Wisconsin Public Television University Place Wednesday Nite at the Lab  Jennifer Laffin, "Testing Chromosomes to Detect Diseases in Children"  November 17, 2010  cc >> Tom Zinnen: Welcome, everybody, to Wednesday Nite at the Lab. I'm Tom Zinnen. I work here at the University of Wisconsin-Madison Biotechnology Center. I also work for UW Extension Cooperative Extension. And on behalf of our partners at the Wisconsin Alumni Association and Wisconsin Public Television, welcome to Wednesday Nite at the Lab. We do this every Wednesday night, 50 times a year. Tonight I'm delighted to be able to welcome Jennifer Laffin to Wednesday Nite at the Lab. She'll be talking about her work at the Wisconsin State Laboratory of Hygiene and telltale clues using genetic tests to diagnose diseases in children. Jennifer is from Kaukauna, Wisconsin, and got her BS degree here in genetics in 1996. She went over to Iowa City and got her PhD in genetics in 2003 and came back to Madison with a fellowship in 2006 and then as an assistant professor in 2008, and just this year she became co-director of the clinic. 

I look forward to her talk. Please join me in welcoming Jennifer Laffin to Wednesday Nite at the Lab. [APPLAUSE] >> Jennifer Laffin: Thank you. Thank you, everyone. It's a pleasure to be here. So I'm Jennifer Laffin. I'm co-director of the Clinical Genetics Laboratories at the State Lab of Hygiene. The State Lab of Hygiene houses the Clinical Genetics Laboratories at the UW. So we have sections in cytogenetics, molecular genetics and biochemical genetics. So that covers all the areas of clinical genetic laboratory testing. 

So it's a really great place to be right now. And in the audience is my co-director, Dr. Raca. We oversee the cytogenetic section and the molecular genetic section. So tonight I'm just going to talk a little bit about the types of testing that we do, and then give you some examples of the types of cases that come through our door. So you can kind of get a feel of what happens to a specimen, how do we use our different techniques and how do we use our tools to try to get at some answers for our patients. And then learn something as well about the biology and kind of feed the research and feed the clinical side as well. So all the sides kind of work together to try to bring better patient care. So the types of testing, the types of specimens that we work on, so we do everything from prenatal testing where we take amniotic fluid specimens, chorionic villi specimens, so little placental biopsies, to oncology testing. So we do a lot of testing of patients that have leukemias, lymphomas and tumors to try to identify genetic markers that can be both prognostic and drive treatment as well. And we also do a lot of testing for individuals who have developmental issues. 

We do testing for fertility, anything that has an genetic aspect. And genetic aspects, as you're learning more and more, is hitting all fields of medicine. So genetics is really growing and growing fast. So tonight I'll be talking about cytogenetics and molecular genetics and how we have an opportunity here to use our research tools as well as our clinical tools to help our patients. So I'll start with a little primer in cytogenetics. So cytogenetics 101. So we have the opportunity to work at all levels or all resolutions of genetics. So we start with chromosomes which are composed of our DNA protein and they carry our genetic material in our cells. And then go all the way down to molecular so we can look at bases as well. So cytogenetics is the study of structure and function of chromosomes-- Yeah? >> Could you tap your microphone? Thank you. 

That activates it. >> Okay, sorry. Okay, so in order to see chromosomes we need living tissue that's actively dividing. So if you remember the types of specimens that we have, we get blood specimens where you hope that cells wouldn't be actively dividing in your peripheral blood and we also get tumor specimens as well which are actively dividing, amniotic fluid specimens actively dividing. So we need living tissues in order for it to go. So we have cells that we either induce to go into mitosis or into cell division or we look at cells that are already actively dividing. So here we have a couple diagrams of just the cell cycle, what happens. And here's, on the right hand side, we have a diagram of really what's happening looking at the DNA inside of those cells. So at interphase we have DNA that's all strung out so that you can get activation of genes and silencing of genes, and then as cells go through mitosis, we need those chromosomes to condense and then normally what would happen is the chromosomes that condense line up at the metaphase place and then be split into the two daughter cells. So what we do is we need to capture those chromosomes at their most condensed state. 

