Polymerase chain reaction (PCR) is a laboratory technique for amplifying small quantities of deoxyribonucleic acid (DNA) into amounts large enough for detailed genetic analysis, creating a very large quantity of the originally extracted material.1 The method was conceived in April 1983 by Kary Mullis, who arrived at the idea one Friday night while driving to his cabin in the Anderson Valley, Mendocino County, California.2 Mullis is credited with the initial concept that led to PCR,3 and he was awarded the 1993 Nobel Prize in Chemistry for the technique.4 Modern forensic DNA work was made possible by PCR.5
In forensic science, PCR is used to profile DNA extracted from minimal biological material. In David Berryman’s September 22, 2002 Radio National interview on Polymerase Chain Reaction (PCR) crime-scene contamination, broadcast on Radio National (ABC), David Berryman explained that forensic DNA profiling commonly uses PCR amplification of variable regions of DNA called short tandem repeats (STRs), because forensic samples such as a single hair or the DNA in fingerprint cells contain very little material.6 Henry C. Lee said that most forensic DNA techniques are powered by PCR, a process permitting faithful reproduction of very small amounts of DNA into large amounts of identical copies.7 He said PCR allows investigators to obtain a DNA profile from postage stamps, coffee cups, and single hairs.8
The Anomaly Physical Evidence Group (APEG) directed its discussions with Bill Chalker toward using PCR amplification and sequencing of mitochondrial DNA on a hair sample from the Peter Khoury alien abduction case to test the credibility of abduction accounts.9 Preparing the samples involved a prolonged extraction step at elevated temperatures, fragmentation of the hair, and repeated boiling and freezing with dry ice to release the DNA from its protein structure before purification and PCR amplification.10 UFO Research New South Wales assisted with funding the preliminary PCR “human consensus” DNA comparisons.11 A preliminary mitochondrial DNA PCR profiling comparison against the human consensus, carried out on both Peter Khoury’s hair sample and a sample from Mike Wood’s daughter as part of the mitochondrial DNA PCR profiling comparison of Mike Wood’s daughter’s hair sample against human consensus, found both within the normal human range but different from each other.12 Vivian Khoury’s hair sample did not produce PCR amplification products, which analysts concluded might have been due to chemical treatment.13 A specialised ribosomal PCR DNA profiling was also undertaken on samples from Betty Hill’s dress, as part of Betty Hill’s 1961 dress DNA analysis, to extract non-human DNA alongside standard assessments.14 Kary Mullis did not personally carry out the PCR testing of the Khoury hair samples described in Chalker’s account.15
According to Hair of the Alien, four PCR products were obtained in total from this work: two from the tall blond alien hair and two from Peter Khoury’s own hair, but none from his wife’s hair, which may have been chemically treated and had never come into direct contact with the alien hair.16 The same source states that a 2-centimetre portion of the thin blond hair shaft, cut from just above the root, required extensive grinding to release its DNA, and that no contaminating DNA was found on any outer part of the sample.17 According to the source, PCR studies of that hair shaft, part of the hair-shaft mitochondrial DNA hypervariable region I analysis of Peter Khoury’s alien hair sample (Phase One, 1998-1999), revealed unusual mitochondrial DNA of Chinese genetics carrying a rare 16,108 C-to-T mutation, recorded as the Chinese Mongoloid mitochondrial DNA substitution found in Peter Khoury’s alien hair shaft.18 According to the same source, both sides of the mitochondrial hypervariable region I were amplified by PCR for 37 or 33 cycles respectively, while a nuclear region coding for the CCR5 gene was amplified for 40 cycles.19 A later stage of testing, the Phase Two hair-root DNA analysis of Peter Khoury’s alien hair sample revealing Basque/Gaelic mitochondrial DNA, found that two different samples from the hair root gave two different mitochondrial DNA sequences within the same PCR run: a 16255C genotype for six of seven clones from the soft root, and a 16304 sequence from the hard keratin at the base of the shaft, matching the shaft result from the first round of testing.20
David Berryman has drawn attention to the potential for deliberate contamination of crime scenes with PCR products.21 In his Radio National interview, Berryman said PCR has contamination issues in the laboratory and that a crime scene can be deliberately contaminated using PCR products as the diagnostic test.22 He said this does not require planting hairs or a fingerprint, but can be done by covering the whole scene with DNA PCR products.23 He said it requires only easy access to a publicly available PCR kit and amplifier, comparable in difficulty to buying a litre of milk at a supermarket.24 He said the PCR reaction itself requires no special skill or significant resources and can be performed at home on a stove using publicly available information.25 He said that, based on his data, squirting PCR products around a crime scene is enough to contaminate it virtually beyond any possibility of recovering a genuine DNA profile.26 He was concerned that while considerable effort had gone into resolving PCR-related contamination issues in laboratory settings, the impact of deliberate PCR contamination in the field, at crime scenes, seemed largely ignored.27 Bill Chalker said he considered it certain that Peter Khoury had no knowledge, in 1992, of the forensic PCR contamination techniques only later discussed in the field.28 Chalker also said he considered it very difficult, though not impossible, to fabricate a PCR “brew” containing the same rare genetic substitutions found in the Khoury hair sample, given the rarity of the profile and its unusual attributes near the root and at the shaft.29 Debate continues about the factors that might affect the accuracy of standard PCR-based DNA profiling.30
PCR also featured in the investigation of the 2000-09 Gary Lowrey alien claw discovery. According to Hair of the Alien, six separate rounds of DNA analysis, using different and sometimes novel approaches, were carried out on the claw sample, including a new PCR reaction using novel primers targeting the most conserved DNA sequences on Earth, in order to resolve the case.31 The source notes that molluscs, particularly snails and slugs, secrete a thick mucus containing inhibitors of many enzymes fundamental to molecular biology, including PCR enzymes.32 It states that rigorous DNA analysis using a novel set of oligonucleotide primers for PCR ultimately resolved the identity of the claw sample.33