Enhanced efficiency of P-element mediated transgenesis in Drosophila: Microinjection of DNA complexed with nanomaterial

نویسندگان

  • Madhavi Sonane
  • Ritu Goyal
  • Debapratim K. Chowdhuri
  • Kristipati Ravi Ram
  • Kailash C. Gupta
چکیده

The efficiency of genetic transformation technology to generate stable transgenics depends upon the successful delivery of plasmid DNA in embryonic cells. The available gene vectors facilitate efficient plasmid DNA delivery to the cellular milieu but are exposed to nuclease degradation. Recent in vitro studies suggest encapsulation of plasmid DNA with nanomaterial(s) for better protection against nucleases. Therefore, in this study, we tested if complexing of free plasmid DNA with linear polyethylenimine (LPEI, 25 kDa) based nanoparticle (LPN) enhances the efficiency of transformation (transgenesis) by using Drosophila based germ-line transformation technology. Here, we show that the LPN-DNA complex not only enhances the efficiency of this transgenic technology at a DNA concentration of 0.04 μg/μl but also reduces the DNA quantity required to generate transgenics by ten folds. This approach has potential applications for other types of transgenesis and nucleic acid injection methods in Drosophila as well as other popular genetic model systems.

برای دانلود متن کامل این مقاله و بیش از 32 میلیون مقاله دیگر ابتدا ثبت نام کنید

ثبت نام

اگر عضو سایت هستید لطفا وارد حساب کاربری خود شوید

منابع مشابه

Techniques for augmentation of exogenous DNA uptake by ovine spermatozoa

Sperm mediated gene transfer can be an inexpensive and simple method in animal transgenesis; however its efficiency is poor, mainly due to the spermatozoa’s lesser uptake of exogenous DNA. In the present study, the effects of lipofection and other augmentation techniques, such as sperm freezing and spermatozoa treatment with triton X100 and DMSO, on exogenous DNA uptake by sheep spermatozoa and...

متن کامل

Genomic DNA damage in mouse transgenesis.

Creating transgenic mammals is currently a very inefficient process. In addition to problems with transgene integration and unpredictable expression patterns of the inserted gene, embryo loss occurs at various developmental stages. In the present study, we demonstrate that this loss is due to chromosomal damage. We examined the integrity of chromosomes in embryos produced by microinjection of p...

متن کامل

The Meganuclease I-SceI Containing Nuclear Localization Signal (NLS-I-SceI) Efficiently Mediated Mammalian Germline Transgenesis via Embryo Cytoplasmic Microinjection

The meganuclease I-SceI has been effectively used to facilitate transgenesis in fish eggs for nearly a decade. I-SceI-mediated transgenesis is simply via embryo cytoplasmic microinjection and only involves plasmid vectors containing I-SceI recognition sequences, therefore regarding the transgenesis process and application of resulted transgenic organisms, I-SceI-mediated transgenesis is of mini...

متن کامل

Hyperactive self-inactivating piggyBac for transposase-enhanced pronuclear microinjection transgenesis.

We have developed a unique method for mouse transgenesis. The transposase-enhanced pronuclear microinjection (PNI) technique described herein uses the hyperactive piggyBac transposase to insert a large transgene into the mouse genome. This procedure increased transgene integration efficiency by fivefold compared with conventional PNI or intracytoplasmic sperm injection-mediated transgenesis. Ou...

متن کامل

Generation of transgenic rats with YACs and BACs: preparation procedures and integrity of microinjected DNA.

The aim of the present study was to investigate differences in the methods for preparing a large DNA fragment to be used for making transgenic rats from the standpoint of transgenic production efficiency and integrity of the introduced gene. In yeast artificial chromosome (YAC) transgenesis, three methods for preparing DNA for microinjection were compared: amplification of YAC in yeast (AMP), a...

متن کامل

ذخیره در منابع من


  با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید

برای دانلود متن کامل این مقاله و بیش از 32 میلیون مقاله دیگر ابتدا ثبت نام کنید

ثبت نام

اگر عضو سایت هستید لطفا وارد حساب کاربری خود شوید

عنوان ژورنال:

دوره 3  شماره 

صفحات  -

تاریخ انتشار 2013