ライフサイエンスコーパス: minigene

1 APII, and the splicing factor SF2/ASF at the minigene.

2 TTC repeats into an intron of a GFP reporter minigene.

3 ing and translational inhibition from a DICE-minigene.

4 tive splicing and exon inclusion from a CD44 minigene.

5 n 7 inclusion in mRNAs derived from the SMN2 minigene.

6 sis of a 51-nt internal exon in a three-exon minigene.

7 der the control of a human Cathepsin G (hCG) minigene.

8 ative poly(A) site selection on the reporter minigene.

9 atory elements in the alpha-tropomyosin test minigene.

10 e endogenous gene, as well as from an IKBKAP minigene.

11 elta147) when co-expressed as an ER-targeted minigene.

12 ess-induced alternative splicing of the MDM2 minigene.

13 to be an independent event localized to the minigene.

14 licing efficiency of the introns of the MDM2 minigene.

15 cts the splicing of a damage-responsive MDM2 minigene.

16 protein 1 and regulates splicing of reporter minigenes.

17 nsfection of G(o1) minigenes but not G(i1/2) minigenes.

18 er copy-number dependence than observed with minigenes.

19 g RNA analysis of human samples and splicing minigenes.

20 ion of the SC35 binding site in the PKCdelta minigene abolished RA-mediated utilization of 5' splice

21 , which, when introduced into the human CFTR minigene, abolished exon 9 skipping.

22 ion of a 395 bp upstream region in the MOR23 minigene abolishes expression.

23 DC pulsed with TAT-minigene also efficiently induced Flu-M1-specific T cell

24 Induction of the AUA minigene also stimulates both leftward and rightward fra

25 iates with cyclin D1b mRNA (transcript-b) in minigene analyses and with endogenous transcript in pros

26 Minigene analyses demonstrated that this alteration prec

27 Bioinformatic and minigene analyses identified signals that could modify t

28 Minigene analysis showed that the resultant net loss of

29 To test this idea, we constructed a CFTR minigene and replicated exon 9 skipping associated with

30 ecrease exon 10 inclusion in a wild type tau minigene and rescue the increase in exon 10 splicing cau

31 , we cloned a heterologous splicing PKCdelta minigene and showed that inclusion of PKCdelta exon 9 is

32 ge-specific alternative splicing of the MDM2 minigene and that the splicing factor SRSF1 binds exon 1

33 n IIIb inclusion in transcripts derived from minigenes and from the endogenous FGF-R2 gene.

34 eceptors was further combined with G protein minigenes and pharmacological intervention, along with c

35 Transfection of cells with single-exon IIIb minigenes and single-exon IIIc minigenes revealed that m

36 of COL2A1; construction of splicing reporter minigenes and transfection into cultured cells; and RT-P

37 hin two different internal exons in a 3-exon minigene, and millions of successfully spliced transcrip

38 n, the dominant negative Galpha(q)-(305-359) minigene, and pretreatment with pertussis toxin inhibite

39 ion on pre-mRNA splicing was studied using a minigene approach.

40 Here we show that the mouse and human minigenes are regulated similarly by conserved elements

41 cient to induce exon 7 skipping in the mouse minigene as in the human SMN2.

42 ependent exon 6 exclusion from fas mRNA in a minigene assay in cells.

43 Genetic advances include the use of the minigene assay to confirm pathogenicity of splice site m

44 itative real-time polymerase chain reaction, minigene assay, and flow cytometry.

45 Using a splicing reporter minigene assay, we demonstrated that RBM4 promoted exon

46 Using a minigene assay, we showed that c.790A>G altered CIZ1 spl

47 Minigene assays were performed for two non-canonical spl

48 his variant completely prevented splicing in minigene assays, and resulted in exon skipping and an in

49 CLRN1 minigene-based analysis confirmed the splicing of an abe

50 t Wt1 promotes exon inclusion using a Vegf-a minigene-based splicing assay.

51 We show that a DUX4 minigene, bearing only the homeodomains and C-terminus,

52 We also identified two dipeptide coding minigenes between the Shine-Dalgarno sequence and start

53 nd could be blocked by transfection of G(o1) minigenes but not G(i1/2) minigenes.

