Science

Research Techniques

Industry Experience

LIMs
SOPs

QiaCube for Human Blood cfDNA
QiaSymphony for Buccal (Father DNA) and Human Blood cfDNA
gDNA Extraction & Isolation (Using QiaGen DNA Kits)
Aseptic Transfer
Pipetting

 

Academic Research Experience

E.Coli (HB 101)
Yeast (Saccharomyces Cerevisiae)
Bacteria/Yeast Culturing

Competent Cell Prep

  • I understand how to test and produce comp. cells for experiments

Competent Cell Test
Successful

 

Bacterial DNA Extraction / Purification
– Experienced in using :

  • QiaGen and Invitrogen Mini / Maxi -Prep kits
  • QIAquick Gel Extraction Kit
  • QIAquick PCR purification Kit

Time Management

  • Able to manage multiple on-going experiments at a time with minimal-no time wasting
  • Timers and checklists style approach for planned experiments and neccessary functions

Polymerase Chain Reaction

  • Designed and ran too many to count PCR reactions to amplify DNA, or gene sequence
  • Understand the main functions of the different phases of PCR, and how many cycles dependent on importance of the fidelity of target, however, sequencing can always be done
  • Also used PCR to verify gene orientation (5′ or 3′) after a transformation if it was a blunt end (non-directional / sticky end)
  • Used Taq polymerase generally, but I have also used reverse transcriptase in the case of template RNA

Nano Drop (Spectrophotometry)

  • In both my coursework and internships, I have used the NanoDrop extensively to find the concentration of a PCR product so I can calculate the correct ratios for subsequent experiments
Primer Design
  • Designed many primers for PCR when I needed to PCR a target gene from a genome/template, or to check orientation of my insertion after a ligation/transformation
Laboratory Notebook Keeping

Snippet of my personal NoteBook of experiments.
Featured picture of SFGate article.
Property of UCSF.

  • Checklist style format of accomplishing each experimental step
  • Detailed notes of balanced restriction digests
  • Knowledge of how to use NEB book to use proper buffers
  • Able to set up PCR reactions and calculate their elongation times
  • Diagnostic electrophoresis gels ran to confirm PCR product
  • Able to set up agarose gel for gel extraction of product
  • Able to calculate ratios of vector:insert for subsequent ligations

Project Presentation / Reporting

  • I am accustomed to weekly meetings where powerpoint presentations of the weeks findings are briefly and concisely reported to the laboratory to keep every one up to speed
  • Working in a lab, results on one part of a project can have an impact on your laboratory co-worker’s project, so communication between groups is key
  • I also have experience in preparing for and presenting a project infront of a large group of peers (200+) from the competitions that were my internships
Combinatorial Cloning
  • Understand, and have worked with single-shot ligation transformation procedures: ex:
Multi-part Cloning:
– Used multi-part cloning strategy based on Type IIs restriction enzyme AarI to generate, in a single ligation, a vector containing:
  • Promotor
  • Fluorescent Protein Tag
  • Negative Effector
  • Recruitment Domain (synthetic leucine zipper)
  • Transcriptional Terminator
To do that we made two families of “Donor” and “Acceptor” vectors, based on the yeast expression vector pRS315.
The 5 donor vectors contain a single part (RFP tag, Negative Effectors YopJ, YopH, or OspF; Synthetic Leucine Zippers for recruitment, “EE” or “RR”) cloned between two AarI sites, with distinct 4-base overhangs.  (named A, B, C, or D, as shown in the Figure below).
The 4 acceptor vectors contain a Promotor (pSte5, pCyc, pAdh, or pFig; the first three are constitutive with low/med/high expression levels respectively, and the last one is induced by activation of the mating pathway), two AarI restriction sites (each one with a distinct 4-base overhang, named A and D), and a transcriptional terminator (red circle); as shown in the figure below.
Each final construct was then made by ligation into a given acceptor vector, an RFP tag, followed by a negative effector and a leucine zipper, resulting in the following constructs.

Cloning of the Genome-Integrated Scaffold fused to the Recruitment Domain

Synthetic scaffold constructs fused to a leucine zipper (EE or RR) were made by cloning 500 bp of the Ste5 promotor together with the Ste5 open reading frame (ORF) as a single BgIII/BamHI fragment into pRS305.
Zipper sequences were cloned in, then cloned with BamHI/NotI sites and included a TAG stop codon immediately following their coding sequences.
The Adh terminator region was cloned as NotI/SacI fragments.  BgIII and SacI were then used to move the synthetic scaffold cassettes into the BamHI and SacI sites of the M4366 multiple cloning site (MCS).

Cloning of the Genome-Integrated Scaffold fused to the Recruitment Domain

Synthetic scaffold constructs fused to a leucine zipper (EE or RR) were made by cloning 500 bp of the Ste5 promotor together with the Ste5 open reading frame (ORF) as a single BgIII/BamHI fragment into pRS305.
Zipper sequences were cloned in, then cloned with BamHI/NotI sites and included a TAG stop codon immediately following their coding sequences.
The Adh terminator region was cloned as NotI/SacI fragments.  BgIII and SacI were then used to move the synthetic scaffold cassettes into the BamHI and SacI sites of the M4366 multiple cloning site (MCS).

Flow Cytometry (FACs)

  • Knowledge in growing cells to early log phase and then halting their growth (ie: Cycloheximide) and then dispensing them into 96-well culture plates for high-throughput sampling.
  • 10,000 cell readings were counted for each reading, and GFP was the fluorescence measured by exciting at 488 nm with a 100 mW Coherent Sapphire laser.  mean fluorescent values reported are the result of at least 3 independent values.

    The graphs in the slide seen below were generated by me with FlowJo after running the Flow Cytometry experiment.
     

Solutions Oriented Approach

  • I am a creative problem solver.  If something is not available for the next step, I will utilize what I have to achieve the goal while maintaining necessary precautions.
  • I’ve picked up a few materials-and-time-saving-tricks that I could show you in regards to Eppendorfs / Parafilm / small diagnostics
  • I can run a 10 minute quick gel diagnostic that uses minimal product to test whether a PCR was successful or not from a neat trick I learned from UCSF that I could show you if the materials for the setup are available
Culturing 15x transformations, but the laboratory didn't have the necessary mounts. I made my own and it worked beautifully

Culturing 15x transformations, but the laboratory didn’t have the necessary mounts.
I made my own and it worked successfully

 

Happy Hour

  • I love drinking with co-workers when it’s allowed, but not on the job >;)

 

RESEARCH AWARDS

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