U.S. flag
      An official website of the United States government, Department of Justice.
        Here's how you know
      Here's how you know**Official websites use .gov**
            
            A **.gov** website belongs to an official government organization in the United States.
      **Secure .gov websites use HTTPS**
            
            A **lock** (
            A locked padlock
            ) or **https://** means you’ve safely connected to the .gov website. Share sensitive information only on official, secure websites. Menu

      
        close

            
            
                                              About NIJ
        
                    
  
          About NIJ
      
    
          The NIJ Director
          How NIJ Is Organized
          Mission and Guiding Principles
          Research and Development Process
          Strategic Framework
          Assessments of NIJ
          Annual Reports & Reports to Congress
          Staff Directory
          Funding & Awards
        
                    
  
          Funding & Awards
      
    
          Current Funding
          Forthcoming Funding
          Expired Funding
          Awards: Listing of Funded Projects
          Guidance for Applicants and Awardees
          Fellowship & Student Programs
          Travel Scholarships
          NIJ Challenge Program
          Library & Multimedia
        
                    
  
          Library & Multimedia
      
    
          NIJ Journal
          Podcasts
          Publications Listing
          Multimedia Listing
          Rated Practices on CrimeSolutions
          Rated Programs on CrimeSolutions
          Software & Databases
          Datasets
          Patents
          Topics
        
                    
  
          Topics
      
    
          Article Listing
          Corrections
          Courts
          Crime Prevention
          Crimes
          Drugs and Crime
          Equipment and Technology
          Forensic Sciences
          Youth Justice
          Justice System Reform
          Law Enforcement
          Tribal Crime and Justice
          Victims of Crime
          Events
        
                    
  
          Events
      
    
          NIJ 2024 Research Conference
          Scheduled Events
          On-Demand Events
          Research Meetings & Workshops
          NIJ-Hosted Online Training

                            
                                About
          Contact Us
          Subscribe
          Topics A-Z
                  Share
FacebookXLinkedInRedditPinterestEmail Home
                            Library & Multimedia
 MPS Analysis of the mtDNA Hypervariable Regions on the MiSeq With Improved Enrichment
NCJ Number
              252233

Author(s)
          Mitchell M. Holland, Laura A. Wilson, Sarah Copeland, Gloria Dimick, Charity A. Holland, Robert Rever, Jennifer A. McElhoe
Date Published

July 2017
Length
              13 pages
Annotation
              This article describes an improvement in the enrichment of massively parallel sequencing (MPS) analysis of the mtDNA hypervariable regions on the MiSeq.
Abstract
              The non-coding displacement (D) loop of the human mitochondrial (mt) genome contains two hypervariable regions known as HVR1 and HVR2  that are most often analyzed by forensic DNA laboratories. The MPS protocol from Illumina (Human mtDNA D-Loop Hypervariable Region protocol) uses four sets of established PCR primer pairs for the initial amplification (enrichment) step that spans the  hypervariable regions. Transposase adapted (TA) sequences are attached to the 5’-end of each primer, allowing for effective library preparation prior to analysis on the MiSeq, and AmpliTaq Gold DNA polymerase is the enzyme recommended for amplification. The amplification conditions were modified by replacing AmpliTaq Gold with TaKaRa Ex Taq HS, along with an enhanced PCR buffer system. The resulting method was compared to the recommended protocol and to a conventional on-MPS approach used in an operating forensic DNA laboratory. The modified amplification conditions gave equivalent or improved results, including when amplifying low amounts of DNA template from hair shafts, which are a routine evidence type in forensic mtDNA cases. Amplification products were successfully sequenced using an MPS approach, addressing sensitivity of library preparation, evaluation of precision and accuracy through repeatability and reproducibility, and mixture studies. These findings provide forensic laboratories with a robust and improved enrichment method as they begin to implement the D-loop protocol from Illumina. Given that ExTaq HS is a proofreading enzyme, using this approach should enable improved analysis of low-level mtDNA heteroplasmy. 29 references (Publisher abstract modified)
      
  
          Additional Details
Agencies
        NIJ-Sponsored


  
  
        
  Award
        Assessing the Effects of DNA Damage on Next Generation mtDNA Sequencing
    
  
  
Date Published: July 1, 2017
    
          Downloads
        
                      HTML

Side navigation
  

        

  
  
              NIJ Journal
    
      
            
            
            
      Podcasts
    
      
            
            
            
      Publications Listing
    
      
            
            
            
      Multimedia Listing
    
      
            
            
            
      Rated Practices on CrimeSolutions
    
      
            
            
            
      Rated Programs on CrimeSolutions
    
      
            
            
            
      Software & Databases
    
      
            
            
            
      Datasets
    
      
            
            
            
      PatentsRelated Topics
        
                Law enforcement
        Justice system
        Forensic sciences
        Grants and funding
        Research



  
Similar Publications
        
                      White Paper – Information Sharing Between Medicolegal Death Investigation Offices And Organ And Tissue Recovery Organizations
                  Technical Note – Analysis Of Marijuana And Marijuana Products
                  Technical Note – Ultra-High Performance Liquid Chromatography Photo Diode Array Ultraviolet Single Quadrupole Mass Spectrometry (UHPLC-PDA UV-MS)
 Top

      National Institute of Justice

        
        
                        About NIJ
                    About NIJ
      
    
                  How NIJ Is Organized
      
    
                  NIJ Director
      
    
                  Research Disclaimer
      
    
                  Funding & Awards
                    Funding & Awards Home
      
    
                  Awards Listing
      
    
                  Current Funding
      
    
                  Guidance for Applicants and Awardees
      
    
                  Resources
                    Events
      
    
                  Podcasts
      
    
                  Publications
      
    
                  Topics
      
    
                  Multimedia
      
    
                  Reuse Policy
      
    
                  Contacts
                    Contact NIJ
      
    
                  Staff Directory
      
    
                  Submit a FOIA Request
      
    
                  File a Civil Rights ComplaintFacebook
          
                      X
          
                      YouTube
  

      810 7th St. NW, Washington, D.C. 20531
      
              Secondary Footer link menu
            Accessibility
              Plain Language
              Privacy Policy
              Legal Policies and Disclaimer
              No FEAR Act
              Freedom of Information Act
              USA.gov
              Justice.gov