Mahkamah Tinggi Jenayah 3 KL
Di hadapan Yang Arif Dato’ Mohamad Zabidin Mohd Diah
Pihak-pihak:-
PP : Semua hadir
PB : SN, Ram Karpal (KS, Datuk Param Cumaraswamy, Dato’ CV Prabhakaran, Marissa, Radzlan tidak hadir)
WB : Zamri Idrus (for Complainant)
Expert for defence: Prof. David Wells, Dr. Brian McDonalds
AI hadir
[9.01 a.m.]
MY: Hari ini ditetapkan untuk menyambung pemeriksaan balas SP5. Izinkan kami untuk memanggil saksi.
Q: Dengan izin YA. We stop yesterday at the chart of electropherogram, can we start today by you explaining what is electropherogram?
A: An electropherogram is a graphical presentation of the DNA results after analysis which means the separation by electrophoresis. Electropherogram is from the word electrophoresis, its undergone electrophoresis during
separation. And the results which had been presented by the electropherogram and for this particular case, electropherogram is represented in four colors which used to detect the DNA fragment. And the name of those all
respective name loci are at top and the DNA type are reflected by peaks.
Q: This electropherogram, how does it operate in your laboratory? Is it a system or, a software?
A: It is a combination of a system which separate and detect the DNA fragment and there is a software called Gene Mapper which is stored in genetic analyzer and that converts the signals into digitize forms and the Gene
Mapper uses a leather, a DNA fragment of non-sizes, called an allele- leather. It acts as measuring tape, where the sizes of DNA fragment of DNA types are being size and the results are being depicted in the peaks form. It was
presented together with this report.
Q: In other words, is it true if the allele that you got in your Appendix., you obtained from the reading of the electropherogram?
A: Exactly.
Q: In this case, have you printed out the electropherogram of the results that you have done; the DNA analysis?
A: Yes. This is required to interpret the DNA results.
NB: At this juncture, YA, may I tender set of the electropherogram graphs, that being identified first by the witness which consists of 36 pages.
Dr. Seah, is this electropherogram chart that involves in this case?
A: Yes.
Q: And this is pertaining to this case?
A: Yes.
Q: How did you able to identify that this belongs to the particular case and it involves the sample that you have analyze in this case?
A: That’s the wrong sheet before that one, and there is laboratory number which is the unique identifying number: 6334/08 is typed into the wrong sheet and the exhibit number is typed in as well, the exhibit number will be at
the top panel under the name sample file.
Q: The first page of the electropherogram would be which sample of the exhibits?
A: This is the electropherogram DNA profile from swab B.
Q: From the second page?
A: From swab B1.
Q: Third page?
A: From B2. For B2- No result and that’s why the peaks are not detected at all.
Q: Page 4?
A: of the DNA profile from swab B3.
Q: There is a word re-amp there. What does this mean?
A: Re amplifications. This is the second implication, where the first amplification of a complete profile was not successfully obtained. And it was later being assessed for the cause of the Incomplete DNA profile, and the
assessments may indicate that there’s insufficient DNA template, in which case probably more DNA template were being used, or the assessment may indicate inhibitions, in which case procedure and steps will be taken to
overcome the [] either by cleaning up the DNA extract or by method like [] the DNA template which will overcome the inhibitions.
Q: And this is as what that you have explained as with regards to PCR technique that you have adopted?
A: No, we will go through the same technique, the same PCR. But at this time, we want to understand what was problem that caused an incomplete DNA process.
Q: So re-amplifications is a normal process?
A: It is a normal process. It is to correct whatever incompleteness of the first one. Not all of the PCR will be successful at the first amplifications, and there are problems which can only be seen after the re amplifications, and
most of the problems are then recognized and being rectified. Re-amplification is then carried out to correct those problems, and usually result in the most successful DNA profile.
Q: We move on to page 5?
A: DNA profile from swab B4.
Q: And it involves re-amplifications as well?
A: Yes.
Q: Page 6?
A: Electropherogram from swab B5 because of the detection of the presence of semens of this swab, differential extraction will be fall into two fractions, the non-sperm fraction, sometimes called the female fractions and the
sperm fraction which is reflected by B5 (small M).
Q: Where is that? Page 7?
A: Yes. Therefore B5, there would be two amplifications; one was for the sperm extract and one for non-sperm extract.
Q: Page 6 would be for non-sperm extract? And Page 7 will be for sperm extract?
A: Yes that’s correct.
Q: So there are 2 pages involving sample B5, which are page 6 and 7. We move on to page 8.
A: For page 8, again this is the electropherogram of the DNA profile from swab B5. This has no DNA profiles, no peaks in this electropherogram.
Q: Page 9, which sample is this?
A: This is the electropherogram for the swab B6 and swab B6 differential extraction was carried out despite the non detecting the presence of semens. And the female fraction or non-sperm fraction has a complete profile, and
the next page, page 10, there is no DNA from the sperm extract, there are no peaks.
Q: Page 11?
A: From swab B7 and B7, differential extraction is carried out.
Q: Page 12
A: B7 (f) for the non sperm extract.
Q: B7 (m)?
A: Again, B7 (m) is an incomplete profile, we assessed it, and we recognize the problem of inhibitions and DNA extract was being cleaned up to remove the inhibitors, and second amplification is carried out and depicted at
the word re-amp. This is at page 12.
Q: Page 13?
A: 13 is the continuity of the electropherogram from page 12, which is still the sperm extract from the swab B7. Page 14, is the DNA profile from swab B8. Page 15- sperm extract which is B8 (m). B8 (m) was in two pages.
Page 17 – the DNA profile from swab B9. Page 18 and 19- sperm extract which is B9 (m).
Q: Move on.
A: Page 20, is the electropherogram for the blood stain specimen, B10.
Q: Page 21?
A: We have printed out one of this allelic leather.
Q: What is allelic leather?
A: This is the leather contains of DNA fragment of a non sizes and their function is to size the DNA from the test samples and it is acts basically like measuring tape, so it would be able to size the DNA from the test
samples, and the software would access the separation of the allelic leather to ensure that the separation is satisfactory, is reliable, and when the software has access that leather will performing successfully, which the signals is
sufficiently strong to be used as allele leather or as a weapon. It then allows the allelic leather to size the test sample.
Q: Page 22?
A: This separation was based on chemistry, it uses the 4 colors for the test of DNA fragments and that various colors are also adopted by the leather. Mine is in black and white, but for those with colors electropherogram,
there are green, yellow (yellow is depicted in black), blue and the red. These are the 4 types which used during PCR, which allows DNA fragment to be detected in the genetic analyzer. This is the technology that is being used.
There’s a leather design for each locus. If you look at the allelic leather again there’s a leather range which being analyzed for the test sample.
Q: Page 23?
A: It is so called negative control. This is the quality control that is being used. A negative control means, during PCR, another tube of negative control is being set up where instead of putting in a DNA, equal volume of the
template is being used without any DNA which is usual [] water and that will ensure that there is no amplifiable DNA in the PCA mixture. There is no contamination coming out from the DNA mixture. The negative control ensures
that there ought not to be any peaks coming from the negative control. If there is peak coming out, there’s mean there might be a contamination presence, and therefore the [] remaining test sample will not being read, it will be
abandoned.
