Genome Assignment

Chromosome Maps

1. WHICH CHROMOSOME DID YOU CHOOSE? Chromosome 10 2 & 3. STATE THE NUMBER OF GENES AND BASE PAIRS ON THE CHROMOSOME YOU CHOSE. Contains over 1400 genes and 130 million base pairs 4. LIST ONE GENE WHICH IS LOCATED ON THIS CHROMOSOME. PAHX gene 5. STATE THE NORMAL FUNCTION OF THE GENE YOU LISTED IN #4. The normal function of PAHX is the production of PAHX which is an enzyme that is needed for metabolism of phytanic acid.

Introduction to BLAST

6. WHAT IS THE TOP SEQUENCE DESCRIPTION MATCH FOR YOUR QUERY SEQUENCE? Homo sapiens CFTR promoter region (LOC111674463) on chromosome 7 7. IS THIS A SEQUENCE FOR A PROTEIN OR ANOTHER PART OF THE GENE? IF IT IS “ANOTHER PART OF THE GENE”, EXPLAIN ITS PURPOSE. It is for another part of the gene, specifically the promoter region of the cystic fibrosis transmembrane conductance regulator (CFTR) gene. The purpose of this sequence is to stimulate cAMP binding, activating transcription factors, and CCAAT/enhancer binding proteins. 8. WHAT DOES THE ENCODED PROTEIN ASSOCIATED WITH THE ABOVE SEQUENCE DO IN THE BODY? Search the PubMed site at www.ncbi.nlm.nih.gov/entrez/query.fcgi?DB=pubmed to answer this question. Under “Article types,” choose “Review”. CITE THE PAPER YOU USED TO DETERMINE THE PURPOSE OF THE ENCODED PROTEIN. This encoded protein has many functions. The CFTR gene promoter helps to maintain levels of CFTR gene expression. Additionally, it is an epithelial chloride channel and part of the ATP-binding cassette (ABC) membrane transporter family. McCarthy VA, Harris A. The CFTR gene and regulation of its expression. Pediatr Pulmonol. 2005 Jul;40(1):1-8. doi: 10.1002/ppul.20199. PMID: 15806593. 9. A MUTATED FORM OF THIS GENE IS RESPONSIBLE FOR A WELL-STUDIED DISEASE. WHAT IS THAT DISEASE? A mutated form can lead to the disease Cystic fibrosis 10. ON WHAT CHROMOSOME IS THE GENE LOCATED? This is found on Chromosome 7 11. WHAT SPECIES (STATE THE SCIENTIFIC NAME) OTHER THAN HOMO SAPIENS ALSO HAS A 100% IDENTITY (Ident) FOR THIS SEQUENCE? USE THE TOP SEQUENCE LISTED, BUT DO NOT USE THE PREDICTED SEQUENCES. Pongo abelii 12. WHAT IS THE COMMON NAME FOR THIS SPECIES? Sumatran orangutan

13. DOES IT SURPRISE YOU THAT THIS SPECIES ALSO HAS A 100% SIMILARITY IN IDENTITY? WHY OR WHY NOT? No, because orangutans among many other species are closely related to homo sapiens. 14. DESCRIBE THE FIRST MATCH THAT HAS LESS THAN 100% QUERY COVER BUT IS NOT PREDICTED OR HOMO SAPIENS. STATE THE SCIENTIFIC AND COMMON NAMES. Saimiri boliviensis boliviensis, also known as the Black-capped squirrel monkey is the first query at less than 100%. 15. HOW MANY GAPS OCCUR BETWEEN THE TWO SEQUENCES (THE ONE YOU SUBMITTED AND THE FIRST ONE THAT HAS LESS THAN 100% QUERY COVER)? Gaps: 1/119 16. WHAT IS A GAP IN SEQUENCE ALIGNMENTS? According to Wikipedia, gaps are inserted spaces between residues that allow identical or similar sequences to align. FOR EACH, STATE WHAT THE GENE IS (#17-20). Give the description of the gene and gene product. You do not need to state the organism source. 17. NM_145556 TAR DNA binding protein (Tardpb); it enables RNA polymerase II sequence specific DNA binding activity and pre-mRNA intronic binding. 18. NM_013444 Homo sapiens ubiquilin 2 (UBQLN2), mRNA; it is a gene that codes for a ubiquitin-like protein (ubiquilin) and has been shown to bind ATPase and function in ubiquitination machinery. 19. NM_001010850 FUS RNA binding protein; it encodes protein component of hnRNP – heterogeneous nuclear ribonucleoprotein complex which is involved in splicing and export 20. KJ174530 Superoxide dismutase 1 (SOD1); is a gene that codes for a protein that binds copper and zinc ions and helps destroy free superoxide radicals found in the body 21. Search Google to answer the following: WHAT DISEASE IS ASSOCIATED WITH MUTATIONS OF THE GENES REFERENCED IN #17-#20? WHAT IS A “COMMON NAME” OF THE DISEASE? Mutations in the above genes leads to Amyotrophic lateral sclerosis (ALS) – Lou Gehrig Disease 22. WHAT IS GENBANK? (You can do an Internet search to find this information.) It is an open access database created by three organizations that exchange data daily and officially update every two months.

