only prolonged viral shedding and has the ability to survive in the environment but a minuscule infectious dose is required to produce infection in humans: less than 100 viral particles.

<span class="mw-page-title-main">Reverse transcription polymerase chain reaction</span> Laboratory technique to multiply an RNA sample for study

Reverse transcription polymerase chain reaction (RT-PCR) is a laboratory technique combining reverse transcription of RNA into DNA and amplification of specific DNA targets using polymerase chain reaction (PCR). It is primarily used to measure the amount of a specific RNA. This is achieved by monitoring the amplification reaction using fluorescence, a technique called real-time PCR or quantitative PCR (qPCR). Confusion can arise because some authors use the acronym RT-PCR to denote real-time PCR. In this article, RT-PCR will denote Reverse Transcription PCR. Combined RT-PCR and qPCR are routinely used for analysis of gene expression and quantification of viral RNA in research and clinical settings.

<span class="mw-page-title-main">Real-time polymerase chain reaction</span> Laboratory technique of molecular biology

A real-time polymerase chain reaction is a laboratory technique of molecular biology based on the polymerase chain reaction (PCR). It monitors the amplification of a targeted DNA molecule during the PCR, not at its end, as in conventional PCR. Real-time PCR can be used quantitatively and semi-quantitatively.

TaqMan probes are hydrolysis probes that are designed to increase the specificity of quantitative PCR. The method was first reported in 1991 by researcher Kary Mullis at Cetus Corporation, and the technology was subsequently developed by Hoffmann-La Roche for diagnostic assays and by Applied Biosystems for research applications.

Nucleic acid sequence-based amplification, commonly referred to as NASBA, is a method in molecular biology which is used to produce multiple copies of single stranded RNA. NASBA is a two-step process that takes RNA and anneals specially designed primers, then utilizes an enzyme cocktail to amplify it.

Idaeovirus is a genus of positive-sense ssRNA viruses that contains two species: Raspberry bushy dwarf virus (RBDV) and Privet idaeovirus. RBDV has two host-dependent clades: one for raspberries; the other for grapevines. Infections are a significant agricultural burden, resulting in decreased yield and quality of crops. RBDV has a synergistic relation with Raspberry leaf mottle virus, with co-infection greatly amplifying the concentration of virions in infected plants. The virus is transmitted via pollination with RBDV-infected pollen grains that first infect the stigma before causing systemic infection.

<span class="mw-page-title-main">Nucleic acid test</span> Group of techniques to detect a particular nucleic acid sequence

A nucleic acid test (NAT) is a technique used to detect a particular nucleic acid sequence and thus usually to detect and identify a particular species or subspecies of organism, often a virus or bacterium that acts as a pathogen in blood, tissue, urine, etc. NATs differ from other tests in that they detect genetic materials rather than antigens or antibodies. Detection of genetic materials allows an early diagnosis of a disease because the detection of antigens and/or antibodies requires time for them to start appearing in the bloodstream. Since the amount of a certain genetic material is usually very small, many NATs include a step that amplifies the genetic material—that is, makes many copies of it. Such NATs are called nucleic acid amplification tests (NAATs). There are several ways of amplification, including polymerase chain reaction (PCR), strand displacement assay (SDA), transcription mediated assay (TMA), and loop-mediated isothermal amplification (LAMP).

The FluChip is a low-density DNA microarray for the identification of influenza viruses, originally developed at the University of Colorado at Boulder in the laboratory of Professor Kathy Rowlen in collaboration with the Centers for Disease Control and Prevention (CDC) in Atlanta.

Cell-free fetal DNA (cffDNA) is fetal DNA that circulates freely in the maternal blood. Maternal blood is sampled by venipuncture. Analysis of cffDNA is a method of non-invasive prenatal diagnosis frequently ordered for pregnant women of advanced maternal age. Two hours after delivery, cffDNA is no longer detectable in maternal blood.

<i>Cedillo v. Secretary of Health and Human Services</i> Legal case in US Court of Federal Claims, decided in 2009

Michelle Cedillo v. Secretary of Health and Human Services, also known as Cedillo, was a court case involving the family of Michelle Cedillo, an autistic girl whose parents sued the United States government because they believed that her autism was caused by her receipt of both the measles-mumps-and-rubella vaccine and thimerosal-containing vaccines. The case was a part of the Omnibus Autism Proceeding, where petitioners were required to present three test cases for each proposed mechanism by which vaccines had, according to them, caused their children's autism; Cedillo was the first such case for the MMR-and-thimerosal hypothesis.

Recombinase polymerase amplification (RPA) is a single tube, isothermal alternative to the polymerase chain reaction (PCR). By adding a reverse transcriptase enzyme to an RPA reaction, it can detect RNA as well as DNA, without the need for a separate step to produce cDNA. Because it is isothermal, RPA can use much simpler equipment than PCR, which requires a thermal cycler. Operating best at temperatures of 37–42 °C and still working, albeit more slowly, at room temperature means RPA reactions can in theory be run quickly by simply holding a tube in the hand. This makes RPA an excellent candidate for developing low-cost, rapid, point-of-care molecular tests. An international quality assessment of molecular detection of Rift Valley fever virus performed as well as the best RT-PCR tests, detecting less concentrated samples missed by some PCR tests and an RT-LAMP test. RPA was developed and launched by TwistDx Ltd., a biotechnology company based in Cambridge, UK.