Q: In other word, the negative control is there to make sure there was no contamination from the DNA mixed?
A: Yes
Q: Next page?
A: There is another control called the positive control. And if it detected any difference on it, again, the test result will be abandon. This contains DNA of DNA types. This usually come with PCR [] so called positive control
where the sizing and the genotypes at each of respective loci are already known. Then on page 25, there is another control which call the reagent blank.
Reagent blank is carried out simultaneously with test sample from the very first step. It was carried out from the exactly same step that the test sample went through except there would be no DNA sample, no DNA extract. There is
to make sure that there is no DNA introduce from any of the reagent of chemical or baffles, right from the start of the very first step. Again the reagent blank should have be no peaks, no DNA profile. If there is any, even from a very
minor/small peak, this means that your result cannot be accepted. You have to trace where the contamination DNA come from. It means you have to disposed/throw, to assess where the origin of the contamination came from. And
when that is traced, all the test samples result will be abandon, and it has to be done all over again. This is very particular quality control step which is to enforce by the accreditation audit, that if there is at any time peak presents in
reagent blank, in fact that whole result from the reagent bottle during that period of time would have to be abandon.
Q: Page 26?
A: DNA profiles from the seminal stains from trousers A3. There are 2 spots were used. One is the A3 (a) and one is A3 (b) and. Two sports were analyzed. Page 27 is non sperm extract from spot A3 (a). And the next page,
page 28 is the DNA profile from the sperm extract from A3 (a) and then on page 29 that for the non sperm exact from A3(b) and on page 30, DNA profiles from spot A3 (b).
Q: Move on?
A: Page 31, again for the seminal stain from underwear A6, different extraction is carried out for both spots A6 (a) and A6 (b) and page 32 is the DNA profile from the sperm extract A6 (a).
Q: Page 33?
A: From DNA profile of swab A6 (b) – non sperm extract. That’s on two pages: page 33, and 34.
Q: Page 35?
A: From spots A6 (b) from the sperm extract.
Q: Last page 36?
A: DNA profile from hair A. there is no peaks which means no DNA profiles from hair A.
NB: If there is no objection, may this electropherogram to be tendered and marked as P52.
YA: Yes, council?
KS: The marking of it can be different until cross-examine. There want to go [] lay the ground, perhaps at that point, YA.
KS & MY berbincang.
YA: Now, if you want to object on the ground of admissibility you have to settle it now.
KS: I think we need to lay the ground of the objection, but we cannot do it at this stage.
YA: Now, mark it first. Later on, if you have the ground, then you can object lah.
KS: We cannot lay the ground on the stage on EIC. It is better that we lay it down later.
MY: I understand the concern of KS. In fact we have prepared the authority. My learned friend is talking about the absence of certificate. We are giving it through oral evidence. Sec 90A talks about [read].
YA: So you have agreed that until and unless I just throw the certificate, or I just make it through the oral evidence that the computer is in a good working condition, it cannot be mark as exhibits?
MY: If I may refer to Section 136 of EA, where it can prove this. It is the matter of calling the evidence, in fact we give it through her, or give both to the court. In fact, I asked Puan Noorin if she wants to ask after this.
YA: Continue.
Q: This chart again, how does it produced? Produce from computer?
A: The whole genetic analyze system is a system of a very sophisticated separation system, which is the capillary electrophoresis system, lesser detection and it comes with that software which tides the DNA after the
machines had detected it and this software is called Gene Mapper.
Q: So all this information that you have gathered will be stored in your software?
A: Yes, it is stored in the electronic files.
Q: So, this document now produced by a computer.
A: Yes.
Q: At that time when you using that computer, as well as when you printed out the electropherogram, was the computer in a good working condition or good working order?
A: Yes it was.
Q: Was the computer functioning in the course of it daily use?
A: Yes.
Q: If something wrong with the computer or the software of that matter, would you be able to detect it?
A: Yes, not from the computer that the software will indicate if the conversion or the separation is unsatisfactory. The software is actually a part of the system. It is not a separate software that you should install it to re-do
whatever result that you got from the genetic analyzer.
Q: Did you detect anything wrong on that day?
A: No.
NB: At this juncture, if I may, tender and mark this electropherogram as exhibit.
KS: We have objecting to this. This is the computer generated document; certain requirements need to be fulfilled. Is my learned friend want to conduct trial within trial for the purpose of admissibility? This is not necessary at
this point. We can’t assume that everything is in order because she said it. We haven’t cross examined her yet. Unless, trial within trial, perhaps we should now, go into that stage.
MY: The basis of this….put on us, based on this few authority, the latest is the case of Ahmad Najib bin Aris n PP Fed Court decision, 2009 2 MLJ 73. YA, we anticipated all this thing. Sec 90A of the Evidence Act, what
Ahmad Najib said, you refer to Ahmad Hanafi tab 5. [read]. It will be funny that a certificate issued by somebody subject to no cross – examination, the documents which are produced by a computer which is in good working order
is admissible. And yet, if we have the oral evidence from the particular person, choice of evidence of the generated documents, it is safe to say that it is admissible. At this juncture, admissibility is not the issue. Admissibility we
have complied all the requirements. Just for reference, YA, Ahmad Hanafi’s case is para 31 and 32 of the judgment, tab 5. If I just may read this case [read]. At page 155 at para b. Firstly, in good working order, and secondly, it
was operating properly throughout the material part that the documents was being produced. That was all we need to adduce.
KS: Subject to it been reviewed in cross-examination.
YA: If you can establish that the court is not working properly in cross examination, you can ask the court to expunge it.
KS: Very well.
NB: Marked and tendered as P52. Much obliged.
Q: What was the equipment used apart from the electropherogram in conducting your DNA analysis?
A: Standard equipment which was used by the international DNA Laboratory, [], in carrying out the PCR. We have also the [] which to quantify the DNA.
Q: And were the equipment used operating properly in all aspect throughout your analysis?
A: Yes, internal checks were carried out before each and every run.
Q: Are they calliberated ?
A: The maintenance is run periodically, by the train engineer.
Q: when you were not conducting it, where was the item kept?
A: The exhibits?
Q: Yes, the exhibits.
A: The exhibits were kept back in the freezer and sealed. We might initial again. Each and every time the exhibits kept in the freezer, the same procedure will be done to ensure there will be no contamination, or being
accessed by any other unauthorized personnel.
Q: And have you ever left the exhibits unattended
A: No, I have not left it unattended at any time.
Q: After complete your analysis; what did you do after do the analysis?
A: Sealed by the Jabatan Kimia Malaysia, security lable.
Q: When did you return the exhibits to the IO, DSP Jude?
A: The exhibits together with the report were returned to DSP Jude Blacious at approximately 11.30 am on 7th of July 2008.
Q: Was there a handing over document without DSP Jude on that day?
A: This is documented at page 4 of the report.
Q: You handed over the exhibits together with your report?
A: Yes.
Q: If I may refer, where it states the handing over between you to DSP Juse.
A: Page 4, last paragraph.
Q: You signed on that page it as well?
