遗传学报
遺傳學報
유전학보
ACTA GENETICA SINICA
2003年
3期
215-220
,共6页
雷蕾%刘忠华%王鸿%寇朝辉%吴昱琪%徐营%程勇%朱子玉%夏国良%陈大元
雷蕾%劉忠華%王鴻%寇朝輝%吳昱琪%徐營%程勇%硃子玉%夏國良%陳大元
뢰뢰%류충화%왕홍%구조휘%오욱기%서영%정용%주자옥%하국량%진대원
体细胞%核移植%细胞代数%小鼠
體細胞%覈移植%細胞代數%小鼠
체세포%핵이식%세포대수%소서
somatic cells%nuclear transfer%number of passages%mice
为探讨体细胞来源及培养代数对核移植重构胚发育的影响,实验采用电融合法将小鼠2-细胞胚胎卵裂球、胚胎干细胞(ES)、胎儿成纤维细胞、耳成纤维细胞、尾尖成纤维细胞、睾丸支持细胞和精原细胞以及不同培养代次的胎儿成纤维细胞进行了核移植.结果显示:2-细胞胚胎卵裂球供核重构胚发育最好,囊胚率为7.4%;ES细胞重构胚虽然发育率低,但仍有囊胚出现,比例为0.7%;胎儿成纤维细胞重构胚最高发育阶段为桑椹胚,比例为0.2%;精原细胞重构胚只能发育到8-细胞阶段,比例为0.3%;其他几类细胞重构胚则仅能发育至4-细胞阶段.不同培养代数的胎儿成纤维细胞重构胚除第3代外都可发育到8-细胞阶段,且发育率差异不显著,但第一代细胞重构胚2-细胞发育率(40.7%)显著低于2、3和4代细胞重构胚.结果表明:不同分化程度的细胞核移植后,重新编程的难易程度是不一样的,分化程度越高则重新编程越难;未调整细胞周期的ES细胞由于多数处于S期,所以重构胚发育率很低;体外培养传代有利于体细胞核移植后重新编程.
為探討體細胞來源及培養代數對覈移植重構胚髮育的影響,實驗採用電融閤法將小鼠2-細胞胚胎卵裂毬、胚胎榦細胞(ES)、胎兒成纖維細胞、耳成纖維細胞、尾尖成纖維細胞、睪汍支持細胞和精原細胞以及不同培養代次的胎兒成纖維細胞進行瞭覈移植.結果顯示:2-細胞胚胎卵裂毬供覈重構胚髮育最好,囊胚率為7.4%;ES細胞重構胚雖然髮育率低,但仍有囊胚齣現,比例為0.7%;胎兒成纖維細胞重構胚最高髮育階段為桑椹胚,比例為0.2%;精原細胞重構胚隻能髮育到8-細胞階段,比例為0.3%;其他幾類細胞重構胚則僅能髮育至4-細胞階段.不同培養代數的胎兒成纖維細胞重構胚除第3代外都可髮育到8-細胞階段,且髮育率差異不顯著,但第一代細胞重構胚2-細胞髮育率(40.7%)顯著低于2、3和4代細胞重構胚.結果錶明:不同分化程度的細胞覈移植後,重新編程的難易程度是不一樣的,分化程度越高則重新編程越難;未調整細胞週期的ES細胞由于多數處于S期,所以重構胚髮育率很低;體外培養傳代有利于體細胞覈移植後重新編程.
위탐토체세포래원급배양대수대핵이식중구배발육적영향,실험채용전융합법장소서2-세포배태란렬구、배태간세포(ES)、태인성섬유세포、이성섬유세포、미첨성섬유세포、고환지지세포화정원세포이급불동배양대차적태인성섬유세포진행료핵이식.결과현시:2-세포배태란렬구공핵중구배발육최호,낭배솔위7.4%;ES세포중구배수연발육솔저,단잉유낭배출현,비례위0.7%;태인성섬유세포중구배최고발육계단위상심배,비례위0.2%;정원세포중구배지능발육도8-세포계단,비례위0.3%;기타궤류세포중구배칙부능발육지4-세포계단.불동배양대수적태인성섬유세포중구배제제3대외도가발육도8-세포계단,차발육솔차이불현저,단제일대세포중구배2-세포발육솔(40.7%)현저저우2、3화4대세포중구배.결과표명:불동분화정도적세포핵이식후,중신편정적난역정도시불일양적,분화정도월고칙중신편정월난;미조정세포주기적ES세포유우다수처우S기,소이중구배발육솔흔저;체외배양전대유리우체세포핵이식후중신편정.
In order to study the effects of different donor cells and passages on development of nuclear transfer embryos,we constructed embryos by electrofusing several kinds of donor cells into enucleated MⅡ oocytes from Kun Ming (KM) mouse.These cells include 2-cell embryonic blastomeres,KMW embryonic stem (ES) cells,fetal fibroblast,ear fibroblast,tail tip fibroblast,sertoil cells and spermatogonia.Meanwhile,we compared the effects of passage numbers of fetal fibroblast cells on developmental competency after nuclear transfer.We found that 7.4% of reconstructed embryos from 2-cell embryonic blastomeres and 0.7% from ES cell could develop to blastocyst in vitro;embryos from fetal fibroblast could only develop to morula stage with the rate of 0.2%;embryos from spermatogonia could only develop to 8-cell stage and the rate was 0.3%;embryos respectively from ear fibroblast,sertoli cell and tail tip fibroblast could only develop to 4-cell stage.Although 2-cell development rate of embryos reconstructed from fetal fibroblast in first passage was significantly lower than those from the 2nd,the 3rd and the 4th passage,embryos from different passages could develop to 8-cell stage except the 3rd passage.The result indicated that it is more difficulty for terminally differentiated cell nuclei to be reprogrammed in enucleated MⅡ oocytes than for low differentiated cell nuclei.The reason of low development rate from ES cells maybe that most of ES cells was at S stage of the cell cycle,which out of coordination with MⅡ oocytes.We could conclude that culture and passage of donor cells might be benefit to nucleus reprogramming.