So we poison those microtubules that would normally connect and then pull the chromosomes to their daughter cells. So we depolymerize those microtubules and then hold those cells in metaphase. So after we've done that, we bloat the cells a little bit. So we use a hypotonic treatment that swells the cells and helps with chromosome spreading. So what needs to happen is we need to put those chromosomes on a microscope slide so we can actually visualize them. So we swell the cells and then we fix them so they stay put. So that also lyses any red blood cells that might be still floating around but it stops that hypotonic swelling. And then this is the art part of cytogenetics. So it's the slide making and the staining portion. So when we drop the chromosomes there, we have to make sure that everything happens just so so you don't end up with chromosome soup. 

So when you drop the cell suspension on a microscope slide, that the chromosomes don't get spread off into every which way. We need them to stay in a nice little puddle so that we can identify one nucleus, one nucleus, one nucleus. So we can count the chromosomes and look at their banding. But we don't want them so tight that they're all crumpled up and laying on top of each other that you can't really identify which chromosome is which. So we drop the suspension of these fixed cells on to a microscope slide. They need to dry at exactly the right temperature and the right humidity so that you don't get overspreading, underspreading, and that everything will stain and band nice. The slides then get baked and then they're treated with trypsin. So these are pictures from our laboratory of what's happening. So actually the image over here is one of our drying chambers. So a technologist will be there with a patient's specimen, drop the slide, put the slide in the drying chamber, this has exactly the right humidity and temperature so that the chromosomes dry nicely, and then we can check spreading and concentration at the microscope right next door here. 

But then after they're baked, then they go through a process of trypsin, so that eats the proteins a little bit, and then stain. So we use the Giemsa stain. And then they get cover-slipped and then sent out to our technologists for analysis. So what happens most often, now we have a slide scanner, an automated slide scanner that will go through and try to look for metaphases for us, but we do often for like prenatal specimens sit at the scope and look through the whole entire slide looking for the best metaphase spread. So this image right up here is what you would see under the microscope. So the cell is spread and each of these are individual chromosomes. So there should be 46 chromosomes, and then a technologist will go through and find one, one, make sure you have both chromosome ones and go band for band and make sure that all the bands are in the right orientation and that they're actually present. So there aren't deletions or duplications or extra pieces of that. So then we also the capabilities of or we have software that allows us to electronically cut out the chromosomes and then arrange them in a karyogram. So right here you can see all the chromosomes. 

We have chromosome 1 through 22 are autosomes. And then males have an X and a Y and females have two Xs. So with every case that we have we got through and we make sure that everything is as it should be or if there's a particular abnormality that we're expecting, that that's there. So the next step up in resolution that we have, we have fluorescence in situ hybridization and then more molecular genetic techniques. So fluorescence in situ hybridization, or FISH, allows us to look at a very specific part of the genome. So what we can do is we can take the piece of the genome that we're interested in, we can label it with a fluorescent probe and then stick it to the genome. So you make that little fluorescent probe and then you can hybridize it, you can hybridize it either to interphase nuclei and count the dots or you can hybridize it to a metaphase spread and actually see which chromosome that it's hybridizing to. And there are a lot of different probe types that we can use to answer very specific questions. So we have whole chromosome paints. Those don't get used as often anymore now that we have a ray comparative genomic hybridization. 

But if there's a particular piece of a chromosome that you really suspect is coming from a particular chromosome, like for example this image up here, it was suspected that we know that the banding was consistent with chromosome 13, is that extra material that's attached to that chromosome 13, is that from the Y chromosome? So you can paint that and see that yes, in fact, there is chromosome 13 right here and that this red material right here came from chromosome Y. There are centromere enumerator probes. So we can look and count to see how many of each chromosome there are. We use those often in amniotic fluid specimens. So looking for the common trisomies. So 13, 18, 21 and we can look for the sex chromosomes very quickly without looking for rearrangements, just know that there are two 21s, two 13s, two 18s, and so on and so forth. And then locus specific indicators in the cocktails of multiple probes can be manipulated in different ways that we can look for very specific chromosomal rearrangements as well as microdeletion syndromes and duplications. So this example, these examples down here, this is from the DiGeorge region. So this is a recurrent microdeletion syndrome. So we have a piece of DNA that's very specific to the region that is commonly deleted in the 22Q deletion syndrome. 