A: Yes. He signed it as well. Signature of acknowledgements of us were there.
Q: What was the condition on the seals of the exhibits when you handed it over to DSP Jude.
A: The seals were intact, the envelopes and packages were in good condition.
Q: Did you put in any package or packet?
A: In a carrier bag, in Jabatan Kimia Malaysia’s carrier bag.
Q: Is this the plastic bag that you have given to Jude?
A: Yes. This is the bag which was sealed to store the exhibits.
Q: Can we have it mark, as P 53?
Q: What have you put on this plastic bag?
A: The laboratory number. PJ FOR 6334/08-0-2.
NB: That would be all for my EIC. Much obliged Yang Arif.
CROSS – EXAMINATION OF SP5
RK: YA, we want to start cross but as we informed yesterday, we were told that we can view the exhibits, and mark the marking and so on. We’ll need to do that as part of our cross. I wonder if YA would want us to do now
before we start cross.
YA: Can you start first without referring to it, and then you can view the exhibits later.
Q: You informed the court that the document that you referred to was electropherogram. Those were produced by a machine. What was the machine?
A: The genetic analyzer.
Q: What brand?
A: It is a applied bio-system machine, model 3130, SL
Q: This electropherogram is produced by this machine. Do you agree that this graphs is essential to your computer?
A: Yes.
Q: What are the documents, can you inform us, that form the basis of your conclusion, apart from these graphs? Can you itemize it, one by one?
A: First would be the test carrying out form the presence of the seminal stains.
Q: No, I’m talking about the documents. Are there any other documents that form the basis and essential to your conclusion?
A: No, these are the documents which draw the basis of the results.
Q: What software is used for this machine?
A: Gene Mapper.
Q: Who installed this software?
A: By [] Bio System, by their engineers.
Q: Not installed by you?
A: No
Q: Do you assume that this software is patented?
A: No, this is not assumption.
Q: Do you have the personal knowledge on how the software works?
A: No
Q: The end result of what had been produced by the machine, the graphs is the result and product of this software?
A: Yes
Q: Does the software come with a manual?
A: Yes
Q: Does the manual tell us how to operate the software?
A: Yes.
Q: I assume the manual will come with the machine itself, is that correct?
A: Yes.
Q: Do you have any engineer in your department, in Jabatan Kimia Petaling Jaya in charge of handling this software in the event of any irregularities and so on?
A: They are not engineers; they are the analysts that are trained.
Q: Technician?
A: Yes.
Q: They are the people in charge in running of this software?
A: That’s correct.
Q: I put it to you that you have no involvement in the operating of the software. Agree?
A: Not exactly.
Q: You have no involvement in installing the software. Agree?
A: That’s correct
Q: I take you to your EIC earlier. Correct me if I’m wrong, but my notes said that the information received from the graphs stored in electronic files.
A: That’s correct.
Q: The computer is functioning in ordinary cause. Would it be correct that you said that the computer is functioning in its ordinary cause because you believed that the software is running properly?
A: The software is in fact running properly. It is not my belief.
Q: You are not a person in charge in this software are you? Are you the expert in the setting up of the software?
A: No I’m not.
Q: So is the running of the software within the ambit of your expertise?
A: No.
Q: You have no technical no how about this running of this software?
A: That’s correct.
Q: So you assume that it was running well?
A: I’m not assuming.
Q: You said that there is another group of technician who handled the running of the software, dealing with irregularities and so on. So you are just assuming that it was running properly?
A: I disagree.
Q: The graphs, 36 pages in all? Can you just confirm how many pages in total?
A: 36 pages
Q: And it was came out, produced one by one?
A: Yes.
Q: Did you see it printed out one by one?
A: Yes.
Q: Was there anybody else at that time
A: Yes.
Q: Who were they?
A: Yes, my fellow colleague who assisted me, Edriany.
Q: Is this Edriany a chemist as well?
A: She is a scientific officer.
Q: So at no time before you started analysis in this case, did you question the regularity of this software? Did you at all at this exercise question it?
A: No
Q: Can I just in general term referred you to your report P25. Your report is accompanied by the STR result. Three attachments in your Appendix 1?
A: Yes.
Q: How was this appendix produced?
A: This went out from the electropherogram.
Q: This appendix is also part of the machine?
A: Yes.
Q: What is STR stands for?
A: It stands for short tendem repeats.
Q: You testified yesterday that there were certain alleles that belonging to Male Y, the complainant and also the unknown contributor.
A: That’s wrong. That’s only for swab B5. It’s not all way in summary.
Q: Swab A3, look at the 1st appendix, just for an example, look at locus D8S1179, you have conducted the examination of seminal stain from trousers A3(a) and (b). You found the presence of alleles 12, 13?
A: Yes
Q: Those alleles, 12 and 13, where did you get the information from?
A: This extracted from the data, from the electropherogram.
Q: Those alleles come from the reference sample of the complainant?
A: It cannot be assessed just on one locus.
Q: I’m asking you as an example first. 12, 13 come from the analysis from the reference sample of complainant, is that correct?
A: I think the question is out of context.
Q: Was that reference sample given by the complainant?
A: Yes.
Q: What was that reference sample?
A: That was B10, blood stain specimen.
Q: So what was done to that blood stain specimen?
A: Extraction of the blood specimen for the DNA profiling.
Q: The purpose of that DNA profiling is to determine the DNA contributor isn’t it?
A: Yes
Q: We come to the conclusion of who the contributor is by the allele?
A: Yes by the comparison of the entire DNA profiles.
Q: What was the complainant profile as the result of your analysis?
A: It is documented in page iii of appendix 1, laSt column, B10.
Q: Go to first locus, the complainant has 12,13 genotypes, right?.
A: Yes.
Q: I take you to page 3 of your report, para iv. [read] – ‘concordant with being contributed by the donor of blood stain B10 AND Male Y’. What does it means?
A: Concordant means that DNA types match with being contributed.
Q: in your evidence yesterday, you also used the word complainant’s alleles a number of times?
A: I don’t use the word complainant’s alleles.
Q: DNA matches what exactly?
A: B5. It matches with being contributed by the blood stain specimen’s donor.
Q: Have you heard of the term not excluded?
A: Yes
Q: That is commonly used when referring a DNA profile
A: When a firm conclusion cannot be form.
Q: What does it means to your mind?
A: You can’t exclude, and neither can you include conclusively.
Q: What is the difference between the term not excluded and the term you used here, concordant?
A: I used not excluded if it matches only a small number of loci, probably less than 10 where I cannot conclusively said that he is the contributor, and yet I cannot excluded him as a contributor so I used the term not
included.
Q: In order to jump to the conclusion, you have to carry out further test.
A: No, it comes from assessment.
Q: Purely on that, no further test?
A: No, that’s the ultimate test result.
Q: Would it be easy and usual to provide the statistical report to support your conclusion?
A: Yes. It is usual.
Q: What is the purpose of that statistical report?
A: It is to give vantage to the evidence.
Q: Did you carry out the statistical report here?
A: No, I give that for the conclusion of A3(a)..
Q: You did for the first one?
A: Yes, in the first para says ,”matching with B10”
Q: Appendix page i?
