Dr. Mitchell M. Holland

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  www.post-gazette.com/pg/09041/947946-298.stm - [Cached Version]
  Published on: 2/10/2009    Last Visited: 2/10/2009  

  The two-year program -- one of just a handful in the country -- leapfrogs students over entry-level jobs in forensic DNA labs, said Mitchell Holland, associate director of Penn State's forensic science program. Students gain a broader science background than they would if they went straight from getting their bachelor's degree to working in a lab.
  
  "It accelerates their knowledge and preparation and allows them to get a broader view of the entire process," said Dr. Holland.

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  www.sciencemasters.com/PSMProgramList/InstitutionsbyReg - [Cached Version]
  Published on: 9/4/2007    Last Visited: 9/4/2007  

  Forensic Science, Contact: Mitch Holland, Associate Director

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  live.psu.edu/story/40779 - [Cached Version]
  Last Visited: 8/12/2009  

  Mitchell Holland
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  Mitchell Holland
  
  A Penn State DNA researcher who helped to reveal the mystery identity of the Vietnam Unknown Soldier, along with the soldier's sister -- herself a military officer -- will tell both the scientific and the personal stories behind the discovery of the soldier's identity during a talk Tuesday at the National Museum of Health and Medicine in Washington, D.C.
  
  Mitchell Holland, a member of the DNA discovery team who now is a Penn State professor of forensic science, and Col.
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  Holland held various positions at the Armed Forces DNA Identification Laboratory (AFDIL) in Rockville, Md., including Scientific Laboratory Director from 1993-2000.

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  www.scubeindia.com/SoftGenetics/genemarker_hid.html - [Cached Version]
  Last Visited: 3/6/2009  

  Developed in collaboration with Mitchell Holland, Ph.D., Associate Director of The Pennsylvania State University's Forensic Science Program. Review Dr. Holland's GeneMarkerHID Poster.

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  www.bioke.com/products/product+information/softgenetics - [Cached Version]
  Published on: 5/31/2007    Last Visited: 5/31/2007  

  Developed in collaboration with Mitchell Holland, Ph.D., Associate Director of The Pennsylvania State University's Forensic Science Program.

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  www.samreesesheppard.org/beacon.html - [Cached Version]
  Published on: 1/26/2000    Last Visited: 9/21/2000  

  And by the time it was the state's turn to challenge Tahir's findings by calling its own DNA expert -- Dr. Mitchell Holland, chief of DNA for the Armed Forces Institute of Pathology in Rockville, Md. -- the state had thought better of it.
  
  Hence they had no DNA expert at all.

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  760 KFMB - [Cached Version]
  Published on: 1/5/2002    Last Visited: 7/30/2003  

  DR. Mitchell Holland: Director of Bode Technology forensic DNA lab, Holland testified that, using both nuclear and mitochondrial DNA, his lab was able to conclusively identify blood and hair found in Westerfield's RV as having come from Danielle.

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  ABRF 2009 : Confirmed Speakers - [Cached Version]
  Published on: 2/7/2009    Last Visited: 9/1/2009  

  Mitchell Holland, Pennsylvania State University

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  Adelaide IMC: newswire/9726 - [Cached Version]
  Published on: 12/11/2004    Last Visited: 2/19/2006  

  "I think they did the right things," said Mitchell Holland, a professor of forensic science at Pennsylvania State University.

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  Applied Biosystems - BioBeat - U.S. Department of... - [Cached Version]
  Published on: 12/2/2002    Last Visited: 8/21/2003  

  Dr. Mitchell Holland, chief forensic scientist at AFDIL, will lead the efforts to analyze mtDNA from bone fragments of servicemen killed in conflicts this century and supply confirmatory evidence to help identify as many of the previously classified unknown soldiers as is possible.To undertake such a large-scale DNA sequencing operation-one that could potentially involve hundreds of thousands of sequencing reactions-Holland has added ten PE Biosystems ABI PRISM® 377 DNA Sequencers with 96-lane upgrades to his forensics lab in Rockville, Maryland, and equipped the 29 scientists involved in the project with the ultra-sensitive ABI PRISM® BigDye Terminator Cycle Sequencing Kits.
  