So we can look for that and you can see in this specimen here's the normal 22. You have the green is the control probe that kind of marks that chromosome as this is chromosome 22. And then the red is that region that is commonly deleted in this syndrome. So you can see there's a normal 22 and the 22 over here has that region deleted. We can also, so as far as the manipulation of these, you can take locus specific probes and arrange them so that you can look for particular fusions or particular breaks in a gene. So if you know a gene is commonly broken in a particular disease, you can take two probes and then look for the break apart in the abnormal cells. The molecular genetics is a lot less visually fancy at the very beginning. The end is still really cool. So we generally start with a blood specimen from our patients. We get rid of the red blood cells and keep the white blood cells where all the DNA is stored. 

And then some of our assays do require RNA but most of our assays require DNA. And we precipitate out those proteins, and then you can precipitate the DNA out of solution. So you can watch that DNA come right out of solution. So if you think each cell in your body has about six feet of DNA, each nucleated cell has about six feet of DNA, and it's really fun to just watch that ball up and then be used for a really fancy assay like sequencing. So this is just some examples. I won't go into each of these techniques individually because that will probably take an entire night in and of itself, but we can go on and we can look at things like epigenetic effect. So this gel up here is from our Prader-Willi Angelman assay that looks for differences in methylation at a particular region on chromosome 15. This is an example of array comparative genomic hybridization where we can look for copy number changes. So we can look for very small deletions and duplications within the genome. We can count the number of copies of a particular gene using quantitative PCR or quantitative RT-PCR. 

We commonly use southern blotting. We use southern blotting for fragile X testing. And then of course there's sequencing where you look base for base at a particular region of interest. And then we do fragment analysis as well. So first and foremost, our goal is to provide the best patient care that we can. So the clinical laboratories, above all, patient care is it what we're about. So our testing is used for treatment and prognosis, stratification into the different studies, we have a lot of studies that the university participates in and we do the clinical testing for those. And then, as part of our academic mission, we do have research projects on the side. So as we learn things, the exciting thing about genetics is we get a lot of results, sometimes, especially the scanning techniques where we don't know exactly what it means. There's something not right and we have an opportunity to really take the next step, in some cases, and really look at is this the gene involved, what does it do and what can we do about it, is this something that other patients might be interested in as well. 

So this brings in new techniques and new testing into the clinical laboratories. So I'm going to start with a few, I'll have a few case reports or case examples of the types of things that we have come through our lab. So the first case is a five-year-old girl who presented to her family practice clinic with leg pain. So in this case there's a lot of other clinical features that I won't get into in depth, but they did a CBC and they found many abnormal B cell lymphocyte blasts. So the diagnosis was B cell acute lymphoblastic leukemia. But now the diagnosis doesn't just end there. So you take it the next step further. They had requested cytogenetics to look for common rearrangements that happen in leukemias so that you can look at different treatment options, you can talk about prognosis and do other stratifications into potential studies and so forth. So we did her chromosomes and this is what we found. Can anybody identify what is wrong with this chromosome? Yeah? >> It looks like in one of the chromosome 14 samples has been a large deletion. 

>> Right. So there's something wrong with the chromosome 14. It looks like there's a whole bunch of it that's gone. Anybody else? >> How about 10. >> 10, chromosome 10, right. So there's something very wrong with chromosome 10. And if you compare the banding of the abnormal 10 with the banding of 14, you can see that those two chromosomes have switched material. So there's a translocation on 10 and 14. There are two other abnormalities. >> 16. 

>> 16. You guys are good. This is good. Right, so 16. 16 has material at the bottom of it. >> 9. >> And 9. Great. You guys are hired. So 16 has some material on the bottom of it that doesn't look quite right, but then when you look at the top of chromosome 9 you can see that 9 and 16 have swapped material. 

