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\begin{figure}[ht]
\raggedright
\includegraphics[width=0.75\textwidth]{figures/test.eps}
\end{figure}
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\begin{document}
\tableofcontents
\include{introduction}
\include{material_methods}
\bibliographystyle{plain}
\bibliography{references/my_library}
\end{document}
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\chapter{Material and Methods}
\section{Material}
\subsection{test}
In table \ref{tab:chemicals_buffers_media} used buffers and media as well as their composition are listed.
\begingroup
\footnotesize
\begin{longtable}[l]{lrllp{4.99cm}}
\caption{test.} \\
\toprule
\textbf{test} & & \textbf{test} & & \textbf{test} \\
\midrule
\endfirsthead
\caption{test (continued)} \\
\toprule
\textbf{test} & & \textbf{test} & & \textbf{test} \\
\midrule
\endhead
\textit{Media} \\
\cline{1-1} \\
2x YT & 31g & 2x yeast tryptone mix & & per liter \\
& \\
TYH \cite{cull_biotinylation_2000} & 20g & tryptone & & per liter; pH 7.2 calibrated with KOH \\
& 10g & yeast extract & & \\
& 11g & HEPES & & \\
& 5g & NaCl & & \\
& 1g & MgSO$_{4}$ & & \\
&\\
2x YT-Agar & 15.5g & 2xYT & & per 500mL \\
& 1\% & glucose & & w/v \\
& 1.5\% & agar & & w/v \\
&\\
\textit{SDS-PAGE \& Western Blot} \\
\cline{1-1} \\
SDS electrophoresis buffer & 25mM & TRIS & & \\
& 192mM & glycine & \\
& 0.1\% & sodium dodecyl sulfate & & w/v \\
&\\
Upper TRIS & 125mM & TRIS & & pH 6.8 \\
& 0.4\% & sodium dodecyl sulfate & & w/v \\
&\\
Lower TRIS & 375mM & TRIS & & pH 8.8 \\
& 0.4\% & sodium dodecyl sulfate & & w/v \\
&\\
5x SDS Sample Buffer & 175mM & TRIS & & pH 6.8 \\
- reducing - & 50\% & glycerol & & v/v \\
& 10\% & sodium dodecyl sulfate & & w/v \\
& 5\% & $\beta$$-$ mercaptoethanol & & v/v \\
& 0.15\% & bromphenol blue & & w/v \\
&\\
5x SDS Sample Buffer & 175mM & TRIS & & pH 6.8 \\
- non-reducing - & 50\% & glycerol & & v/v \\
& 10\% & sodium dodecyl sulfate & & w/v \\
& 10mM & iodacetamid & & v/v \\
& 0.15\% & bromphenol blue & & w/v \\
&\\
Wet Western Blot Buffer & 48mM & TRIS & & pH 9.2 \\
& 39mM & glycine & & \\
& 20\% & methanol & & v/v \\
&\\
\textit{Agarose Gels} \\
\cline{1-1} \\
TAE Buffer & 40mM & TRIS & & \\
& 5mM & sodium acetate & & \\
& 5mM & acetic acid & & w/v \\
& 1mM & Na${_2}$*EDTA*2H${_2}$O & & \\
&\\
6x DNA Loading Buffer & 0.25\% & bromphenol blue & & w/v \\
& 0.25\% & xylene cyanole & & w/v \\
& 40\% & sucrose & & w/v \\
&\\
10x Orange G DNA Buffer & 4.42mM & Orange G & & w/v \\
& 30\% & glycerol & & w/v \\
&\\
\textit{IMAC Purification} \\
\cline{1-1} \\
TBSU-400 & 100mM & TRIS & & pH 8 \\
& 400mM & sodium chloride & & \\
& 6M & urea & & \\
&\\
TBSU-W & 100mM & TRIS & & pH 8 \\
& 400mM & sodium chloride & & \\
& 20mM & imidazole & & \\
& 6M & urea & & \\
&\\
TBSU-E & 100mM & TRIS & & pH 8 \\
& 400mM & sodium chloride & & \\
& 250mM & imidazole & & \\
& 10\% & glycerole & & \\
& 6M & urea & & \\
&\\
\textit{Thiol Assay} \\
\cline{1-1} \\
Citrate/EDTA/Urea & 100mM & citrate & & \\
& 0.2mM & EDTA & & \\
& 6M & urea & & \\
& \\
TRIS/Urea & 500mM & TRIS & & pH 4.5 \\
& 8M & urea & & \\
& \\
\textit{Ribosome Display} \footnote{these buffer where prepared with ribosome display quality water. See method for further details.} \\
\cline{1-1} \\
10x TBS & 500mM & TRIS & & pH 7.4 at 4$^{\circ}$C and sterile filtered \\
& 1.5M & sodium chloride & & \\
& \\
TBS-T & 50mM & TRIS & & pH 7.4 at 4$^{\circ}$C \\
& 150mM & sodium chloride & & \\
& 0.05\% & tween 20 & & v/v \\
& \\
Washing Buffer & 50mM & TRIS & & pH 7.4 at 4$^{\circ}$C \\
& 150mM & sodium chloride & & \\
& 50mM & magnesium acetate & & \\
& 0,05\% & tween 20 & & v/v \\
& \\
Elution Buffer & 50mM & TRIS & & pH 7.4 at 4$^{\circ}$C \\
& 150mM & sodium chloride & & \\
& 25mM & EDTA & & \\
& \\
\textit{ELISA} \\
\cline{1-1} \\
pNPP Buffer & 50mM & NaHCO${_3}$ & & \\
& 50mM & MgCl${_2}$*6H${_2}$O & & \\
& \\
pNPP Solution & 3mM & pNPP & & dissolved in pNPP buffer \\
& \\
PBS & 137mM & sodium chloride & & \\
& 3mM & potassium chloride & & \\
& 8mM & Na${_2}$HPO${_4}$*2H${_2}$O & & \\
& 1.5mM & KH${_2}$HPO${_4}$ & & \\
& \\
PBS-T & 1x & PBS & & \\
& 0.1\% & tween 20 & & w/v \\
& \\
PBS-TB & 50mM & TRIS & & pH 7.4 at 4$^{\circ}$C \\
& 0.1\% & tween 20 & & w/v \\
& 0.2\% & bovine serum albumine & & w/v \\
& \\
\bottomrule
\label{tab:chemicals_buffers_media}
\end{longtable}
\endgroup
\subsection{Constructs, Plasmids \& Strains}
\subsubsection{test}
test
\subsubsection{Plasmids}
\paragraph{test}
This vector was provided by the host lab and its map is shown in figure \ref{fig:pat222}. It carries an avi tag and the protein D. The avi-tag is recognized and biotinylated by a biotin ligase. Protein D acts as a fusion protein and strongly increases the solubility of its fusion partner. Protein D is a head protein of the lambda phage and coupled to a his-tag makes it a perfect fusion partner for any protein to highly express and purify it. Next to other available fusion proteins, e.g. glutathione-S.transferase (GST) or thioredoxin \cite{berndt_thioredoxins_2008}, protein D does not have any enzymatic activity. Therefore there should be no concern on modifications or whatsoever of the protein of interest \cite{forrer_high-level_1998}. In this study it carried the designed domain constructs for overexpression into inclusion bodies \cite{kane_formation_1988} and purification via IMAC.
