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\title{\bf Introduction to the Bioconductor marray package : Classes  structure component (short)}

\author{Yee Hwa Yang$^1$ and Sandrine Dudoit$^2$}

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\begin{center}
1. Department of Medicine, University of California, San Francisco, {\tt jean@biostat.berkeley.edu}\\
2. Division of Biostatistics, University of California, Berkeley.\\
\end{center}


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\section{Overview}

This document provides a short tutorial on the basic class definitions
and associated methods used in the {\tt marray} package.  To load the
{\tt marray} package in your R session, type {\tt library(marray)}. 
<<eval=TRUE, echo=TRUE>>=
library(marray)
data(swirl)
@

\section{Microarray classes}

The three main classes for cDNA microarray data are:

\begin{description}
\item
{{\tt marrayLayout}:} 
This class is used to keep track of important layout parameters for
two--color cDNA microarrays. It contains slots for: the total number of
spotted probe sequences on the array, the 
dimensions of the spot and grid matrices, the plate origin of the
probes, information on spotted control sequences (e.g. probe sequences
which should have equal abundance in the two target samples, such as
housekeeping genes).   

\item
{{\tt marrayRaw}:} This class represents pre--normalization intensity
data for a batch of cDNA microarrays. A {\it batch} of arrays consists
of a collection of arrays with the same layout ("marrayLayout"). The
class contains slots for the green (Cy3) and red (Cy5) foreground and
background intensities, the layout of the arrays, and descriptions of
the target samples hybridized to the arrays and probe sequences spotted
onto the arrays.

\item
{{\tt marrayNorm}:} This class represents post--normalization intensity
data for a batch of cDNA microarrays.  The class contains slots for the
average log--intensities $A = \log_2 \sqrt{RG}$, the normalized
log--ratios $M = \log_2 R/G$, the location and scale normalization
values, the layout of the arrays, and descriptions of the target samples
hybridized to the arrays and probe sequences spotted onto the arrays.
\end{description}

Other classes are {\tt marrayInfo} which can be used to represents the
Target or the Probes information.  The function {\tt slotNames} can be
used to get information on the slots of a formally defined class or an
instance of the class. For example, to get information of the slots for
the {\tt marrayLayout} class or on the slots for the object {\tt swirl}
use

<<eval=TRUE>>=
slotNames("marrayLayout")
slotNames(swirl)
@

\section{Creating and accessing slots of microarray objects} 

{\bf Creating new objects.} The function {\tt new} from the {\tt
methods} package may be used to create new objects from a given
class. For example, to create an object of class {\tt marrayInfo}
describing the target samples in the Swirl experiment, one could use the
following code  

<<eval=TRUE>>=
zebra.RG<-as.data.frame(cbind(c("swirl","WT","swirl","WT"), c("WT","swirl","WT","swirl"))) 
dimnames(zebra.RG)[[2]]<-c("Cy3","Cy5")
zebra.samples<-new("marrayInfo",
		    maLabels=paste("Swirl array ",1:4,sep=""), 
		    maInfo=zebra.RG,
		    maNotes="Description of targets for Swirl experiment")
zebra.samples
@

Slots which are not specified in {\tt new} are initialized to the
prototype for the corresponding class. These are usually "empty", e.g.,
{\tt matrix(0,0,0)}. In most cases, microarray objects can be created
automatically using the input functions and their corresponding widgets
in the {\tt marrayInput} package. These were used to create the object
{\tt swirl} of class {\tt marrayRaw}.

{\bf Accessing slots.} Different components or slots of the microarray
objects may be accessed using the operator {\tt @}, or alternately, the
function {\tt slot}, which evaluates the slot name. For example, to
access the {\tt maLayout} slot in the object {\tt swirl} and the {\tt
maNgr} slot in the layout object {\tt L}:

<<eval=TRUE>>=
L<-slot(swirl, "maLayout")
L@maNgr
@

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\section{Basic microarray methods}\label{smethods}

The following basic methods were defined to facilitate manipulation of
microarray data objects.  To see all methods available for a particular
class, e.g., {\tt marrayLayout}, or just the print methods

<<eval=FALSE>>==
showMethods(classes="marrayLayout")
showMethods("summary",classes="marrayLayout") 
@

\subsection{Printing methods for microarray objects} 

Since there is usually no need to print out fluorescence intensities for
thousands of genes, the {\tt print} method was overloaded for microarray
classes by simple report generators. For an overview of the available
microarray printing methods, type {\tt  methods ? print}, or to see all
print methods for the session 

<<eval=TRUE>>=
showMethods("print")
@

For example, summary statistics for an object of class {\tt marrayRaw},
such as {\tt swirl}, can be obtained by {\tt summary(swirl)} 