54 When introduced as a minigene by retroviral transduction into human breast ca

55 pattern of LH2, both endogenously and in the minigene, by the RNA-binding splicing proteins TIA-1 and

56 ly, we have shown that use of the C-terminal minigenes can specifically block receptor activation of

57 Using minigenes carrying deletion mutations within intron 11,

58 ct of high-level expression of two different minigenes carrying either the rpoB-2 or the ndhF-2 editi

59 HEK293T cell transduction with minigenes carrying exon 39 showed that the splice defect

60 Using minigenes carrying linker-scanning mutations within exon

61 Transfection of minigenes carrying mutations associated with DRRD (G118R

62 Using a CoAA minigene cassette, we find that the switched alternative

63 in D3-induced expression of a growth hormone minigene cassette.

64 and rapid approach based on a modular hybrid minigene combined with antisense oligonucleotides to ena

65 The shark kappa clusters are minigenes consisting of a simple VL-JL-CL array, where V

66 In transgenic mice, a minigene construct containing 10 kb of upstream and 1.5

67 An FLT1 minigene construct containing exon 13, 14 and the interv

68 COS-1 and HepG2 cells transfected with a F5 minigene construct containing the patient's mutation exp

69 ption of dimerization using a TM4 peptide, a minigene construct encoding TM4, or by mutation of TM4,

70 The truncated alpha1(XII) collagen minigene construct MXIINC3(-), driven by the mouse alpha

71 Mice immunized with a minigene construct of hgp100(25-33) rejected B16 melanom

72 A minigene construct showed that this mutation alters spli

73 s faster and less sensitive to a PLCbeta1-ct minigene construct than inhibition of PIP(2)-sensitive K

74 We have engineered a human tau minigene construct that was designed to allow alternativ

75 gulate alternative splicing using a splicing minigene construct transfected into the oligodendrocyte

76 tion of Bcl-x pre-mRNA, we developed a BCL-x minigene construct which conferred the same ratio of Bcl

77 moter upstream of a human lipoprotein lipase minigene construct with a glycosylphosphatidylinositol a

78 e development using a recombinant protein or minigene construct, a series of truncated recombinant RA

79 equences, when transferred to a heterologous minigene construct, inhibited splicing but only when clo

80 rted at different positions within the FGFR2 minigene construct.

81 Human FIX minigene constructs containing a simian virus 40 (SV40)

82 In vitro assays were performed with minigene constructs containing ABCA4 exon 39.

83 Minigene constructs containing the entire intron 4 seque

84 In Drosophila melanogaster S2 cells, using minigene constructs designed to produce nearly identical

85 ssays between Xenopus laevis and X. borealis minigene constructs injected into oocytes.

86 Minigene constructs of the HD locus confirm the regulato

87 AT3 tumors in syngeneic rats, we constructed minigene constructs that report on alternative splicing.

88 interpretation of data solely obtained with minigene constructs to study the effects of sequence var

89 re assessed by transfecting COS-7 cells with minigene constructs.

90 n of an alternative exon is through reporter minigene constructs.

91 tic), a finding reproduced in vitro by using minigene constructs.

92 and a collection of experimentally clarified minigene constructs.

93 us MYPT1 as well as stably transfected MYPT1 minigene constructs.

94 DNA ends (RACE), and transient expression of minigene constructs.

95 is of potential SRSF10 binding motifs in two minigene constructs.

96 d accumulation of both mRNA expressed from a minigene containing exon 12 and the endogenous G6PD mRNA

97 nusoidal endothelial cells by transfecting a minigene containing the EIIIA exon and its flanking intr

98 COS cells transfected with a minigene containing the intronic +14 mutation produce ex

99 pha-spectrin gene transcripts derived from a minigene containing the IVS 22 +5 mutation.

100 sgenic mouse model using a 2.9-kilobase HD-5 minigene containing two HD-5 exons and 1.4 kilobases of

101 quence could have dual roles, we transfected minigenes containing a series of 2-bp mutations in the 1

102 this effect was not specific to ETR-3 since minigenes containing either of the two motifs were respo

103 tigated by introducing translatable chimeric minigenes containing sequence -20 to +6 surrounding the

104 Minigenes containing X. laevis spacer sequences are only

105 Mutational analysis of an LH2 minigene demonstrated that 2 of the 4 Fox binding motifs

106 ation with a VV-e7r-expressing ubiquitinated minigene, demonstrated that memory CD8 T cells alone cou

107 rnative splicing, we systematically screened minigenes derived from the GABA(A)Rgamma2 and human beta

108 demonstrated that alternative splicing of a minigene-derived transcript to express ACF65 was enhance

109 hereas mice immunized with the mgp100(25-33) minigene did not develop protective tumor immunity.

