GENETIC CHAPTER 23 GENOMIC II: FUNCTIONAL GENOMICS, PROTEOMICS AND BIOINFORMATICS

ALTERATIONS THAT AFFECT THE PROTEOME -.... .... (CHAP 12): ** -Most important alteration that occurs in .... -A single ... ... is spliced into more than one version -.... is often cell specific or related to environmental conditions -... EDITING (CHAP 12): ** -Much less ... than alternative splicing -Leads to changes in the ... .... of mRNA after the mRNA is made

-ALTERNATIVE SPLICING -eukaryotes -pre-mRNA -Splicing -RNA -common -coding sequence

CHROMATIN IMMUNOPRECIPITATION -... .... (...) CAN DETERMINE IF .... CAN BIND TO A PARTICULAR REGION OF DNA IN THE ... OF LIVING CELLS: ** -.... in living cells that are binding to specific sites in the DNA are covalently cross-linked to those DNA sites using formaldehyde -Cells are lysed and .... broken into small pieces -.... is used to precipitate the protein of interest -DNA is chemically ... from the cross-links -The DNA is .... with PCR -The sequence of the DNA is identified directly or by using it as a probe on a microarray (.... .. ....)**

-CHROMATIN IMMUNOPRECIPITATION (ChIP) -PROTEINS -CHROMATIN -Proteins -DNA -Antibody -freed -amplified -(ChIP-chip assay)

MASS SPECTROMETRY IS USED TO IDENTIFY PROTEINS -THE NEXT STEP IS TO .... A GIVEN SPOT ON A TWO-DIMENSIONAL GEL WITH A ... ...: -A spot is .... out from the gel -The ... is eluted from the gel in a purified form -The amino acid sequence of the protein is revealed via a technique called .... ... ...:** -Two .... are used: -The first measures the ... of a given peptide (that was generated from protein digestion) -The second analyzes the peptide .... it has been digested into even smaller fragments, one amino acid at a time

-CORRELATE -PARTICULAR PROTEIN -cut -protein -tandem mass spectrometry -spectrometers -mass -after

CHROMATIN IMMUNOPRECIPITATION CONT... -TO IDENTIFY .... BOUND BY THE PROTEIN, RESEARCHERS CAN USE: -... -DNA ...

-DNA -PCR -microarray

DNA MICROARRAYS -THE ... ..... ON A MICROARRAY CAN BE EITHER: -.... by PCR and then spotted onto the microarray -Synthesized .... on the microarray itself -A single ... contains tens of thousands of different spots in an area the size of a postage stamp -The relative ... of each spot is known -The .... for making DNA microarrays is quite amazing: -It involves spotting technologies that are quite similar to the way that an inkjet printer works -Once a .... microarray has been made, it is used as a hybridization tool

-DNA FRAGMENTS -Amplified -directly -slide -location -technology -DNA

A MICROARRAY CAN IDENTIFY GENES THAT ARE TRANSCRIBED -RESEARCHERS DEVELOPED A TECHNOLOGY CALLED .... ....(ALSO CALLED GENE ...): ** -This technology makes it possible to .... thousands of genes ....** -A DNA microarray is a small silica, glass or plastic .... that is dotted with many sequences of ....:** -Each of these ... corresponds to a known ...: -These fragments are made synthetically -These sequences of DNA act as .... to identify genes that are transcribed

-DNA MICROARRAYS -CHIPS -monitor -simultaneously -slide -DNA -sequences -gene -probes

INTRODUCTION -THE GOAL OF ... .... IS TO ELUCIDATE THE .. OF GENETIC .... IN A GIVEN SPECIES: ** -In most cases, it aims to understand gene .... -The entire collection of .... that an organism can make is termed the ....** -The goal of ... is to understand the functional roles of the ... of a species:** -It aims to understand the .... among many different proteins -BIOINFORMATICS: ** -Is the analysis of .... information using a mathematical/.... approach: -Often aimed at extracting ... from genetic data

-FUNCTIONAL GENOMICS -ROLES -SEQUENCES -function -proteins -proteome -proteomics -proteins -interplay -biological -computational -information

TWO-DIMENSIONAL GEL ELECTROPHORESIS -THE TECHNIQUE INVOLVES TWO DIFFERENT ... ....EXPERIMENTS: -The first separates by pH/.... interactions (isoelectric focusing) -The second separates by .... (mass)

-GEL ELECTROPHORESIS -charge -size

SEARCHING DATABASES FOR HOMOLOGOUS SEQUENCES -IN GENERAL, A STRONG CORRELATION IS TYPICALLY FOUND BETWEEN .... AND ...: -.... between genetic sequences can be identified by computer programs and databases -A powerful tool for predicting the ... of genetic sequences -In 1990, Stephen Althschul, David Lipman and their colleagues developed an approach called BLAST** -.... ... ... ... ...** -.... between the query sequence and each matching sequence is given an E-value (Expect value): -Represents the number of .... that the match or a better one would be expected to occur purely by random chance in the entire database

-HOMOLOGY -FUNCTION -Homology -function -Basic local alignment search tool -Relationship -times

CREATING COMPUTER DATA FILES -Entering the data into the computer file is done: -.... (that is, typing) -Usually by .... -Reading data directly from a ... ladder -THE GENETIC ... CAN BE .... IN MANY WAYS: -Does a sequence contain a ...? -Where are ... sequences, such as promoters and splice sites? -Does a sequence encode a ....? -If so, what is the ... ...sequence of that polypeptide? -Is a sequence .... to other sequences? -What is the evolutionary .... between two or more genetic sequences?