A: On page 3, the first indication where it matches and probability …
Q: You made the statistical report for this?
A: Yes.
Q: Why did you do that?
A: Indicate common origin and the level of certain peaks and necessary to do that.
Q: It would be important for the report?
A: Yes, important when giving the evidence to court.
Q: This statistical report, what did you called it?
A: Statistical evaluation.
Q: What does it look like?
A: The used of it… (SAKSI TIDAK SEMPAT HABISKAN JAWAPAN).
Q: It forms part of your report?
A: The figures…(SAKSI TIDAK SEMPAT HABISKAN JAWAPAN).
Q: Can we call it something?
A: Statistical evaluation.
Q: This forms part of the report. Would it be correct to say that without the benefit of the statistical evaluation, it would not be able to produce your report?
A: No,.
Q: You don’t need to do that?
A: You only need that to match one against the other.
Q: Was the statistical report necessary for the production of P25?
A: Yes.
Q: Did you give it to the police?
A: No.
Q: But you agree that statistical evaluation would be important in assist us evaluating your result?
A: Yes.
Q: I take you to appendix again. Locus D8S1179, seminal stain from trousers, the first column, can you take us to the electropherogram which indicates this conclusion? Sorry, B5 and not A3. That would be page ii of
Appendix.
A: Yes.
Q: Let’s look at the locus D8S1179 for the non sperm extract. You had dentify 12, 13 allele as the complainant’s.
A: Yes.
Q: Does the evidence at page ii, does it solely indicate or identify a contribution of person allele 12 and 13?
A: No. as I said earlier, that assessment cannot be done just on one locus.
Q: 12 13 14 15 alleles. You’ve identified that 12 and 13 were from the complainant.
A: Yes.
Q: There’s a possibility of a third party on that locus?
A: Yes.
Q: Can we have a look at B5. There are 4 peaks, there must be at least 2 contributors right?
A: Yes.
Q: If there are more than 2 peaks, must be two or more persons.
A: Yes.
Q: Less than two peaks is one person?
A: Yes, sometimes… it cannot be more than one person. If 1 peaks, can be 1 person or can be more than 1 person if both had homozygote in. That’s what I’m saying that it cannot be done just based on one locus, must
based on the whole results.
Q: Looking at this D8, would you agree that there was a possible the contributions of 10 contributors?
A: No.
Q: 10 genotypes?
A: [no answer]
Q: From this locus, can we say that it could be 12, 14 as well?
A: Yes if we have no other information.
Q: Can I suggest another combination [GIVES SOME COMBINATION OF THE ALLELES].
A: Yes
Q: What does that mean?
A: Ten possible per mutation, we are doing it mathematically.
Q: From this locus, in layman’s term, [12, 13] being one of them, [15, 15] being another one of them, out of ten combinations, what was that mean?
A: They are possible per mutation. In the absence of any other information.
Q: Looking at D8 alone, what can you [12, 13] is not excluded from this combination?
A: Possible.
Q: In other words, it doesn’t tell us for certain that 12, 13 is the contributor?
A: Yes if just based on one locus.
Q: Your final assessment based on entire profile?
A: Yes
Q: Any other method or statistical evaluation carried out to arrive at your conclusion?
A: Yes, by looking at the relative peaks heights.
Q: Apart from that, any other method to come to your conclusion?
A: Yes, putting it into our software and assessing whether the respective parties could be the contributors?
Q: The software had produce something?
A: Yes, another assessment.
Q: You cannot say just an assessment. It is important. You did it for reasons.
A: Yes
Q: So, we should know what the result is, isn’t it?
A: No.
Q: So you want us to just go on this electropherogram, that’s all? Even though there are other methods?
A: It is the standard procedure
Q: There are other sources, we don’t know what it is because we are not be given them. You agree that this is important right?
A: This could be an alternative too
Q: This is important, if not, you wouldn’t do it otherwise, don’t you?
A: Not for this because there is an unknown contributor.
Q: Generally, you will employ other method?
A: That is if statistical evaluation, if you want to assess the statistical probability of certain individual in that mixture.
Q: So you have gone one step further isn’t it?
A: Yes, but here we have unknown male.
Q: But can we do that in this particular stage?
A: Not at this stage where the..
Q: No, I asked you, can you list out other sources apart from this 36 pages electropherogram? Lets make it simple. You have your report, support by your Appendix. Then you have your electropherogram produces by
software that we are not familiar with it. Right?
A: I don’t think that it would be appropriate to answer..
Q: Are you familiar with that software?
A: It’s not a matter of familiarity here.
Q: So you are not familiar with it?
A: I don’t think that..
Q: That’s why you have technicians to worry about it.
A: it is part of the work process, having people trained in certain area.
Q: You have a certificate in running of the software?
A: Yes.
Q: Can you produce it?
A: is not with me now.
Q: Very well. Going back to the locus D8, you were telling us that apart of this 38 pages graphs, there are other statistic evaluation. And this was also produced by the same machine?
A: No, it is not. It is by different machine, different software
Q: What is the name of the other machine?
A: DNA view.
Q: The first machine was applied bio system machine.
A: Yes, Applied bio system genetic analyzer.
Q: That is responsible for producing the graphs?
A: Yes.
Q: DNA view is responsible in?
A: Doing statistic evaluation.
Q: Produces other documents?
A: Yes.
Q: Both of these machines, there are different software. We go to DNA view. What is the name of that software?
A: DNA view.
Q: Are you trained to run this software?
A: Yes
Q: Are you familiar with that software, so whatever had been produced by that software, you can assure us that it is correct?
A: Yes.
Q: In that statistical evaluation, how many pages?
A: 3 pages.
Q: Can you confirm that those 3 pages form parts of your report?
A: Yes.
Q: Those 3 pages contributed to the end product and essential to your report?
A: Yes.
Q: Are they called as statistical evaluation? Those three pages separate documents?
A: Separate documents.
Q: Their title?
A: Calculation of match probabilities.
Q: Now back to locus D8, of B5. You would have to calculate the match probability?
A: Not for this profile.
Q: Which profile you count the match probability?
A: A3.
Q: You have to exclude certain alleles?
A: For A3 (a) and )b)? No, all alleles are computed.
Q: You did the calculation of match probabilities?
A: Yes.
Q: What is the purpose of this?
A: To compare whether it matches by coincidence from a randomly selected random person and that would give the level of certainty.
Q: Without the benefit of that, you wouldn’t give the level of certainty?
A: Yes
Q: So we don’t know what is contained in the documents. We don’t know whether there is match probability or not?
A: No, that’s not being put in the right way. I have to do it statistically. We have a paper for the compilation of database. For the match probability of certain loci, we know that it should be certain level, and that level will
indicate the certainty of the match. The exact computation is 500 and 700. it can go higher than that, or slightly lower than that. Therefore, we put it in the exact figure.
Q: Need to do the calculation
A: Yes, because the court needs to know the exact figure.
Q: Did you carried out the match probability of B5?
A: No.
A: Did you carry out the match probability to other sample apart from A3?
B: No.
Q: Page 6, would you agree that locus D8 does not tell you that there is a person in this mixture of [12, 13] allele?