110 competing 5' splice sites in an alpha-globin minigene, direct hnRNPH/F-regulated alternative splicing

111 The expression of a FVII minigene directed by a segment of the native FVII promot

112 A hybrid minigene driven by cytomegalovirus promoter mimicked 1B-

113 ransgenic line of mice carrying a tyrosinase minigene driven by the dopachrome tautomerase (Dct) prom

114 cinar cells in transgenic mice by creating a minigene driven by the rat elastase I enhancer/promoter.

115 CD8(+) T cell lines generated to minigene-encoded CTL epitopes secreted IFN-gamma and TNF

116 5-HT was antagonized by the expression of a minigene encoding a peptide scavenger of Gbetagamma subu

117 Here, we engineered a DNA minigene encoding seven H-2(b)-restricted Cpn CTL epitop

118 n PTLDKVLEV was pulsed onto cells, or when a minigene encoding this sequence was used to artificially

119 Using minigenes encoding defined T-helper epitopes from lympho

120 esponses by vaccinating rhesus macaques with minigenes encoding fragments of Gag, Vif, and Nef.

121 ma and beta-arrestin proteins; expression of minigenes encoding the carboxyl terminii of the G protei

122 ed whether vaccinating rhesus macaques with "minigenes" encoding fragments of Gag, Vif, and Nef resul

123 ith AGA at the test position; induction of a minigene ending in AGA stimulates bypassing at the latte

124 We find that induction of a minigene ending in AUA stimulates bypassing at an AUA co

125 Minigene experiments demonstrated an important role for

126 Minigene experiments explain disease-associated mutation

127 Reporter minigene experiments investigating representative exons

128 ional Ub from a poly hemagglutinin-tagged Ub minigene expressed from the Hprt locus.

129 sense mutations W1001X and R1014X using hERG minigenes expressed in HEK293 cells or neonatal rat vent

130 e containing ECR2 reproducibly directed lacZ minigene expression in mesoderm.

131 A minigene expression system confirms both the effect of t

132 temporal neocortex tissue and in a cellular minigene expression system.

133 h delivered one exon of a beta-galactosidase minigene flanked by donor or acceptor splice sequences.

134 We created minigenes for two of these genes, the CFTR and MTMR1 gen

135 Expression of minigene fragments from beta-arrestin1 or STAM1, known t

136 e constructed various tissue-specific unc-52 minigenes fused to a gene for green fluorescent protein

137 In contrast, DC pulsed with minigene fusion protein lacking TAT were either poorly r

138 A TAT-minigene fusion protein was generated, encoding both the

139 ally, the haplotype- and allele-specific ACE minigenes generated similar splicing patterns in both AC

140 Minigenes generated with pSpliceExpress show the same re

141 Transfection of a G(i1/2) minigene had no effect on thrombin-stimulated [Ca(2+)](i

142 We expressed minigenes harboring the mutations in cell lines to demon

143 ertion of another RNAPII pause site into the minigene has a similar effect on exon IIIb silencing.

144 evis spacer sequences are only dominant over minigenes having complete X. borealis spacers if a space

145 of the free Gbetagamma-specific sequestering minigene hbetaARK1(495) also inhibit EGLT-mediated CDC42

146 In addition, a Gbetagamma minigene, hbetaARK1(495), inhibits VPF/VEGF-stimulated H

147 Functional assays using splicing reporter minigenes identified the proteins hrp36 and hrp38 and th

148 in an immunogenic context; that is, as a DNA minigene in a bacterial carrier system.

149 a calcitonin/calcitonin gene-related peptide minigene in a hormone-dependent manner.

150 urvival, whereas re-introduction of an LRP-1 minigene in a mouse LRP-1-deficient fibroblast cell line

151 vel of mutant mRNA compared to the wild-type minigene in an NMD-dependent manner.