  Program Begins at the Punchbowl in Hawaii
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  Then, in a lab at CILHI, forensic scientists cut a specimen of bone from the remains and send a sample of it to AFDIL where Holland's group extracts remaining fragments of DNA from the mitochondria of bone cells.
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  "At AFDIL, we do the mitochondrial DNA analysis, then we send our results to CILHI, and it is their responsibility to combine this information with other circumstantial data in order to recommend an identification," says Holland.
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  "If you have a dental match, then you do not need mtDNA analysis," notes Holland, who has overseen all of the DNA analysis projects.
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  "We hope that number will go up as the word gets out there and more family members come forward and provide a sample," notes Holland.
  
  Production-Scale Human Identification
  
  AFDIL has been using molecular techniques to identify people since 1991.However, in the last five years-since they began using mtDNA analysis more regularly-their scale of operation has increased tremendously from making just a few identifications eight years ago to now processing hundreds of skeletal elements (i.e., bones and teeth) and assisting in the identification of greater than 100 individuals per year.
  
  To meet the DNA sequencing demands of such a large-scale human identification project, Holland has increased the number of forensic research scientists working in his lab from 32 to 40; 29 of these scientists are working solely on projects that require mtDNA analysis.A few of these scientists were hired specifically to help develop the family reference database.
  
  As part of AFDIL's recent expansion, Holland has also added 10 new ABI PRISM® 377 DNA Sequencers with 96-lane upgrades.According to Holland, eight of these new sequencers are dedicated to performing sequencing related to the analysis of mtDNA samples.They are key to simplifying the creation of mitochondrial DNA profiles.
  
  BACK TO TOP
  
  To make a mitochondrial DNA profile, Holland's group first performs the polymerase chain reaction (PCR) to pluck relevant mtDNA base-pair sequences from a pool of DNA extracted from a blood or bone sample.
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  When making mtDNA profiles, scientists in Holland's lab encounter DNA samples that range in quality from pristine-such as those extracted from whole blood samples-to extremely degraded samples; those from older sets of skeletal remains.The approach the scientists take when making profiles differs depending on the quality of the DNA sample.
  
  "With pristine DNA, it is possible to amplify the entire mtDNA control region (approximately 1200 nucleotides long), then bombard it with multiple sequencing primers to generate the sequence data," notes Holland.When all relevant mtDNA fragments or amplicons have been sequenced, a pattern of base-pair sequences emerges that becomes an individual's mitochondrial DNA profile.
  
  To visualize the sequence of DNA bases that make up an individual's mtDNA profile, the scientists load the sequencing products in one of the 96 lanes of the 377 DNA sequencer.Like an automated word processor, the 377 system decodes the strings of DNA bases in each lane and spells out a long sentence using a four-letter alphabet-the four DNA bases: adenine (A), thymidine (T), cytosine (C), and guanine (G).
  
  "The 96-lane upgrade has greatly increased our throughput, and the 377 system's new neural net tracking system does a very good job of tracking lanes," notes Holland.
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  There are only a few copies of mitochondrial DNA left in these bones," Holland explains."We need to guard against them being contaminated, so we can extract those few copies and make them available for PCR."
  
  Over time, DNA degrades, so that only small size fragments can be extracted from bone and amplified.To obtain mtDNA profiles from skeletal remains, Holland's group must therefore choose a greater number of small fragments throughout the control region.
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  "Over time, the evolution of sequencing kits has continued to improve the quality of results and allow for more of this work to be done, because you don't have to repeatedly go back and replicate what's been done," Holland says."The BigDye terminators have increased the quality of our sequencing data, which reduces the number of reactions that we might have to re-run.Also, the increased sensitivity of the dyes gives us better results from weaker amplicons."
  
  "Our recent upgrade to 377 systems with 96-lanes and our use of the BigDye terminators is increasing our throughput by about 25 percent," Holland continues, "But we anticipate that they will have an even larger impact on this project as we address other bottlenecks in mtDNA analysis."
  
  Customized Software Helps Analyze Data
  
  Extraction of DNA from bone is not the only bottleneck in the identification process.According to Holland, the analysis of resulting sequencing data is also an extremely time consuming process.To streamline data analysis, AFDIL has developed custom mtDNA sequence analysis software with GeneCodes and Wagner Associates that recognizes whether a sequence is a forward or reverse sequence and where it is in the control region.It then aligns sequences based on the mtDNAs genome numbering system, will identify differences from a consensus sequence, and provide a mtDNA profile or an overall case consensus sequence.
  
  Scientists at AFDIL have just finished validating this software and they are planning to first use it to generate the family reference database, which will help with their efforts to identify servicemen missing in the Korean War.
  
  "In the future, when we have cases in which there is no information associated with a set of remains, we'll use the reference database we're developing to provide hits that suggest a presumptive identification," Holland says.

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