A: B5 has its own contributor, it is swab taken from non individual and therefore we can put it in the profile of non contributor which is the donor of blood specimen.
Q: Does this locus tell you that there is a person of nature [12,13]?
A: Yes.
Q: Are there other possibilities?
A: We cannot view it per se that this is the DNA profile from the swab which is taken from non person, and therefore we have non contributor here. If this stain taken from a surface and from an item not from individual, then
your agreement will be varied.
Q: You’ve got a complainant here.
A: Yes, this was taken from him.
Q: So, you just assuming therefore [12.13] is his?
A: Not assuming.
Q: 8 others possible contributors?
A: No, that s how the per mutation works, you know.
Q: Let’s take out Male Y and the complainant. What about other possibilities? 8 or 9?
A: there are 8 persons but not reflected here.
MY: YA, we have come though this. The witness has agreed that there could be many, but also she used the word per mutation so on. But it must base on the whole thing. That’s why she couldn’t agree because she cannot
look at the locus in isolation. He is now repeating the same questions.
RK: I’m not repeating. She doesn’t answer my question.
Q: But from the information given, you assuming, isn’t it? That there is [12,13]?
A: Yes. It is a swab taken. I’m not just assuming.
Q: Let’s go to the next locus, I take you to locus D2IS1338. There are 4 peaks there. The first one is the stutter, the small one. We have 18, 19, 20, 23 alleles from this locus. How many contributors? Two?
A: Possibly.
Q: Is there 24 allele there?
A: There’s no 24 there.
Q: So if you were look at this, you cannot tell us that we have allele 24?
A: Yes.
Q: At face value, can we say that there is contributor?
A: Again, Yang Arif, this is not the way. He’s going to locus by locus.
Q: Can you conclude that there is a contributor with 20 and 24 only?
A: If you are asking me can the [20, 24] individual be indicated in this locus, there would be dropout. It could be indicated because you are choosing D2S1338. D2 is locus with the large amplicon, meaning that it is larger
than the remaining loci.
Q: So you say that 24 could drop out?
A: Yes.
Q: Are you excluded other alleles from dropping out?
A: []
Q: So you didn’t analyze the sample further to determine drop out?
A: I didn’t.
Q: I put it to you that the evidence of locus D8 doesn’t tell us for certain that a contributor with [12,13] allele, or [15,15] allele contributed to it?
A: I disagree.
Q: I put it to you that there are possibilities that the other combination of contributors reflected in locus D8 cannot be excluded contributors?
A: Yes.
Q: I put it to you that this evidence of locus D8 does not tell us without a doubt that any particular allele belong or is contributed by a particular person? Agree?
A: Yes. If the question is based per se on the thing you asked.
Q: I put it to you further that evidence of locus D8 again does not tell us without a doubt that a contributor of the locus were those with 12,13,15 allele. Do you agree?
A: Disagree
RK: YA, we need to look at the exhibits as we agreed yesterday.
YA: Another aspect of question la. 20 minutes cukup?
RK: Half an hour.
YA: We’ll assemble at 11.20 a.m.
[10.52 a.m.] Stand down.
[11.23 a.m.] Pihak-pihak masuk ke Kamar Hakim.
[11.27 a.m.]Pihak-pihak keluar dari Kamar Hakim.
Kes ditangguhkan kepada 3.00 petang kerana Peguambela meminta masa untuk memeriksa eksibit.
[3.15 p.m.]
Sambung pemeriksaan balas SP5.
Q: You told us yesterday that your lab in Jabatan Kimia Malaysia in PJ is accredited by ASCLD, right?
A: Yes.
Q: There must be guideline set out as a result of this accreditation for DNA profiling.
A: Yes. There would be written procedures.
Q: Is this internal guideline for Jabatan Kimia Malaysia or are they general guidelines?
A: No. They are the procedures for the quality system which is the ISO system.
Q: You told us that the ASCLD accredited Jabatan Kimia Malaysia, right?
A: Yes.
Q: Does ASCLD has its own accredited guideline?
A: Yes.
Q: Is it documented in any document?
A: Yes. They have the ASCLD lab menu and the ASCLD lab criteria.
Q: Is it updated periodically?
A: Yes. The ASCLD lab current quality system is based on ISO 17025.
Q: This guideline that you’ve just told us about are they applied equally across abroad? For DNA sampling.
A: Yes
Q: You told us that you had relied on certain machines in this case. Two machines.
A: Yes. Instrumentations is used.
Q: And they are manufactured overseas. And does this manufactures have their own guideline?
A: Yes, they have. In fact they are the supplier of the instrumentations applied by our system, and their system has been validated for forensic use.
Q: So, do both the machines have guidelines? Do they come with manual which lay out how to carry on profiling and so on?
A: Yes. They told there’s a menu on operation, checks and calibration.
Q: What about the PCR kit? Do they have guidelines also?
A: Yes. They have written instructions.
Q: You told us about ASCLD guidelines earlier. Do they correspond to the manufactures of the PCR kits?
A: Yes. They have recommended procedures for use of reagent and check and controls that has to be placed in carrying out PCR.
Q: Are these standard guidelines?
A: Yes, they are.
Q: Bearing in mind those guidelines that we talked about just now, the machines guidelines, the PCR guidelines, the ASCLD guidelines. There are many guidelines in the regulated fields. Can you tell us what is a reporting
threshold? Is there such a thing as reporting threshold?
A: Yes. Based on our validation studies carried out, we recommended a reporting threshold of 50 RFU, for single source DNA profile.
Q: Relative fluorescent unit?
A: Yes.
Q: How are peaks recorded and reported according to the RFU units?
A: Once the peaks has been assessed and accepted, the RFU is not documented in the report or in our appendix. The RFU will be called and they are on the electro-phoreogram as printed out.
Q: So from that information is the RFU?
A: Yes.
Q: You said the threshold is 50 RFU. Is that the minimum amount before you report the peak?
A: Yes. For Jabatan Kimia Malaysia, that would be the minimum threshold before it is considered as an allele.
Q: So, anything below that is consider what?
A: Anything below that would be reported but would not be considered for statistical evaluation.
Q: Would this include the stutter?
A: The stutter has its own range.
Q: So, there are rules for determining stutters?
A: Yes. They have their own rules.
Q: Do you know what the manufactures generally recommend as a reporting threshold?
A: They do not generally lay any fix ceiling on the reporting effect, but they recommend that each lab carry their own validation studies to determine the minimum threshold.
Q: So they leave it to the labs?
A: They recommend the lab do their own internal validation studies.
Q: You told us the lowest peak is 50 RFU. Not below 50 RFU? At least 50 RFU?
A: To be constant for statistical value, at least 50 RFU.
Q: Is it applied to all peaks?
A: Yes. That applies to all peaks.
Q: Also for homozygous peaks?
A: Yes, even for homozygous peaks. But we are more conservative for homozygous which reads at 50 RFU that we interpret with caution although..
Q: You are saying that you still apply 50 RFU for homozygous peaks?
A: Yes. But with careful consideration.
Q: Why careful consideration?
A: Because there could be allele dropout.
Q: So, what would normally be the threshold for homozygous peaks?