152 ate that the unplanned expression of the hGH minigene in CollagenVI expressing mesenchymal cells can

153 Splicing assays using a transfected TP53 minigene in combination with siRNA knockdown of SRSF3 sh

154 Expression of a minigene in LX2 containing the AZIN1 slow-fibrosis SNP y

155 xpressed in the same cell type as the unc-52 minigene in order to regulate its expression, supporting

156 findings by analyzing the effect of the hGH minigene in TgC6hp55 transgenic mice which express the h

157 enerated by the ectopic expression of an ADA minigene in the gastrointestinal tract of otherwise ADA-

158 asts as well as into transcripts from a SMN2 minigene in the motor neuron cell line NSC34.

159 vates exon inclusion of a cardiac troponin T minigene in transient transfection assays in an MSE-depe

160 short:long isoform-expression ratios of DRD2 minigenes in cell culture.

161 nisms are typically studied using artificial minigenes in cultured cells, conditions that may not acc

162 the potential use of RAD51-targeted ribozyme minigenes in tumour radiosensitisation.

163 ctivate exon inclusion of cardiac troponin T minigenes in vivo via muscle-specific splicing enhancer

164 tivated MSE-dependent exon inclusion of cTNT minigenes in vivo.

165 the MHC class I-dependent CD8(+) T cells to minigene-included Cpn CTL epitopes, rather than by pan-D

166 The minigene integrated 0.98 Mb upstream of Sox9 and was acc

167 s substantiated by transfection of the G(o1) minigene into HMECs, which led to a blockade of thrombin

168 ting of a single-copy gp91(phox) therapeutic minigene into one allele of the "safe harbor" AAVS1 locu

169 trastromal injection of a lumican expression minigene into the corneal stroma of Lum-/- mice.

170 e this hypothesis, we first transfected LDLR minigenes into SH-SY5Y neuroblastoma cells and found tha

171 s are here recapitulated by MOR23 and M71 OR minigenes, introduced into mice.

172 Therefore, introduction of a functional CD45 minigene is sufficient to overcome the principal severe

173 ability to direct this process by the use of minigenes is relevant to the design of vaccines and unde

174 into an intron in the APRT gene or the HPRT minigene, long tracts of CTG/CAG repeats (more than abou

175 To this end, we designed a minigene (M), and in combination with a series of deleti

176 e approaches, termed the "heterozygous" and "minigene" methods, we generated mice in which Cre-expres

177 exon 10-skipped CYP24A1 splice variant; in a minigene model the latter was attenuated by a functional

178 Expression of one of the minigenes modestly interfered with translation of ycbK.

179 Using high-throughput genetics, 5560 minigene molecules were assayed for splicing in human HE

180 Minigenes of 9 kb and as short as 2.2 kb are selectively

181 A panel of minigenes of varying splicing potential were integrated

182 idenced by investigating the effect of G(o1) minigenes on thrombin-stimulated stress fiber formation

183 n 7 inclusion in mRNAs derived from the SMN2 minigene or from endogenous SMN2.

184 icing errors of either the mutated human DDC minigene or the mouse artificial splicing construct in v

185 Manipulation of the UCE, in a reporter minigene or via random mutations in the genomic context

186 sigma, and sigma-2, encoded by 35 functional minigenes or clusters.

187 nd LFA-3, together with either p540 and p865 minigenes or the full-length hTERT, effectively stimulat

188 s were found when autologous hFIX cDNA, hFIX minigenes, or hPC minigenes were used as reporters in th

189 tion, these findings highlight the generated minigene panel as a flexible platform for the query of s

190 ylation was equivalently detected across the minigene panel, irrespective of splicing and H3K36me3 st

191 patS minigenes patS4 to patS8 encode PatS C-terminal 4 (GSGR)

192 and an engineered alpha-SNAP binding-domain minigene peptide blocked basal and evoked vWF secretion.

193 i) response to the same extent as with G(o1) minigene peptide, suggesting that this G(o)-mediated [Ca

194 We have now used the 'minigene' phenomenon to ascertain whether depletion of t

195 transcripts, the transfection of the A or C minigenes produced a large amount of the alternatively s

196 l peptide (PA(224-233)) expressed as a viral minigene product formed a sizeable cytosolic pool contin

197 rther, co-transfection of SC35 with PKCdelta minigene promoted selection of 5' splice site II.

198 The GFP_(GAA*TTC)(560) minigene recapitulates the molecular hallmarks of the mu

199 HITS-CLIP and minigene reporter analyses indicate that these polyadeny

200 of that SNP on alternative splicing using a minigene reporter assay.