<span class="mw-page-title-main">Molecular diagnostics</span> Collection of techniques used to analyze biological markers in the genome and proteome

Molecular diagnostics is a collection of techniques used to analyze biological markers in the genome and proteome, and how their cells express their genes as proteins, applying molecular biology to medical testing. In medicine the technique is used to diagnose and monitor disease, detect risk, and decide which therapies will work best for individual patients, and in agricultural biosecurity similarly to monitor crop- and livestock disease, estimate risk, and decide what quarantine measures must be taken.

<span class="mw-page-title-main">Reverse Transcription Loop-mediated Isothermal Amplification</span>

Reverse transcription loop-mediated isothermal amplification (RT-LAMP) is a one step nucleic acid amplification method to multiply specific sequences of RNA. It is used to diagnose infectious disease caused by RNA viruses.

The Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines are a set of protocols for conducting and reporting quantitative real-time PCR experiments and data, as devised by Bustin et al. in 2009. They were devised after a paper was published in 2002 that claimed to detect measles virus in children with autism through the use of RT-qPCR, but the results proved to be completely unreproducible by other scientists. The authors themselves also did not try to reproduce them and the raw data was found to have a large amount of errors and basic mistakes in analysis. This incident prompted Stephen Bustin to create the MIQE guidelines to provide a baseline level of quality for qPCR data published in scientific literature.

References

  1. ↑ "Stephen Bustin". ResearchGate . Retrieved 24 September 2013.
  2. ↑ "Stephen Bustin's Biography". Selectbiosciences. 2011. Retrieved 12 August 2013.
  3. ↑ McGee, Patrick (10 May 2007). "How Reliable is Your qPCR Data?". Drug Discovery & Development. Retrieved 12 December 2013.
  4. 1 2 Bustin, Stephen (2013). "Definitive qPCR" . Retrieved 12 August 2013.
  5. ↑ Perkel, Jeffrey M. (1 December 2013). "PCR: Past, Present, & Future". The Scientist . Retrieved 21 February 2014.
  6. ↑ "Stephen Bustin". Anglia Ruskin University. Retrieved 30 September 2016.
  7. ↑ "Stephen Bustin". Member. Academia Europaea. Retrieved 2 November 2024.
  8. ↑ "The future of qPCR". American Association for the Advancement of Science. 30 September 2010. Retrieved 11 August 2013.
  9. ↑ Nolan, T.; Hands, R. E.; Bustin, S. A. (2006). "Quantification of mRNA using real-time RT-PCR". Nature Protocols. 1 (3): 1559–1582. doi:10.1038/nprot.2006.236. PMID   17406449. S2CID   10108148.
  10. ↑ Mueller, R.; Bustin, S. A. (2005). "Real-time reverse transcription PCR (qRT-PCR) and its potential use in clinical diagnosis". Clinical Science. 109 (4): 365–379. CiteSeerX   10.1.1.328.8607 . doi:10.1042/CS20050086. PMID   16171460.
  11. ↑ Bustin, S. (2000). "Absolute quantification of mRNA using real-time reverse transcription polymerase chain reaction assays". Journal of Molecular Endocrinology. 25 (2): 169–193. doi: 10.1677/jme.0.0250169 . PMID   11013345.
  12. ↑ Bustin, S. A.; Benes, V.; Garson, J. A.; Hellemans, J.; Huggett, J.; Kubista, M.; Mueller, R.; Nolan, T.; Pfaffl, M. W.; Shipley, G. L.; Vandesompele, J.; Wittwer, C. T. (2009). "The MIQE Guidelines: Minimum Information for Publication of Quantitative Real-Time PCR Experiments". Clinical Chemistry. 55 (4): 611–622. doi: 10.1373/clinchem.2008.112797 . PMID   19246619.
  13. ↑ "The MIQE Guidelines" . Retrieved 11 August 2013.
  14. ↑ "Most Cited Articles". Clinical Chemistry . Retrieved 9 September 2013.
  15. ↑ "The MIQE Guidelines: M inimum I nformation for Publication of Q uantitative Real-Time PCR e xperiments".
  16. ↑ Marx, Vivien (2013). "PCR: living life amplified and standardized". Nature . 10 (5): 391–395. doi: 10.1038/nmeth.2443 . PMID   23629414. S2CID   8605063.
  17. ↑ Bustin, Stephen (8 December 2008). "Fading Claims of MMR Link to Autism". The Guardian . Retrieved 11 August 2013.
  18. ↑ Fitzpatrick, Michael (4 July 2007). "'The MMR-autism theory? There's nothing in it'". Spiked Online . Retrieved 12 December 2013.
  19. ↑ Feinstein, Adam (2010). A History of Autism: Conversations with the Pioneers. Blackwell Publishing. p. 227.
  20. ↑ Quilliam, Rebecca (11 March 2015). "Mark Lundy retrial: DNA on shirt matched to victim". New Zealand Herald . Retrieved 30 August 2015.
  21. ↑ Galuszka, Jono (9 March 2015). "Mark Lundy murder retrial: Day 20". Stuff.co.nz . Retrieved 30 August 2015.
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