So you have two translocations in this karyotype. So then my job is to take it the next step farther. So it's fine to say we have two translocations, but now we need to know are there any important genes at those break points that we can tell right now, have any of these translocation occurred before. Do you have a question? >> What causes translocation? >> What causes translocation? There are probably a lot of things that cause translocation. So you could have mutation that causes abnormal crossing over. You can have breaks and rejoining. It depends, a lot of things depend on where and how you see it. When you see it in an oncology specimen, it can sometimes just be sheer chaos. So there are molecular mutations that happen that cause abnormal swaps. There can be repetitive regions that get deregulated. 

There are all kinds of reasons that translocations happen. But there's not one particular reason why they happen. So there are some, I'll talk about one recurrent translocation in humans, but other than that there's not really a whole lot of recurrent translocation just in general. So there are some that happen over again in specific types of cancer, but those mechanisms aren't really well known. Yeah? >> Are they limited to certain chromosome numbers or types? >> No, they're not really limited to certain chromosome numbers or types. So there are certain cases like chronic myeloid leukemia. So that has a 9-22 translocation. That is the indicator for that particular disorder. But in most cases, there are a whole host of translocations that have been seen in this disorder and that disorder, especially when you start talking about oncology. And I didn't bring any really crazy karyotypes. 

But there are some, like myelomas you can have karyotypes that have 100 chromosomes and all kinds of translocations. Especially solid tumors, you can have chromosomes you can't identify. You know it's chromosomal material but you can't really figure out where the material came from. So in some cases it's kind of predictable, but predictable we don't exactly know why. But it keeps it exciting and it helps us identify, so with all these different types of translocations that helps us identify new genes to go after to see, okay, if we don't physically see a break in a translocation in them, are there molecular things happening that we can't see at a chromosomal level. So if we start going after the genes at the break points, can we identify things earlier and faster so that we can intervene with treatments a little bit sooner. So when I looked at these two translocations, there were four genes of interest. So the 9-16 translocation had the PAX5 gene at the chromosome 9 break point, and it had core binding factor at the chromosome 16 break point. So my hypothesis was with this that this is a translocation that helped solidify it as a B cell lineage. So if this translocation was present, it was going to look like a B cell type abnormality. 

And then there was a 10-14 translocation. And at that translocation we had, okay we're good? There were two genes the BMI gene and then the T cell receptor alpha delta. So the T cell receptor alpha delta we see commonly we have a clinical probe that we use for it involved in T cell disorders, but if it was in conjunction with this BMI-1 gene, could that be just what's fueling the proliferation. So now this is a good hypothesis for the step forward. So we can now take this out of the clinical realm and, of course with the consent of our families, we can take this into the research realm and really look at these genes and try to figure out, make the first step, make sure that those are the genes that are actually involved and then move that out further into the research realm to try to figure out how important are these genes in the disease and can we learn something from the way that they're functioning. So towards that, we were able to make some FISH probes. So we were stepping it up one more notch in resolution. And in this one we did see right off the bat that our T cell receptor has been translocated. So you see here this is a normal cell. The five prime end of the gene and the three prime end of the gene are together. 

And then over here this is a metaphase. You can see the normal chromosome 14 right here, and then you can see the abnormal, that little tiny chromosome 14, the derivative 14 and then the derivative 10. So the three prime end of the gene is now on chromosome 10. So you can see that in both metaphase and in the interphase cells. So the next question was, well we know that TCR is involved in the translocation, so the question is, is the BMI one gene also involved? So we took a look. You can go online and you can find back clones that are specific to your region of interest. You can label them up and you can look to see how your clone reacts in your particular specimen. So we took this RP11 clone that's near the BMI-1 gene, and then we labeled that in blue. And then we looked to see is that with the TCR gene at all. And you can see here, here's the normal 14. 