201 In one gene (MAGOH), using minigene reporter assays, we demonstrated that the combi

202 Using a minigene reporter, we show that Sam68 controls expressio

203 Here, using a minigene reporter, we show that the nrg alternatively sp

204 dulate alternative exon inclusion from a MAG minigene reporter.

205 Two minigene reporters were constructed: One produces green

206 * and 33 can be recapitulated in transfected minigene reporters.

207 ant vaccinia virus expressing epitope V as a minigene, resulted in the induction of weak, but reprodu

208 ection of this cell line with a murine Xylt2 minigene results in the production of recombinant chondr

209 High level expression of plasmid-borne minigenes results in the sequestration as peptidyl-tRNA

210 of Akt2(-/-) cells with a PKCbetaII splicing minigene revealed defective betaII exon inclusion.

211 gle-exon IIIb minigenes and single-exon IIIc minigenes revealed that mutation of terminal sequences o

212 Minigene RNA splicing studies in BV2 microglial cells es

213 man fibroblasts (HFFs) expressing a Target 1 minigene RNA, which contains the required splicing and P

214 Analyzed with minigenes, SMN1C6T displayed a approximately 20% increas

215 lly, changing the promoter alters both FGFR2 minigene splicing and the MAZ4 effect.

216 to prioritize VUS and developed a cell-based minigene splicing assay to confirm aberrant splicing.

217 ns and demonstrates functional activity in a minigene splicing assay.

218 LW/MW exon 3 haplotypes by semi-quantitative minigene splicing assay.

219 Reticulocyte RNA and functional minigene splicing assays in heterologous cells revealed

220 Minigene splicing assays revealed a novel splicing event

221 Using a minigene splicing construct containing an SVA, we observ

222 Mutating iE(B) in minigene splicing reporters abrogated intrasplicing, as

223 In vitro minigene splicing studies qualitatively replicated these

224 Through chimeric minigenes splicing assay we investigated the unique elem

225 Using bacterial artificial chromosome-minigene stable cell lines, CRISPR/Cas9 enhancer-deleted

226 o levels comparable to that of the wild-type minigene, suggesting that hERG nonsense mutations are su

227 Correction of tau splicing occurs in a tau minigene system and in endogenous tau RNA in neuronal ph

228 of T1945+6C on splicing was studied using a minigene system and on function by heterologous expressi

229 Using an in vitro minigene system to analyze splicing events, we found red

230 In this manuscript we use an inducible minigene system to determine the time course of ASO acti

231 We used a minigene system to study two frameshift mutations, R1032

232 Using a CFTR minigene system, we studied TG tract variation and obser

233 at exon-intron boundaries using an in vitro minigene system.

234 Using a novel tyrosinase minigene-tagged Sleeping Beauty transposon-mediated muta

235 A dominantly acting K14-agouti minigene tags both rearrangements, which enables these b

236 Using a minigene that includes exon 4, intron 4, exon 5, intron

237 region, we replaced the region with a 659-bp minigene that linked the three cis-elements (MG-GFP).

238 DM2 splicing regulation by utilizing a novel minigene that mimics endogenous MDM2 splicing in respons

239 We have designed a G alpha(s) minigene that retains the signals required for G alpha(s

240 Using an intron 8-inclusion minigene that supports NMD, we demonstrated that mutant

241 this issue of Neuron, Vassalli et al. use OR minigenes that coexpress histochemical markers and show

242 ling in endothelial cells, we have developed minigenes that encode an 11-amino acid C-terminal peptid

243 NMD was studied using minigenes that support NMD.

244 ted in the cDNA construct to generate a CD45 minigene, the LFA-1 promoter was able to drive reproduci

245 A of tRNA cognate to the last triplet of the minigene, thereby limiting protein synthesis for lack of

246 classical memory responses were presented as minigenes, they induced clear memory inflation.

247 For the CFTR minigene this regulation was demonstrated to be dependen

248 We delivered these minigenes through combinations of recombinant Mycobacter

249 ls (LS8) were transfected with an amelogenin minigene to increase amelogenin synthesis, the transfect

250 We used a series of HIV-1 reporter minigenes to demonstrate that Tat's role in splicing is

251 In this study, we screened 366 human miRNA minigenes to determine their effects on the major signal

252 Several recent reports use fluorescence minigenes to image alternative splicing events in living

253 h the aid of p115RhoGEF-RGS, G(12) and G(13) minigenes to inhibit G(12/13), we found that the G(12/13

254 describe a system for targeting proteins and minigenes to lysosomes.