A: It depends to because this is a multiplex essay and each locus would have certain characteristics and we would be more conservative with the loci with larger amplicons. We will be more conservative with the loci with
lower amplicons and less conservative with loci with larger amplicons. Because larger amplicons are..
Q: I’m talking threshold. So, in general terms?
A: For homozygous in general terms?
Q: Yes. What would be the threshold? In terms of RFU.
A: We lay out a threshold of 50 RFU minimum. Even for heterozygous and homozygous. But the homozygous assessment comes with a particular loci which that homozygous was observed. And there would be careful
consideration of homozygous occurring at the loci with shorter amplicons than those loci with larger amplicons.
Q: So, this 50 RFU for all the sort of peaks? That is your guideline?
A: Yes. Our written guidelines.
Q: For Jabatan Kimia Malaysia?
A: Yes.
Q: You mentioned drops-out earlier. Can you tell us very briefly what is drop-outs?
A: It is the expected value that is expected to be seen in DNA profile is absent and there are many causes for the allele drop-outs. One of the main causes is …
Q: Before we go into causes, how would you know that there is a drop-out?
A: Normally you will be able to detect it if you have a reference specimen.
Q: A reference specimen?
A: A reference profile or a reference specimens.
Q: In other words, would it be correct to say that your way of determining if there is a drop-out is based on information given to you?
A: Based from information from reference.
Q: But you are not forming your conclusion directly from the evidence before you, in terms of a graph.
A: You wouldn’t be able to do that. It is too un-scientific to assess allelles without reference. That would be assumptions without a reference.
Q: You have come across drop-outs as you go along, right?
A: Yes.
Q: In order to determine what drop-outs are, you are telling us that you rely on information given to you. Reference sample.
A: Not given to me. Which was used to analyse.
Q: It is not part of the evidence before you. It comes from a different source.
A: Yes.
Q: That information was given to you.
A: Yes.
Q: So you rely on that information?
A: Yes.
Q: So, you are influenced by that information. Is that correct?
A: No. That is not influence. That is the scientific way of deduction.
Q: You are given a reference sample?
A: Yes.
Q: Is information given to you?
A: Yes.
Q: Quite independence is what before you. What is before you is the graphs and all that, isn’t it?
A: Yes.
Q: That is the evidence independence, right?
A: Yes.
Q: You are being given a reference sample.
A: Yes.
Q: Another independent source, right?
A: Yes.
Q: So that influenced your mind, isn’t it? You are relying on that, aren’t you?
A: I disagree.
Q: So the reference sample matters nothing to you?
A: It matters, but you are putting it in a wrong context.
Q: I put it this way, if you receive a reference sample, you are going to act on the reference sample, right?
A: Yes.
Q: When you act on the reference sample, youl’ll apply the knowledge of the reference sample that is the evidence before, isn’t it?
A: Yes. I have to use that.
Q: And the reference sample you received in this case is which one? Which one is the reference sample of the complainant?
A: B10.
Q: I take you to the appendix of your report. The reference sample is at page 3, isn’t it?
A: Yes.
Q: The last column. B10 – bloodstained specimens labeled “Mohd Saiful Bukhari bin Azlan”. So you got all the alleles there?
A: Yes.
Q: For the first locus, 12, 13, 29, 32, 11 and so on.
A: Yes.
Q: How many loci are there?
A: A total of 15 STR plus the [].
Q: And you come out with Saiful’s allele as a result?
A: DNA profile. Yes.
Q: So, when you studied the graphs in this case you have in mind already Saiful’s allele, don’t you?
A: Yes.
Q: You were not independent when you embark on this exercise, don’t you? You were influenced by Saiful’s allele, don’t you?
A: I disagree. There is no bias here.
Q: Is it possible to determine drop-outs without the benefits of the reference sample?
A: No. I’m not able to. You are calling it drop-outs because you are referring to the the samples.
Q: So, there is no other exercise in determining drop-outs, right?
A: Yes. There will be no exercise unless you have a reference sample.
Q: That is what your experience has taught you?
A: Yes.
Q: Are you aware of an organisation of International Society for Forensic Genetics?
A: Yes. I’m a member of that organisation.
Q: Are you aware of papers published by it?
A: Yes. Because as a member I received the issues, every issues.
Q: In those publication by the society, are you aware of the procedures in determining drop-out?
A: Yes.
Q: Are you aware that those procedures are not adopted in this case? Do you agree?
A: I disagree.
Q: You have adopted procedures of drop-out?
A: Yes. We have our own system of procedures.
Q: I’m asking you, in relation of this publication, were these steps employed in this case?
A: We have our own procedure.
Q: When you say “we”, is that Jabatan Kimia Malaysia or the society?
A: Jabatan Kimia Malaysia.
Q: The organisation has list out certain procedure of determining drop out, right? Were those procedures adopted to treat or identified drop-outs in the particular case?
A: I am unable to answer the question unless you outline what are the procedures listed by the organisation.
Q: Were international procedures adopted in the identification of drop out in this case? Apart from the one adopted in Jabatan Kimia Malaysia.
A: Yes, we have adopted international procedures.
Q: What are they?
A: Perhaps you can outline it. I’ve forgotten. This is a 2006 paper.
Q: I’m asking you, were international standard adopted in the identification of drop-outs. Drop outs were identified in this case, right? So, were the criteria from this international standard adopted and employed in this particular
case?
A: I’ve said We have our own procedures.
Q: You don’t want to answer the question, isn’t it? I think you have made up your mind. Everything is now recorded. So, you don’t want to answer my question?
A:: Yes. I don’t want to answer the question because it is very general way of putting specific…
Q: Fair enough. Next question. Your lab, the Jabatan Kimia Malaysia lab. We’ve talked about how drop-outs will occur. So let put them aside now. How about situation where drop-outs will not occur?
SP5: Can I just ask a question, YA?
RK: No, you can’t ask question.
SP5: No. Because it’s…
RK: You can ask questions to your counsel. She’ll…
SP5: No. Because that questions that has been asked could be applied to a low copy template DNA profiling.
RK: We don’t need to go into that.
YA: If you don’t understand the question, you ask him to repeat. If you want clarification, you ask.
RK: The question just now is clear, right?
SP5 Yes, allele drop-outs for what?
YA: So, your question again?
RK: I’ve gone past that. I’m going to a different question now. I think what the witness is trying to ask me is in relation to what I’m asking her earlier. I’ve past that. I’m going to another question now.
Q: In relation to your lab, Jabatan Kimia Malaysia lab in PJ. Are there procedures to determine how drop-outs might not occur?
A: No. Not that I’m aware of how to avoid drop-outs.
Q: Are you aware for T-value? Drop-out threshold?
A: We don’t adopt it.
Q: Are you are aware of it?
A: Yes.
Q: What is the T-value?
A: It is threshold value where drop-out is assumed to have occurred.
Q: Who has recognize this T-value?
A: Again, I have feeling this is about low copy template DNA. This is not the case here. It is not the profiling here.
Q: I’m not asking you in what context it is. I’m asking you…
A: YA, this is also the exercise used in…
Q: Was there drop-outs in this case?
A: Yes.