Introduction to Swiss-Prot to Study Protein Sequences

23. WHAT IS cDNA? How can we obtain cDNA in the lab?

Stands for complementary DNA. It is obtained by using reverse transcriptase in vitro with specific mRNA

Enter the following sequence: ACATTTGCTTCTGACACAATTGTGTTCACTAGCAACCTCAAACAGACACCATGGTGCATCTGACTCCTGAGGAGAAGTCTGCCGTTACTGCCCTGTGGGGCAAGGTGAACGTGGATGAAGTTGGTGGTGAG

GCCCTGGGCAG

24. USING THE SAME PROGRAM YOU USED IN THE INTRODUCTION TO BLAST ABOVE, WHAT IS THE SEQUENCE MATCH?

Homo sapiens partial HBB gene for hemoglobin beta chain, exon 1, isolate 04593664

Now access the Expasy translate tool at https://web.expasy.org/translate/.

25. WHAT IS AN OPEN READING FRAME?

An open reading frame are “spans of DNA sequence between the start and stop codons”

26. ALL OF THE PROPOSED OPEN READING FRAMES (HIGHLIGHTED IN RED) START WITH THE AMINO ACID “M”. FROM WHAT YOU KNOW ABOUT POLYPEPTIDES, WHAT IS “M”?

M most likely stands for methionine, which is the amino acid coded by the start codon AUG.

27. WHICH 5’ TO 3’ FRAME IS MOST LIKELY TO BE AN OPEN READING FRAME? WHY DID YOU CHOOSE THAT FRAME?

I choose the 5’-3’ Frame 3 because the open reading frame is the longest of all the options and thus seems more likely to code a full protein.

Amino Acid Sequence Comparisons

Person 1/Sequence 1:

MGAPACALALCVAVAIVAGASSESLGTEQRVVGRAAEVPGPEPGQQEQLVFGSGDAVELSCPPPGGGPMGPTVWVKDGTGLVPSERVLVGPQRLQVLNASHEDSGAYSCRQRLTQRVLCHFSVRVTDAPSSGDDEDGEDEAEDTGVDTGAPYWTRPERMDKKLLAVPAANTVRFRCPAAGNPTPSISWLKNGREFRGEHRIGGIKLRHQQWSLVMESVVPSDRGNYTCVVENKFGSIRQTYTLDVLERSPHRPILQAGLPANQTAVLGSDVEFHCKVYSDAQPHIQWLKHVEVNGSKVGPDGTPYVTVLKTAGANTTDKELEVLSLHNVTFEDAGEYTCLAGNSIGFSHHSAWLVVLPAEEELVEADEAGSVYAGILSYGVGFFLFILVVAAVTLCRLRSPPKKGLGSPTVHKISRFPLKRQVSLESNASMSSNTPLVRIARLSSGEGPTLANVSELELPADPKWELSRARLTLGKPLGEGCFGQVVMAEAIGIDKDRAAKPVTVAVKMLKDDATDKDLSDLVSEMEMMKMIGKHKNIINLLGACTQGGPLYVLVEYAAKGNLREFLRARRPPGLDYSFDTCKPPEEQLTFKDLVSCAYQVARGMEYLASQKCIHRDLAARNVLVTEDNVMKIADFGLARDVHNLDYYKKTTNGRLPVKWMAPEALFDRVYTHQSDVWSFGVLLWEIFTLGGSPYPGIPVEELFKLLKEGHRMDKPANCTHDLYMIMRECWHAAPSQRPTFKQLVEDLDRVLTVTSTDEYLDLSAPFEQYSPGGQDTPSSSSGDDSVFAHDLLPPAPPSSGGSRT