So we have the red and the green. And here's that derivative 14. That's the driver. So that's what we're hypothesizing is what's driving the disease, what's making the proliferation of all of these cells. And we have the derivative 10 and a normal 10 over here. So you can see that those markers are there. So the next question was looking at that 9-16 translocation. So what about PAX5 and what about this core binding factor? It also has been involved in myeloid disorder translocations as well. So we know that this core binding factor, this MTG16 is a known culprit in leukemias already. So when we look at clones, so these are clones that we purchased. 

We labeled those up. And when you hybridize those to cells, you can see that that PAX5 and that core binding factor are fused in these cells. So that's what's driving, that's what we're hypothesizing is driving that B cell commitment. So now we've take be it to a step where now it has to be handed off for further research, but we have the opportunity, it's exciting to have the opportunity to really look into these disorders more and get more answers to physician and patient questions. So our second case is kind of a common site in a clinical genetics clinic. So often what the physicians over there see are kids with a lot of developmental delay, no real handles, so when they come in the door there aren't any real handles that their family practice doctors have. They just know they're not meeting their milestones, they're having global issues, but after spending, so the clinical geneticists really spend a lot of time asking very specific questions and trying to get some more answers that might not be readily apparent to a family practice doctor. So in this case we have a five-year-old girl, could be any young age, with a history of she's awake at night, very tired during the day. So her sleep cycles are all off. She has a history of self-injurious behavior. 

So like head banging and things like that. And difficulty toilet training. So these might be phenotypes that really wouldn't be striking to anybody but coming into a clinical genetics clinic, the physician thought this sounds a lot like Smith-Magenis syndrome. So what comes to us is I would like to do a FISH test looking for this very specific deletion on chromosome 17P. So we do that test. And what you can see here, we have, again, so the control probe is in green. So this marks these chromosomes as this is chromosome 17. And then you have your region of interest. So the red probe is for that Smith-Magenis syndrome region. And you can see in this patient that that region is in fact deleted. 

So this is a nice case in that we're able to get answers, but often it's a series of the diagnostic odyssey. So we really need to get better at finding more disorders and finding efficient ways to get at answers for families. So the physician was suspicious of this, but it's important to get a very distinct clinical diagnosis through testing for a lot of reasons. So this opens up access for special education opportunities, speech and physical therapy. There are pharmacological management options for regulating that sleep cycle which makes everybody in the family a lot happier when everybody sleeps at night and everybody is awake during the day. Things you learn to appreciate. And then there's surveillance of very specific things about this syndrome that you would look for during healthcare maintenance that you might not look for if you didn't have a very specific diagnosis. So looking at kidney and thyroid function, monitoring for scoliosis and eye changes. So the next case is meant to highlight how different techniques can be complimentary. How when you look in one area, don't find an answer, you look in another area using a little slightly different view or slightly different resolution you can get the answer that you've been looking for. 

So case three, a 20-year-old male. The phenotype is developmental delay, cataract, high myopia, midface hypoplasia, so a very flattened faces, and spondyloepiphyseal dysplasia. So they have vertebral abnormalities. And so this phenotype was very specific. It was a little bit more severe but it was specific for Stickler syndrome. And the physician had ordered sequencing for Stickler syndrome. So you would think sequencing would find everything. It would find the mutation, it would find exactly what you're looking for. But the sequencing came back normal. And so the next question was, okay, well, the phenotype is more severe than what you would expect so let's look at a whole genome survey for copy number analysis. 

So the requested technique was array comparative genomic hybridization. So just to give you a very brief background of the technique. So this is like a giant FISH assay. So you label your test or your patient, we label our patients in green. You label control or reference DNA in red, and then you cohybridize them in equal amounts to a microscope slide that has on it spots that are very specific to all regions of the genome. So instead of doing, looking at one or two regions of the genome, we can look at 44,000 plus regions of the genome. So each of the spots on the slide has lots of little pieces of DNA. So there are 70 base pair strings printed in little spots. So then when you hybridize your patient and your reference DNA to it, you should get yellow. So whatever is in your test should be in your control if everything is normal. 

