PV92+/-Alu polymorphism in Humans (Lab Report Sample)

Analysis of PV92+/-Alu polymorphism in Humans
Bio 301 Molecular Biology and Biotechnology Lab 5
Professor:
Dec , 2021
Abstract
This lab purposes to analyze and understand the bioinformatics of Alu transposable element at the PV92 locus of the chromosome 16 of the human genome. Alu are primate specific short interspersed retrotransposons. For this lab, in silico PCR simulation was done using specially designed reverse and forward primers of about 50 base pairs flanking both sides of Alu insert region. The NCBI BLAST made this possible, resulting to a 415 base pairs PCR product that on further analysis aligned with the CDH13 gene belongin to the superfarmily of the catherin proteins. Finally, BLAST search for PV92 containing an Alu insertion revealed Homo sapiens sequence ID: AF302689.1 with two separate matching sequences in the NCBI database.
Introduction
The Alu elements are repetitive sequences of DNA belonging to short interspersed elements class of Non-LTR Retrotransposons (Lankenau & Volff, 2009). They are 315 bp in length, contributing about 10% the human genome. Alu elements transcription into into RNA is aided by RNA polymeraseIII ("Transposable elements, inflammation, and neurological disease," 2021). The reverse transcriptase then converts the RNA into a double stranded DNA prior to transposition into new location of the genome.   Due to their nonautonomousity, as all SINEs, Alu elements lack the ability to make DNA copies or to integrate in new locations in the genome. For better functionality, Alu rely on LINE L1 (Lankenau & Volff, 2009). Most Alu elements are approximately 300 base pairs long, with considerable sequence variation. The PV92 locus Human-specific Alu insertion in the chromosome 16 is therefore termed as a dimorphic even ("Transposable elements, inflammation, and neurological disease," 2021) t.
Methodology
NCBI BLAST page was searched followed by a click of Human link on the resulting page. Under Database, GRCh38 p13 reference assembly top level was selected in the drop-down menu for Homo sapiens. In a new Window, the Blackboard Document for PV92 primers.txt was Copy/pasted while in FASTA format into the “FASTA Sequences(s)” box on the Nucleotide BLAST page.
On the Program Selection, the “Optimize for” box was confirmed to shows “Highly similar sequences” On the BLAST line, the box “Show results in a new window.” Was clicked, on the Algorithm parameters, Short queries deselected, Word size set to 24 and BLAST search initiated. Once BLAST Results Page loaded, the “Back to Traditional Results Page” was clicked and the report page printed. The “download” on the top of the blast page was clicked to get Reverse Primer simultaneously with the Forward Primer and select “Hit Table(csv)”
On BLAST Report output page, Graphics was clicked and Homo Sapiens Cadherin 13 typed in the search box. On first BLAST Report output page a click on “Genome Data Viewer” was made and once CDH13 window appeared, the cursor was hovered over the circles and the information recorded.
The PV92 Primer based BLAST analysis was then repeated once more and the Accession number of the GRCh38 Primary Assembly hit recorded. Back on the Nucleotide BLAST box, a general BLAST search was started, Accession Number was entered in Query Sequence box and the nucleotide range values of the PCR segment entered from the highest to the lowest. And the search selected for others. BLAST was clicked and then searched for hits.
Results
https://www.ncbi.nlm.nih.gov/gene/1012
https://www.ncbi.nlm.nih.gov/genome/gdv/browser/blast/?id=NC_000016.10&alignid=Query_4678&rid=F286A2VN016
Discussion
The BLAST found the sequence alignment to be on the Human Chromosome 16. The PV92 forward primer aligned to CDH13 (catherin 13) gene with an excellent 100% identity hit. Additionally, it scored an E-value of 5.4e-5. Based on the range of the two primers, the PCR product produces was 415 base pairs long (82878610-82878195) ("CDH13 cadherin 13 [Homo sapiens (human)] - Gene - NCBI," n.d.).
The catherin 13 gene was found to belong to the superfarmily of the catherin proteins which are mainly found near the surface of the plasma membrane. Instead of a transmembrane domain, it is anchored by a GPO moiety and hence very unlikely to be a cell to cell adhesion protein. Furthermore, it negatively regulates the growth of axons as well as protecting the cells of