255 Whereas transfection of the G or T minigenes transcribed predominantly normal-sized transcr

256 strongest negative effect on editing of the minigene transcript was located -20 nt 5' to the C targe

257 e transcript could explain why expression of minigene transcripts containing the 27 nt sequence did n

258 cells resulted in an increase in LH2 (long) minigene transcripts, accompanied by a decrease in LH2 (

259 (LDLR) exon 12 in female human liver and in minigene-transfected HepG2 cells.

260 in both in vitro complementation assays and minigene-transfected mouse erythroleukemia cells (MELCs)

261 Using a minigene transfection assay for alternative splicing act

262 Plasmid DNA encoding a FN-C/H-II minigene under control of the cytomegalovirus promoter w

263 c mice have been generated that carry a CD45 minigene under control of the human leukocyte function-a

264 Transfection splicing assays of SMN minigenes under hypoxia revealed that hypoxia-induced sk

265 The human growth hormone (hGH) minigene used for transgene stabilization in mice has be

266 CD45 minigenes using the CD45 5' sequences up to 19 kilobases

267 0-fold increase in the Cpn challenging dose, minigene-vaccinated mice had a 60-fold reduction in lung

268 GD2 cross-reactive IgG antibody responses by minigene vaccination with a protective epitope of GD2 ga

269 chieved for the first time using an oral DNA minigene vaccine against murine vascular endothelial gro

270 s for inclusion in a multiepitope peptide or minigene vaccine construct, truncated recombinant Hsp20

271 this model by demonstrating that an oral DNA minigene vaccine induces effective HLA-A2-restricted, CE

272 This minigene vaccine strategy provides a more flexible alter

273 mice was broken by the CEA(691) (IMIGVLVGV) minigene vaccine.

274 pertussis toxin or cells transfected with a minigene vector for Gi inhibited the ISO-mediated RhoA a

275 ARMS2 transcription from minigene vectors carrying different alleles at variants

276 However, cells transfected with minigene vectors for G12 and G13 did not prevent RhoA ac

277 iven thrombin-induced response, we generated minigene vectors that encode the C-terminal sequence for

278 simple presentation of the Ag in the form of minigene vectors.

279 binant vaccinia virus encoding SSIEFARL as a minigene (VvgB(498-505)).

280 vaccinia virus that expressed SSIEFARL as a minigene (VvgB) and UV-inactivated HSV.

281 ents that influenced Bcl-x splicing, a Bcl-x minigene was constructed.

282 An LH2 minigene was designed, validated, and used in Fox-2 over

283 Further analyses showed that the hGH minigene was expressed in several tissues, also leading

284 nd a keratin-14 (K14) promoter-driven agouti minigene was introduced onto the inverted chromosome 15

285 lternative 5' splice site utilization in the minigene was promoted by RA.

286 Using an LH2 minigene, we have compared the regulation of the alterna

287 Also, using an I-SceI-sensitive HPRT minigene, we show that gene correction is more efficient

288 Using Galpha13 or Galpha12 minigenes, we demonstrated that Galpha13, but not Galpha

289 Using splicing minigenes, we identified multiple exonic and intronic sp

290 Using transfected minigenes, we performed a systematic analysis of the seq

291 DC treated with TAT-minigene were efficiently recognized by both Flu-M1 and

292 patS minigenes were constructed and expressed by different de

293 A variety of P element minigenes were examined in transgenic embryos to determi

294 autologous hFIX cDNA, hFIX minigenes, or hPC minigenes were used as reporters in the expression vecto

295 Minigenes were used to validate the functional effect of

296 lved in 5' splice site selection we cloned a minigene, which included PKCdelta exon 10 and its flanki

297 whereas co-injection of a lumican-expressing minigene with a beta-galactosidase reporter driven by th

298 lacement of the TG dinucleotide tract in the minigene with random sequence abolished splicing of exon

299 the deltaenalphaA promoter was maintained in minigenes with different types of introns and polyadenyl

300 The system allows generation of minigenes within one week.