Person 2/Sequence 2:

MGAPACALALCVAVAIVAGASSESLGTEQRVVGRAAEVPGPEPGQQEQLVFGSGDAVELSCPPPGGGPMGPTVWVKDGTGLVPSERVLVGPQRLQVLNASHEDSGAYSCRQRLTQRVLCHFSVRVTDAPSSGDDEDGEDEAEDTGVDTGAPYWTRPERMDKKLLAVPAANTVRFRCPAAGNPTPSISWLKNGREFRGEHRIGGIKLRHQQWSLVMESVVPSDRGNYTCVVENKFGSIRQTYTLDVLERSPHRPILQAGLPANQTAVLGSDVEFHCKVYSDAQPHIQWLKHVEVNGSKVGPDGTPYVTVLKTAGANTTDKELEVLSLHNVTFEDAGEYTCLAGNSIGFSHHSAWLVVLPAEEELVEADEAGSVYAGILSYRVGFFLFILVVAAVTLCRLRSPPKKGLGSPTVHKISRFPLKRQVSLESNASMSSNTPLVRIARLSSGEGPTLANVSELELPADPKWELSRARLTLGKPLGEGCFGQVVMAEAIGIDKDRAAKPVTVAVKMLKDDATDKDLSDLVSEMEMMKMIGKHKNIINLL

GACTQGGPLYVLVEYAAKGNLREFLRARRPPGLDYSFDTCKPPEEQLTFKDLVSCAYQVARGMEYLASQKCIHRDLAARNVLVTEDNVMKIADFGLARDVHNLDYYKKTTNGRLPVKWMAPEALFDRVYTHQSDVWSFGVLLWEIFTLGGSPYPGIPVEELFKLLKEGHRMDKPANCTHDLYMIMRECWHAAPSQRPTFKQLVEDLDRVLTVTSTDEYLDLSAPFEQYSPGGQDTPSSSSSGDDSVFAHDLLPPAPPSSGGSRT

Submit the sequences for comparison.

28. DO YOU SEE ANY DIFFERENCES BETWEEN THE TWO AMINO ACID SEQUENCES? (Look for the absence of an asterisk, which indicates the same amino acid in both sequences.)

Yes, there are differences between the two amino acid sequences.

29. IF YOU SAW DIFFERENCES, WHAT WERE THEY?

In the main comparison, residue 361 has one difference, where Person 1 codes for G and Person 2 codes for R.

Return to the BLAST home page (http://blast.ncbi.nlm.nih.gov/Blast.cgi). Run a PROTEIN BLAST search to identify the polypeptide which you have been analyzing. (You may use either sequence.)

30. WHAT IS THE FUNCTION OF THIS PROTEIN?

Fibroblast growth factor receptor 3 (FGFR3). Its function is bind acidic and basic growth hormones and develop and maintain bones.

31. WHAT HUMAN DISEASE IS CAUSED BY A MUTATION IN THIS GENE?

Mutations lead to craniosynostosis and types of skeletal dysplasia.

32. REFLECT ON ONE THING THAT YOU LEARNED FROM DOING THIS ASSIGNMENT. Please be honest. If you didn’t learn anything, admit it…

This taught me a range of specific things about genetic studies. I did not previously know about things like the GenBank, open reading frames, or many of the genes and proteins listed in this assignment. Understanding each of these gave me a better understanding of genetic libraries and how they are used.

By: Joy Brewer