Q: So, T-value is relevant to drop-out, isn’t it?
A: But, it is of no significant here.
Q: Is T-value relevant to drop-outs?
A: Yes, it is relevant to drop-outs.
YA: Whether it is significant or not, the DPP will ask. And you can explain it later.
Q: What is T-value?
A: It is the threshold value in which where the peak below that is not considered.
Q: Who set this threshold? Where did the T-value comes from?
A: I think it comes from validation studies.
Q: Is that local studies or what?
A: The validation studies can be conducted by any laboratory.
Q: So, it is a safeguard, isn’t it? In relation to drop-outs?
A: It is for better understanding.
Q: Is it adopted in this case? Was the threshold adopted in this case?
A: It is recommended for low copy template DNA.
Q: Was it employed in this case?
A: It is not employed in this case.
Q: Have you lab ever validated the T-value before?
A: No.
Q: It is something new, is that right?
A: It is recommended for people who carry out low template DNA profiling.
Q: You told us that you are aware of the International Society for Forensic Genetics which you are a member of. They come out with publication, do they?
A: Periodically. And it is about 4 issues per year.
Q: Do they come out with the latest development for DNA profiling?
A: They come out with the recommendation in all aspect of DNA profiling.
Q: The machine you informed us this morning, the second machines you called it DNA view machine.
A: It is not a machine. It’s a software.
Q: This software, how long is it? Is it new? How old is it?
A: More than 10 years old.
Q: Quite old, isn’t it?
A: No. It is regularly updated.
Q: Did you update the software?
A: No. It is a software from [] and he regularly updates it.
Q: When was it last updated?
A: The last update is in 2008 or 2007.
Q: So, in the last 3 or 4 years it is not updated?
A: It is not updated in the workings.
Q: What are stutters?
A: Peaks which are observed, which are one repeat unit less than the real peaks of the real alleles and that usually occurs from slippage during PCR.
Q: Can you show the example from one of your graphs.
A: Page 1. You can just take the 1st locus, D8S1179. You observed a small peak just below allele 12.
Q: The three peaks there?
A: Yes.
Q: The first one? The shortest?
A: Yes, that is the stutter for allele 12. And for allele 13 it will not be seen because there’s a [] of allele 12 there. The stutter is usually less than 20% of the real allele.
Q: Stutters are basically [] peaks?
A: Artifacts created during PCR.
Q: So they are negligible?
A: They have to counted for because they are stutters but are not counted as peaks.
Q: Are there guidelines in relation to the identification of stutters?
A: Yes. They recommend validation studies on the system to account for the range of your stutter values were you will consider it as stutter peak rather than real peaks.
Q: Those guidelines can be found? Where?
A: Jabatan Kimia Malaysia has this guideline.
Q: Internal guideline?
A: Yes.
Q: Are there any accredited guideline?
A: You cannot apply a single guideline to a laboratory. It has to be established individually.
Q: Is there any difference in this guideline from the one in Singapore?
A: There will be variations.
Q: So, the stutters will be different?
A: There will be variations.
Q: But generally they are the same?
A: Their ranges might fluctuate. Because it depends on your testing systems.
Q: The concept of stutters are the same throughout the world, isn’t it?
A: Yes.
Q: Are there any guidelines internationally for stutters and how to identify it and so on?
A: Yes, there are.
Q: Does your lab follow any of those guidelines?
A: We do.
Q: Which one are those?
A: They are all in the books now. In fact the stutters is all literature.
Q: Can you name us for example any society which..
A: John Butler has good books on interpretation and whole chapters for stutters.
Q: What are the stutter guidelines for each of the locus?
A: We have established a range for each of the locus.
Q: What is the range? Perhaps you can explain by way of an example.
A: …
Q: Before that, your PCR summary. Can you tell us what is the stutter range for each of the locus?
A: I wouldn’t be able to define the range, but it is in our…
Q: But it is important, isn’t it?
A: It is important.
Q: The range of the locus of your stutter is important to your findings, isn’t it?
A: Yes. It is a guideline.
Q: That’s why you’ve made a record of it?
A: Yes.
Q: It could possible be wrong?
A: It could vary.
Q: Can you say it is free from error?
A: The range could vary. We are only putting range. It could fall out of the range some other time.
Q: Is this in a list or something? Is it in another report?
A: We have a range for stutter where we study and break a range from our studies.
Q: How is it documented?
A: Documented as for D8 as what is the stutter range that is carried out in studies and found to be in this range.
Q: Can I say there is another report?
A: No.
Q: Then what is it called?
A: That is in the validation of this system.
Q: You told us this morning about the statistical evaluation which you have…
A: That is different.
Q: So, there’s a statistical evaluation which you have, which has not been given to us. Now you are telling us there is another document. With the stutter range, which is also important to your finding, isn’t it? You are in
possession of that document? What do you want to call it?
A: It is in our validation filing system. Validation manual.
Q: You have a copy of the validation manual?
A: Yes.
Q: The validation manual is crucial to your finding, isn’t it?
A: It will aid us in our interpretation.
Q: It is necessary, isn’t it?
A: It will help us. It will assist us.
Q: You are in possession of it?
A: Yes.
Q: Can you give us an example of a stutter range.
A: Off hand, I can’t quote here. Because there are 15 here.
Q: Would they go in percentage? In relation to what?
A: They go in percentage. In relation to the peak [].
Q: Look at locus D8. So you’ve got a small stutter?
A: Yes.
Q: There’s a high peak next to it?
A: Yes.
Q: So, the range of percentage is between the stutter and the peak, is that right?
A: Yes.
Q: For the first stutter, can you look at page 1?
A: Yes.
Q: So you got allele 12 for locus D8. The peak is 1654, is that right?
A: Yes.
Q: The next peak is 1627.
A: Yes.
Q: So, at this moment of time, you cannot tell us what the stutter height is?
A: We accept the threshold in the system.
Q: No. At this moment of time. You can’t assist the court in stutter height, can’t you?
A: No. It can be read of.
Q: So, what is the stutter height now?
A: Because it is established as stutter, we have not [].
Q: That information can be found in the validation menu?
A: Yes.
Q: Is there method in which you arrived in the figures of the stutter ?
A: There are procedures for establishing the range for stutters.
Q: What procedures it is?
A: One of the easiest way is to computed it in the population database. This can be carried out concurrently with the population databases where we compute all the stutters of all of the loci for every sample.
Q: Under what guidelines are you being guided when you come to this conclusion?
A: These are straight forward way of measuring because it is just one. So you have to carry out a number of significants samples which is 100 or 200 samples.
Q: Do the manufacturers give guidelines in determining stutters?
A: Stutter range from their studies, but they always recommended that each laboratory does their own stutter range.
Q: So, your lab doesn’t conform to the manufacturers guidelines?
A: We don’t use any stutter range from any other sources except that is carried out in our system.
Q: Can you tell us what overloading is?
A: It means that the amount of the DNA template that is being used is much higher than the optimum amount.
Q: Too much DNA?
A: Yes.
Q: And the results are not accurate?
A: They are still accurate. There would be pull-ups.
Q: So, what would you do as a result of overloading?