And when you have a loss or a deletion in your patient, you can don't have that labeled portion in your patient pool. So what you see coming up is reference. So what you see is red. And these slides, these slides are not scanned by the naked eye, they're scanned by a laser and all of that data is put into a computer and then visually represented for you to do the analysis. So in this diagram here with have the patient has a deletion. So what you see is red, and every other area of the genome is yellow because the patient is reference is equal. So in this particular patient, this is the diagram of what we would see coming off of the computer in the lab. So this is of chromosome 12 and you can see that in this patient there's a big deletion on chromosome 12. It's a 2.73 megabase region, but it includes the gene for Stickler syndrome as well as 19 other genes. So the 19 other genes kind of help identify the severity. 

But with the size of the deletion, if you went in and sequenced that gene, what are you sequencing? You're only sequencing the one allele that's there. The other allele is gone. So you would get normal. So it was a nice complement to testing that was already done. So the next few cases that I'm going to talk about really focus more on prenatal genetics. So we do prenatal diagnosis for prognosis. So a lot of times we'll get specimens in for advanced maternal age. So do we see the presence of the common trisomies that are associated with advanced maternal age. So 13, 18, 21, sex chromosome abnormalities. Can there be, if we do find something, is there a medical intervention. 

So sometimes there are medical interventions for these things. And then inheritance. So what is the recurrence risk. So has there been an abnormal ultrasound. If there's been an abnormal ultrasound, you look at the genetics, can we see anything that would give the parents any idea of what recurrence might be. So this example is one that we see often. So we have phenotypically normal potential parents. So the reason for referral, so in this particular case we have a phenotypically normal male, a partner with a history of miscarriage, that happens often, a family history of multiple pregnancy losses. So his mother had multiple miscarriages. A partner with an ongoing pregnancy. 

Three copies of chromosome 13 detected by prenatal FISH, where is that extra 13 coming from? Those types of things. The requested testing is standard chromosome analysis and a peripheral blood specimen. And what you can see here on this patient is a Robertsonian translocation carrier. So can you all see which chromosome? 13. So chromosome 13 has chromosome 14 attached to it head to head. So this individual has all the genetic material he needs to be phenotypically normal, but when you try to force his chromosomes to go through miosis you get nondisjunction and problems. So the reproductive consequences in this case, so there's risk of affected children, so there's risk of trisomy 13 or 14 in this particular case. And then there are affects of uniparental disomy. So it's important that you get the appropriate amount of DNA from mom and the appropriate amount of DNA from dad. If you don't have that exact mix, and it is chromosome specific so there are some chromosomes that are more affected than others, but if you don't get that mix you can end up with a phenotypic effect because you have too much of your genome that came from mom. 

Even though you have the right number of chromosomes. And then what is the risk to other family members? So is this translocation new in this individual or did they inherit it in a balanced fashion from a parent? Do they have carrier parents? Are there potential siblings that might be affected as well? And it also offers the opportunity for preimplantation genetic diagnosis. So now if we have a very specific abnormality that we can look for, we can look for that in the preimplantation period to increase the chances for a successful pregnancy. The next example is, again, a phenotypically normal pregnant female. The reason for referral is a fetus with growth retardation. So the fetus isn't growing as fast as it should. So the request was for FISH for 13, 18, 21, X and Y, so or standard prenatal panel looking there. And then chromosome analysis. So those are often paired because the FISH analysis doesn't look for physical rearrangements, it just looks for presence or absence or too many of a particular chromosome. So we did the prenatal panel. 

And you can see over here these are the probes for chromosome X and 18. So you can see that this baby is a girl. And then on the right you have the probes for chromosome 13 and 21. So it looks like there's the right number of chromosomes, at least with regard to the common trisomies. When with you look at the chromosomes, we find too many. So you can see at 22 there is this little extra chromosome. So immediately the technologists in the lab suspect Emanuel syndrome. So this looks like the product of an individual who has a common recurrent translocation that's, like the only recurrent translocation in humans, between chromosome 11 and 22. So when this looks like the derivative 22 from that translocation. So just to confirm this, because the banding is not exactly descript on those tiny chromosomes, so we do some extra testing just to make sure that the material on that chromosome does in fact come from chromosome 11 and chromosome 22. 