-Manually -instruments -sequencing -SEQUENCE -ANALYZED -gene -functional -polypeptide -amino acid -homologous -relationship

ALTERATIONS THAT AFFECT THE PROTEOME CONT... -.... COVALENT MODIFICATION:** -.... changes may be necessary to produce a functional protein -... processing; attachment of prosthetic groups, sugars or lipids -Reversible changes that transiently affect the .... of the protein -....; acetylation; methylation

-POSTTRANSLATIONAL -Irreversible -Proteolytic -function -Phosphorylation

TWO-DIMENSIONAL GEL ELECTROPHORESIS IS UESD TO SEPARATE A MIXTURE OF DIFFERENT PROTEINS -ANY GIVEN CELL OF A MULTICELLULAR ORGANISM WILL .... ONLY A SUBSET OF THE .... IN ITS PROTEOME: -THE SUBSET THAT IT MAKES ... PRIMARILY ON THE: -Cell ... -Stage of ... -Environmental .... -A technique in the field of proteomics is two-dimensional ... electrophoresis -It is a .... technique that can distinguish hundreds or even thousands of different proteins in a cell extract

-PRODUCE -PROTEINS -DEPENDS -type -development -conditions -gel -separation

PROTEOMICS -.... EXAMINES THE FUNCTIONAL ROLES OF THE .... THAT A SPECIES CAN MAKE: ** -The entire collection of a species' proteins is its .... -Genomic ... can provide important information about the proteome: -RNA-Seq and DNA microarrays provide insight about the .... of genes -May not provide an ... measure of protein abundance -Genomic insights are often followed up with research that involves ... analysis directly

-PROTEOMICS -PROTEINS -proteome -data -transcription -accurate -protein

FORMATION OF HOMOLOGOUS GENES IN TWO SPECIES -IN THIS CASE, THE TWO SEQUENCES ARE .... BECAUSE THE GENES ARE ... TO EACH OTHER: -These are ...** -They are found in different species and have been ... from the same ... ...

-SIMILAR -HOMOLOGOUS -orthologs -derived -ancestral gene

TWO-DIMENSIONAL GEL ELECTROPHORESIS CONT... -SPECIFIC .... MAY BE OF SPECIAL INTEREST: -.... that are very abundant in a cell type: -May be important for that cells structure or ... -Spots present in only given ....: -Cells exposed to a hormone versus those that are ... -Spots present only in ... cells: -Very common in cancer cells

-SPOTS -Proteins -function -circumstances -not -abnormal

THE PROTEOME IS MUCH LARGER THAN THE GENOME -.... and analysis of an entire genome can identify all the ... that a given species contains -The proteome is ...., however, and its actual size is more difficult to determine -THIS LARGER SIZE IS ROOTED IN A NUMBER OF CELLULAR ...: -Alternative ... -... editing -.... covalent modification -All of these processes increase the number of potential .... in the proteome

-Sequencing -genes -larger -PROCESSES -splicing -RNA -Postranslational -proteinsAL

ANALYSIS OF DNA MICROARRAYS -Cells respond to environmental .... via the coordinate regulation of their ...: -Some genes are turned ... while others are turned off -In the past, .. ....was studied using tools that can analyze the expression of only a few genes at a time: -... make it possible to study the expression of the whole ... under different .... conditions**

-changes -genes -on -gene regulation -Microarrays -genome -environmental

BIOINFORMATICS -The .... has become an important tool in genetic studies: -The marriage between genetics and .... has yielded an important branch of science: .... -.... analysis of genetic sequences usually relies on three basic components: -A computer -A computer ... -Some type of data

-computer -biocomputing -bioinformatics -Computer -program

COMPUTER DATABASES -The amount of .... information that is generated by researchers has become enormous -Large numbers of .... data files are collected and stored in a single location, a ...: -The files in a .... typically contain annotations -They contain the genetic ... and a concise description of it -In addition, they contain other features of significance -The scientific community has created several large databases containing data from thousands of ...

-genetic -computer -database -database -sequence -labs

ORTHOLOGS AND PARALOGS -When two .... genes are found in different species, these genes are termed .... -When ... .... genes are found in a single organism, these genes are termed ....:** -A ... ... consists of two or more copies of homologous genes within the genome of a single organism** -Homologous genes are often ..... and identified using computer data ..., which are described next

-homologous -orthologs -two homologous -paralogs -gene family -analyzed -bases

BLAST -BLAST starts with a genetic sequence and then .... ... sequences in a ... database: -Homology among .... sequences is easier to identify than is DNA sequence homology -Table 23.5 (next slide) shows the results of a database search: -The amino acid sequence of human phenylalanine hydroxylase was used as a "query sequence" by the BLAST program -Within minutes, BLAST can search the ... ... and determine which ... are the closest matches: -Small E-Value indicates that similarity is .... to be due to random events; the genes are likely to be homologous -E-values depend on length of query, number of gaps in alignment, and .... ....

-locates -homologous -large -protein -entire database -sequences -unlikely -database size

HOMOLOGOUS GENES ARE DERIVED FROM THE SAME ANCESTRAL GENE -When comparing genetic ....., researchers sometimes find two or more similar sequences -Example: DNA sequences of the lacY gene in E. coli and K. pneumoniae: ~ 78% of the bases are a perfect match; Fig. 23.9a

-sequences

EXAMPLE: TRANSLATING A DNA SEQUENCE INTO AN AMINO ACID SEQUENCE -Consider a program aimed at .... a DNA sequence: -The geneticist (i.e. the user) has a ... sequence that needs to be translated into an .. ....sequence -DNA sequence of the lacY gene (shown in Figure 23.7) -The user is sitting at a computer that can run the .... program: -The program can .... in all three forward reading frames; the longest reading frame is shown here:

-translating -DNA -amino acid -TRANSLATION -translate