A: You can either [] it down the PCR products and do a second run. Or you can start all over again and do a PCR.
Q: So, manufacturers have this guideline as well? How to handle overloading?
A: Yes.
Q: Does your lab conform to this guideline?
A: Yes, we do.
Q: Why does your lab conform to this guideline?
A: Because when there is overloading, the peaks get too high and they don’t get [], and you’ll see a lot of artifacts. And very [].
Q: So you are selective on the guidelines to follow?
A: We follow the stutter guidelines but the ranges that is being adopted has to be carried out in individual laboratory. But the guidelines on what to do with the overloading there is international guideline.
Q: You mentioned pull-up. Please tell briefly.
A: A pull-up happens when there is overloading. That is because of the spectrum overlap and this electrophoresis of DNA where the separation occurs, the detection is by signals from respective []. And this [] has their own
spectrums and their own laps in the spectrums are like rainbows and there are spectrum overlaps. But the own laps are corrected by the machines through calibration. But when there is overloading, the overlaps become very
significant and the correction would not be able to [] entirely the signals that can be detected by the [] colour ranges and therefore the blue signals can be detected in the green [] as in yellow.
Q: Can this pull out creates a false appearance of peaks?
A: No.
Q: Can you show the sample?
A: Page 20, B10. In B10, there is a straight line which is occurring at the same places down, that is draw a straight line down. If you look at peak D19S433.
Q: The colouring is significant?
A: No. It will make it clearer. It does not matter.
Q: Carry on.
A: The third row. You have that peak 13. If you draw a straight line along this peak 13 up and down, you’ll see a single line up the blue and up in the green area. That shows there is a pull-up in the region. This peak 13 is 8315
[] which is a very strong signal. And that straight line will be [] in all. As you look down where the peaks are high, there will be [] straight up. Therefore if you have a peak in another colour range, that is in the line of the pull-up, then it
will create a [] in the peak shapes and it will require amplification or a re-dilution of the PCR products to enhance the defeiction of the real allele. If you have a straight line again as in the fourth row D5S818, again that first peak is
very strong and if you draw a staright line up, it will show a straight line up which is the pull up.
Q: Are the red lines produced by the machines?
A: Yes.
Q: Are they automatic?
A: This is in the [] software. This is the smarter version. It is from a machine. It give indication of a pull out.
Q: There are 2 types of DNA profiling. One is single and one is the mixtures.
A: They are not two types of DNA profiling.
Q: You have to interpret mixtures sometimes, right? Like in this case. That means there is more than one contributor.
A: Yes.
Q: Do you have any guidelines for mixtures?
A: Yes.
Q: What are they?
A: They are carried out in validation studies. It is in our menu.
Q: Those guidelines are carried out in validation studies? What do you mean? The same document you are referring earlier?
A: No. It’s a document of studying of mixtures. Mixtures interpretation.
Q: What are the guidelines?
A: Interpretation of those mixtures.
Q: What are the guidelines? You told us there is guidelines. Because this is a case involving mixtures. So, what are the guidelines applicable in such scenario?
A: You have to state which particular scenario. Because we have mixtures for major minor which we have drawn up. Mixtures of 3 individue .
Q: Major then.
A: Major minor. For major minor interpretation, you have the [] guidelines where the manor contributor can be inferred, interpreted as a single source. But for the minor contributor, it will have to be interpreted as a mixture.
Q: How do you distinguished between the major and the minor?
A: The major alleles will be at least 50% higher than that of the minor.
Q: So you go by ratios?
A: Yes.
Q: Are the ratios recorded anywhere?
A: What do you mean? How can measures the major minor when the peaks high is greater than 50%.
Q: So you see it visually from the graph?
A: Yes. You can read out the RFU.
Q: Do you know what is peak high balance is?
A: Yes.
Q: What is it?
A: It is the ratio for the peaks in the heterozygote.
Q: What is it?
A: It’s the relative peak high between the minor peak and the major peak.
Q: Is there any guidelines on that?
A: Yes.
Q: What is PHB? Peak high balance?
A: We have established a 60% for normal DNA profile. For a low copy template that ceiling cannot be placed because it will be less than 60%.
Q: Do manufacturers have guidelines on the PHB?
A: Again, they recommend you do your own service.
RK: At this jucture, I’m finished on this area. This is in relation to guidelines and so on. But there might be a few more as we go along. But for now, it is as it is.
YA: Go on to other aspect then?
RK: Other aspects are quite long, YA. Very long.
YA: Kalau boleh habis hari ini, kita sambung hari Isnin.
RK: We can start it fresh. Once we start, it will be difficult on the flow. [] prejudicial of my learned friend.
YA: Not the question of prejudicial. The question is sekarang ni kita ada masa lagi. Our office hour habis pukul 5.
RK: Can we start on Monday at 8.30 a.m.? I can finish it at 11 a.m..
YA: Continue at least until 4.30.
RK: Nevermind. I’ll go on the guidelines again, YA.
Q: We go again to PHB. Manufactures would tell you to do it yourselves.
A: Yes.
Q: Do they not set a PHB guideline?
A: Yes.
Q: What are the guidelines?
A: Their testing system would be different from your testing system. They would have established what is their peak high balance ratio and…
Q: What are the manufacturers guidelines as to PHB? Is it the same as yours, 60%?
A: No. Theirs are different. Their labs adopts various peak highs.
Q: What does the manual say?
A: The manual says you have to do your own.
Q: They don’t specify percentage?
A: No. They recommend you to do your own laboratory system. That cannot be fixed for any laboratory.
Q: So you confirm the manufacturer does not identify the peaks?
A: They do their own.
Q: What is it?
A: I can’t remember, but their recommendation is you do your own.
Q: As a guideline, they have also given a figure, isn’t it? As a guideline.
A: That’s based on their studies. They cannot recommend it on our lab. Their studies indicate certain percentage.
Q What is a mixture ratios?
A: A mixture ratio for the whole profile? That cannot be ascertain. Because the mixture ratio would varies between loci.
Q: For example in this case. You’ve got a mixtures of contributors. In this case not more than two. Is there a ratio that you would come to in determining this ratio?
A: No. That could not be ascertain. That would vary between the loci.
Q: So mixture ratios cannot be determine?
A: No. Because the ratios varies between loci.
Q: Like what?
A: [Hypothetical explanation]
Q: So the next loci is different?
A: Yes. The variation will occur because of the multiplexing.
Q: So, it won’t be the same for all the loci?
A: No.
Q: What degree of variation in the mixtures ratios between loci would you consider in your lab?
A: I don’t understand the question.
Q: What degree of variation in mixtures is acceptable in your laboratory? The variation can’t be too large, isn’t it?
A: I don’t think there is a need for guideline []. Because of the variation, the acceptability issue is not here.
RK: I’ve finished on this topic. I’ve got nothing more.
YA: I take your word there is nothing more on this topic. And on Monday you’ll start a new topic that is completely different from this topic?
RK: I’ve got 5 more topic, YA.
YA: But completely different from today’s topic?
RK: Yes. Completely different. But as I said, there might be overlap. I’ve finished on this one.
YA: Monday continue.
[4.17 p.m.] Adjourned.
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