And through a series of FISH tests, we do, in fact, see that here we see what's in red is 11Q, so it's that long arm. You have your two normal 11s in this individual, but then that little tiny chromosome has 11 material in it. And then when looking for chromosome 22, you can see that it also has 22 material. So it is, in fact, from that 11-22 translocation. So getting that extra chromosome, in this particular case, happens because of three-to-one segregation. So those of you who enjoy looking at segregation, it's always kind of fun to see how chromosomes can potentially segregate. So in this case the segregation usually happens in a three-to-one fashion. So you end up with that extra derivative chromosome. So the 11-22 translocation is a recurrent translocation in humans. The unbalanced segregants are usually from a three-to-one segregation, leading to 47 chromosomes with that extra derivative chromosome. 

So this is just an example, it's not as clear as I'd like it to be, but of what the carrier parent would look like. So they, too, have all the genetic material to be phenotypically normal, but when you try to force their chromosomes to go through miosis, to make gametes, things run amok. And these types of things we see at a reasonable rate when we do fertility testing, for instance. So approximately one in 500 people phenotypically normal has some kind of chromosomal abnormality that would generally go undetected, and then for fertility reasons they do chromosomal work up and we'll find a balanced translocation. And so then that's an answer that you can then look for and that's where genetic counseling comes in and we talk about what the possibilities are there. So the importance of the diagnosis. So when we see translocations like this you can anticipate the medical needs at birth. So in this particular case it's involved with congenital heart disease. They can have kidney abnormalities, diaphragmatic hernias, talking about inheritance. So where did the chromosome come from? How far back can we track it? Are there other people who might be at risk? And then to really get an idea of what the developmental expectations are. 

So with a can we do to give this patient the best that we can? >> What are the diagnoses of these? >> So what's all involved with it? And that's another important thing. So a lot of people go online and see what's involved with Emanuel syndrome, and you'll find a whole list of things. Any genetic disorder, the Internet is really kind of a scary place to go. That's where you go to your genetic counselors and your clinical geneticists because they give you a list of a million things because this individual has this genotype, this individual has this genotype but they also have this phenotype. So the phenotype kind of builds, builds, builds but not one person has all of those things. So there can be a whole host of things to look at. But really the things that you look at are, in this particular case their concerns are with congenital heart disease, kidney abnormalities. [INAUDIBLE] Yeah. So this is a photo that I took from GeneReviews. So you can go to: genereviews.org It's a great place to go to get genetic information, and it just shows a little bit about, you can describe a phenotype in words but looking at pictures will really give you an idea of what things can be different in a phenotype and what things can be the same. 

So looking at phenotype in people is a lot different than looking at phenotype in other models. So there are a lot of things that are different but a lot of things that you can really point out as being similar or the same. And then to try to make those genotype/phenotype correlations, and that's really where a lot of the research in human genetics is going right now to try to identify what those genotype/phenotype correlations are because a lot of times it really isn't well characterized. But those are the questions. Those are the questions that patients and parents want to know. What can I expect? What can I do? How can I fix it? And towards that we have to do more and more work on the abnormalities that we do find to try to identify gene function, how it works in an organism and how it affects development. There's a whole bunch of things that won't be solved by necessarily... >> I have a question. >> Yeah. >> What are the diagnostics of these types of children? >> So these children have exactly that karyotype. 

So they have that extra derivative 22. So they're exactly-- if I go back, can I go back? So they have the karyotype, can I go back? They have this. So when you see Emanuel syndrome, you will see chromosomes that look like this. But the individuals will look a little bit different because they all have a different background, but when you look at just the chromosomes, they'll have the extra derivative 22 from that 11-22 translocation.  So, like I had mentioned before though, too often things are apparently normal. So a lot of work needs to be done. So we have new mutations to be discovered, technology costs need to be reduced. So from the clinical laboratory perspective we want to offer our patients the best and not have to pay a lot of money to get an answer. And then to come up with some testing algorithms that are really efficient. So you can, with the tests that are available, they are very specific. 

So can we narrow it down faster and better to get answers for our patients? So that is all I have this evening. [APPLAUSE] Thank you very much.