\begin{figure}[hp]
\raggedright
\includegraphics[width=0.75\textwidth]{figures/pat222.eps}
\caption{The vector map of pAT222 shows its inhabiting tags, fusion protein, antibiotic resistance as well as restriction sites}
\label{fig:pat222}
\end{figure}
\paragraph{pBirA}
This plasmid carries the \textit{birA} gene and the resistance against Chloramphenicol. \textit{birA} codes for a biotin ligase which transfers biotin molecules onto avidin or streptavidin. It was used together with pAT222 in a double transformation to biotinylate the domain construct \textit{in Vivo} \cite{cull_biotinylation_2000}. pBirA is commercially available from Avidity.
\paragraph{pRDV}
test
\begin{figure}[ht]
\raggedright
\includegraphics[width=0.75\textwidth]{figures/prdv.eps}
\caption{test}
\label{fig:prdv}
\end{figure}
\paragraph{pDST67}
test
\begin{figure}[ht]
\raggedright
\includegraphics[width=0.75\textwidth]{figures/pdst67.eps}
\caption{test}
\label{fig:pdst67}
\end{figure}
\subsubsection{Strains}
XL1-blue is a \textit{E.coli} strain commercially available from Stratagene. It was used for overexpressing the protein of interest as well as for DNA (plasmid) purification.
\subsection{Antibodies \& Enymes}
\subsubsection{Antibodies}
In table \ref{tab:antibodies} all used antibodies are listed.
\begingroup
\footnotesize
\begin{longtable}[l]{L{9cm}L{2.5cm}}
\caption{Antibodies used for detection at western blots, ELISA and FACS experiments.} \tn
\toprule
\textbf{Antibody} & \textbf{Supplier} \tn
\midrule
\endhead
mouse IgG- ${\alpha}$- His${_4}$ & host lab \tn
& \tn
goat Fab2-fragment- ${\alpha}$- mouse IgG H+L - HPR conjugate & host lab \tn
& \tn
mouse IgG- ${\alpha}$- RGSH${_4}$ & host lab \tn
& \tn
goat- ${\alpha}$- mouse IgG - AP conjugate & host lab \tn
& \tn
goat- ${\alpha}$- mouse IgG - Alexa 457 conjugate & host lab \tn
& \tn
\bottomrule
\label{tab:antibodies}
\end{longtable}
\endgroup
\subsubsection{Enzymes}
In table \ref{tab:enzymes} all used enzymes are listed.
\begingroup
\footnotesize
\begin{longtable}[l]{L{5cm}L{3.5cm}L{2.5cm}}
\caption{Enzymes used for cloning and ribosome display with their respective class and supplier.} \tn
\toprule
\textbf{Enzyme} & \textbf{Class} & \textbf{Supplier} \tn
\midrule
\endhead
\textit{Bam}HI & REN & Fermentas \tn
& \tn
\textit{Hind}III & REN & Fermentas \tn
& \tn
\textit{Nco}I & REN & Fermentas \tn
& \tn
T4 ligase & ligase & Fermentas \tn
& \tn
Calf Intestinal Phosphorylase & phosphorylase & Fermentas \tn
& \tn
Vent & polymerase & NEB \tn
& \tn
T7 RNA polymerase & polymerase & Fermentas \tn
& \tn
AffinityScript RT & reverse transcriptase & Stratagene \tn
& \tn
DNaseI & DNA specific endonuclease & Roche \tn
& \tn
\bottomrule
\label{tab:enzymes}
\end{longtable}
\endgroup
\section{Methods}
\subsection{Basic Applications}
The web link gmedina has given you here is teaching you this.yaakov wrote:i couldnt evaluate, what parts of the code are important and causing the problem.
Learn LaTeX easily with newest books:
The LaTeX Beginner's Guide: 2nd edition and perfect for students writing a thesis
The LaTeX Cookbook: 2nd edition full of practical examples for mathematics, physics, chemistry, and more
LaTeX Graphics with TikZ: the first book about TikZ for perfect drawings in your LaTeX thesis
