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increasing reprogramming effi ciency. One can apply fewer genetic manipulations to
preprogram neural progenitor cells in comparison with previously reported somatic
cells. On the other hand, some small molecules may be used as integration factors
instead of certain viral particles. These molecules also promote the reprogramming
process (Shi et al. 2008 ). These approaches can readily translate cell types directly
into a clinical type; such has been successful in neurons and cardiac cells. This
achievement is an essential contribution of chemical biology in stem cell research
that elucidates a number of specifi c advantages in applying small molecules (Jung
et al. 2014 ).
Transposon systems as viral-based methods have an interesting development in
hyperactive transposase enzymes. PiggyBac, Sleeping Beauty, and Tol2 transposon
systems are simple strategies that have been frequently used for various applica-
tions, and the system components can be separated on two plasmids. The fi rst plas-
mid carries an expression cassette for the particular transposase enzyme and the
second plasmid carries the transgene fl anked by inverted terminal repeats (ITRs).
Both plasmids are introduced into cells and the transposase is expressed, followed
by the transposition of the ITR-fl anked transgene into the genome. Importantly, the
transgenes integrate by a cut-and-paste mechanism and all residual plasmid ele-
ments are eliminated by degradation. In general, transposon integrations occur ran-
domly in the genome with no preference for gene-containing regions or promoter
sites. The transposase can be introduced in trans for completely integrated transpo-
son removal, resulting in safe and clean iPSCs (Kumar et al. 2015 ). For instance, the
bovine-derived iPSCs that use the piggyBac and Sleeping Beauty transposon sys-
tems include a different group of reprogramming factors, each regulated by the
chimeric CAGGS promoter and separated by self-cleaving peptide sequences.
Another bovine iPSC line produced by a piggyBac vector that consists of six key
reprogramming genes has been examined in detail, including alkaline phosphatase
expression, morphology, and typical pluripotent hallmarks, such as pluripotency
marker expression and mature teratoma formation in a mouse model. Furthermore,
this iPSC line is able to transfer the Sleeping Beauty transposon in a second round.
These achievements are p romising for germline-competent bovine-derived iPSCs
and provide a strategy for bovine genome genetic modifi cation (Talluri et al. 2015 ).
One obstacle in reprogramming is the introduction of exogenous genetic modifi -
cations in host cells. This obstacle can be overcome by directly transferring repro-
gramming proteins into cells instead of requiring the transcription of inserted genes
by the host cells. A fully somatic reprogramming study on murine fi broblasts
reported a protein transduction method that directly transfers recombinant repro-
gramming proteins. This result served as a signifi cant landmark in generating iPSCs
and has several principal advantages over previous methods (Zhou et al. 2009 ).
However, t his method does suffer some disadvantages, including low effi ciency and
high costs because of the number of required protein factors.
Another trend in nonviral iPSCs is the use of RNA molecules. The major steps
include synthesizing mRNAs encoding Oct4, Sox2, c-Myc, Klf-4, and SV40 large
T cells and introducing these mRNAs into host cells (i.e., human fi broblasts) by
electroporation. Transfecting fi broblasts with this mRNA mixture signifi cantly
P.V. Pham et al.