Electropherograms alleles, missing alleles, and artifacts:


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Electropherograms alleles, missing alleles, and artifacts: Electropherograms are plots obtained from electrophoresis automatic sequencing. Attachment 1 Attachment 2 ATTACHMENT PREVIEW Download attachment 10Electropherogram.pdf Electropherogram 1: Electropherogram 2 Electropherogram 3 Electropherogram 4 Electropherogram 5 Electropherogram 6 Electropherogram 7 Electropherogram 8 Electropherogram 9 Electropherogram 10 ATTACHMENT PREVIEW Download attachment Electropherogram.pdf Week 8: Data Packs Electropherogram 1 1. (2) Electropherogram 1 is missing allele designations for the sample. Compare to the ladder and determine allele designations manually. D3S1358 2. (1) D16S539 Amelogenin TH01 TPOX CSF1PO Below is a miniature copy of just the sample from Electropherogram 1. Circle and name one artifact. Electropherogram 2 1. (3) The OL allele in the blue channel is an artifact. Name the artifact, and explain how you came to this determination. 1 Week 8: Data Packs Electropherogram 3 1. (3) Looking at Electropherogram 3, what kind of conclusions can you make about this profile? Points to think about: Are the STR loci amplified? What do the RFU scales tell you? Did your internal size standard work? Is there anything that you could do to recover additional data? Electropherogram 4 1. (2) In Electropherogram 4, CSF and D7 have visible peaks, but they have not been called. Given that the analysis threshold is 75 RFU, can you explain why the peaks do not have allele designations? 2. (1) List two ways that you can modify this sample’s treatment (through sample preparation and/or reinjection) to ensure that all peaks are called. Be specific. 2 Week 8: Data Packs Electropherogram 5 1. (1.5) The arrows point to an artifact. What is it? 2. (1.5) Electropherogram 5 is reprinted below. ON THIS PAGE, identify one other artifact in the electropherogram. 3 Week 8: Data Packs Electropherogram 6 1. (1.5) This is a COfiler® profile after application of an “analysis method” in GeneMapper but before editing the allele calls. Perform your own analysis to eliminate any allele calls that are artifacts and not actual alleles. Write down the resulting profile, locus by locus, in the table provided below. D3S1358 (1.5) D16S539 Amelogenin TH01 TPOX CSF1PO D7S820 Provide an explanation for the allele calls you eliminated, including the locus and allele call, followed by justification for your change. Example: TPOX, 11: pullup from CSF1PO in green channel Electropherogram 7 1. (3) Review this locus and discuss what you think would cause this genotype. If there are artifacts, how can you correct for them? 4 Week 8: Data Packs Electropherogram 8 2. (1.5) This is an Identifiler® profile after application of an “analysis method” in GeneMapper but before editing the allele calls. Perform your own analysis to eliminate any allele calls that are artifacts and not actual alleles. Write down the resulting profile, locus by locus, in the table provided below. CSF1P0 D7S820 FGA D16S539 TH01 (1.5) D8S1179 D21S11 D2S1338 D3S1358 D18S51 D19S433 TPOX Amelogenin vWA D13S317 D5S818 Provide an explanation for the calls you eliminated, including the locus and allele call, followed by the explanation. Example: TPOX, 11: pullup from CSF1PO in green channel Electropherogram 9 1. (2) How many contributors are in this sample, at a minimum? List the alleles (by locus) that may be shared or have multiple copies. 5 Week 8: Data Packs 2. (1) Give one reason why the alleles vary so greatly in RFU value. How much confidence do you have in the calls of the larger-sized (base-pair) amplicons, based on their peak heights? Electropherogram 10 1. (2) There are multiple uncalled peaks in this electropherogram. Are these true alleles or artifacts? If they are artifacts, what kind are they? 2. (1) If you believe the uncalled peaks to be artifacts and not true alleles, how can these types of artifacts be corrected? 6 Read more

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