<<eval=TRUE>>=
summary(swirl)
@

\subsection{Subsetting methods for microarray objects} 

In many instances, one is interested in accessing only a subset of
arrays in a batch and/or spots in an array. Subsetting methods {\tt "["}
were defined for this purpose.  For an overview of the available
microarray subsetting methods, type {\tt methods ? "["} or to see all
subsetting methods for the session {\tt showMethods("[")}.  When using
the {\tt "["} operator, the first index refers to spots and the second
to arrays in a batch. Thus, to access the first 100 probe sequences in
the second and third arrays in the batch {\tt swirl} use

<<eval=TRUE>>=
swirl[1:100,2:3]
@

\subsection{Methods for accessing slots of microarray objects}


A number of simple methods were defined to access slots of the
microarray classes.  Using such methods is more general than using the
{\tt slot} function or {\tt @} operator. In particular, if the class
definitions are changed, any function which uses the {\tt @} operator
will need to be modified. When using a method to access the data in the
slot, only that particular method needs to be modified. Thus, to access
the layout information for the array batch {\tt swirl} one may also use
{\tt maLayout(swirl)}.\\ 

 In addition, various methods were defined to compute basic statistics
 from microarray object slots. For instance, for memory management
 reasons, objects of class {\tt marrayLayout} do not store the spot
 coordinates of each probe. Rather, these can be obtained from the
 dimensions of the grid and spot matrices by applying methods: {\tt
 maGridRow}, {\tt maGridCol}, {\tt maSpotRow}, and {\tt maSpotCol} to
 objects of class {\tt marrayLayout}. Print--tip--group coordinates are
 given by {\tt maPrintTip}. Similar methods were also defined to operate
 directly on objects of class {\tt marrayRaw} and {\tt marrayNorm}. The
 commands below may be used to display the number of spots on the array,
 the dimensions of the grid matrix, and the print--tip--group
 coordinates.

<<eval=TRUE>>=
swirl.layout<-maLayout(swirl)
maNspots(swirl)
maNspots(swirl.layout)
maNgr(swirl)
maNgc(swirl.layout)
maPrintTip(swirl[1:10,3])
@

\subsection{Methods for assigning slots of microarray objects}

A number of methods were defined to replace slots of microarray objects,
 without explicitly using the {\tt @} operator or {\tt slot}
 function. These make use of the {\tt setReplaceMethod} function from
 the R {\tt methods} package.  As with the accessor methods just
 described, the assignment methods are named after the slots. For
 example, to replace the {\tt maNotes} slot of {\tt swirl.layout}

<<eval=TRUE>>=
maNotes(swirl.layout)
maNotes(swirl.layout)<- "New value"
maNotes(swirl.layout)
@

To initialize slots of an empty {\tt marrayLayout} object

<<eval=TRUE>>=
L<-new("marrayLayout")
L
maNgr(L)<-4
@

Similar methods were defined to operate on objects of class {\tt
marrayInfo}, {\tt marrayRaw} and {\tt marrayNorm}.

\subsection{Methods for coercing microarray objects} 

To facilitate navigation between different classes of microarray
objects, we have defined methods for coercing microarray objects from
one class into another. A list of such methods can be obtained by {\tt
methods ? coerce}. For example, to coerce an object of class {\tt
marrayRaw} into an object of class {\tt marrayNorm}

<<eval=TRUE>>=
swirl.norm<-as(swirl, "marrayNorm")    
@

It is also possible to convert objects of class {\tt marrayRaw} or {\tt
marrayNorm} into objects of class {\tt exprSet} (see definition in the
{\tt Biobase} and {\tt convert} package)

\begin{verbatim}
> library(convert)
> as(normdata, "exprSet")    
\end{verbatim}


\subsection{Functions for computing layout parameters}

In some cases, plate information is not stored in {\tt marrayLayout}
objects when the data are first read into R. We have defined a function
{\tt maCompPlate} which computes plate indices from the dimensions of
the grid matrix and number of wells in a plate. For example, the Swirl
arrays were printed from 384--well plates, but the plate IDs were not
stored in the {\tt fish.gal} file. To generate plate IDs (arbitrarily
labeled by integers starting with 1) and store these in the {\tt
maPlate} slot of the {\tt marrayLayout} object use

<<eval=TRUE>>=
maPlate(swirl)<-maCompPlate(swirl,n=384)
@

Similar functions were defined to generate and manipulate spot
coordinates: {\tt maCompCoord}, {\tt maCompInd}, {\tt maCoord2Ind}, {\tt
maInd2Coord}. The function {\tt maGeneTable} produces a table of spot
coordinates and gene names for objects of class {\tt marrayRaw} and{\tt
marrayNorm}.

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