Compare human colon cancer gene MLH1 with other genes.
- To use ORF finder to translate DNA sequence to protein sequence in all reading frames. -
- To use blastn, blastp, CD search and blast 2 sequence programs for searching and comparison.

1. Compare MLH1 (answer of assignment 2.6) and mutS (answer of 2.7) sequence.
2. Translate the above two gene sequences to protein sequences.
3. Perform protein sequence homology searching for MLH1 in GenBank. Give the 10 highest hits.
4. Compare human MLH1 protein with MLH1 in M. musculus, R. norvegicus and D. melanogaster. Give the pairwise alignment and % of sequence smility.
5. Search the conserve domain (CD) for MLH1. Give the position of the CD, name of CD and Pfam ID number.
6. Show multiple alignment of MLH1 conserve domain with 5 sequences from the top of the CD alignment.

1. 進入"BLAST 2 Sequences"中，將MLH1及mutS的基因序列輸入進行比較，比較的結果為"No significant similarity was found "。

2. 進入BLAST中的"ORF finder "，將MLH1的核酸序列輸入（以FASTA模式），或是輸入GI，按下"OrfFind"，得到搜尋結果，接著按下"Sixframes"，選擇"3 Fasta protein"並按下"View"。便能得到蛋白質序列。 搜尋結果

用同樣方法，可找到MutS的蛋白質序列。 搜尋結果

3. 進入BLAST中的"Standard protein-protein BLAST [blastp]"，將MLH1的蛋白質序列輸入，經過"BLAST!"及"Format!"後得到許多相似的蛋白質序列，取最前面十個即是相似度最高的序列。 Top10Hits

4. 分別將M. musculus、R. norvegicus及D. melanogaster的蛋白質序列輸入"BLAST 2 sequences"中，皆與MLH1比較，得到比較結果。 序列一 序列二 序列三

M. musculus：Identities = 145/151 (96%), Positives = 150/151 (99%)

R. norvegicus：Identities = 649/758 (85%), Positives = 696/758 (91%), Gaps = 3/758 (0%)

D. melanogaster：Identities = 335/751 (44%), Positives = 456/751 (60%), Gaps = 94/751 (12%)

5. 進入BLAST中的"Search the Conserved Domain Database using RPS-BLAST"，將MLH1的蛋白質序列輸入，經過"BLAST!"，得到CD的搜尋結果。 position of the CD：147~327 name of CD：DNA_mis_repair, DNA mismatch repair protein;mutL/hexB/PMS1 family Pfam ID number：01119 網站資料(一) Link

6. 在搜尋multiple alignment的選項中，選擇"up to 5"和"from the top of the CD alignment"，按下"Add"，就能得到"CD Browser"。 搜尋結果