0 22 Unicellular-unilineage unicellular-unilineage JJ 23 37 erythropoietic erythropoietic JJ 38 46 cultures culture NNS 46 47 : : : 48 57 molecular molecular JJ 58 66 analysis analysis NN 67 69 of of IN 70 80 regulatory regulatory JJ 81 85 gene gene NN 86 96 expression expression NN 97 99 at at IN 100 107 sibling sibling NN 108 112 cell cell NN 113 118 level level NN 118 119 . . . 121 123 In in FW 124 129 vitro vitro FW 130 137 studies study NNS 138 140 on on IN 141 154 hematopoietic hematopoietic JJ 155 162 control control NN 163 173 mechanisms mechanism NNS 174 178 have have VBP 179 183 been be VBN 184 192 hampered hamper VBN 193 195 by by IN 196 199 the the DT 200 213 heterogeneity heterogeneity NN 214 216 of of IN 217 220 the the DT 221 229 analyzed analyze VBN 230 234 cell cell NN 235 246 populations population NNS 246 247 , , , 248 250 ie ie FW 250 251 , , , 252 256 lack lack NN 257 259 of of IN 260 267 lineage lineage NN 268 279 specificity specificity NN 280 283 and and CC 284 297 developmental developmental JJ 298 303 stage stage NN 304 315 homogeneity homogeneity NN 316 318 of of IN 319 339 progenitor/precursor progenitor/precursor NN 340 345 cells cell NNS 346 353 growing grow VBG 354 356 in in IN 357 364 culture culture NN 364 365 . . . 366 368 We we PRP 369 378 developed develop VBD 379 390 unicellular unicellular JJ 391 398 culture culture NN 399 406 systems system NNS 407 410 for for IN 411 421 unilineage unilineage JJ 422 437 differentiation differentiation NN 438 440 of of IN 441 449 purified purify VBN 450 463 hematopoietic hematopoietic JJ 464 474 progenitor progenitor NN 475 480 cells cell NNS 481 489 followed follow VBN 490 492 by by IN 493 501 daughter daughter NN 502 506 cell cell NN 507 515 analysis analysis NN 516 518 at at IN 519 527 cellular cellular JJ 528 531 and and CC 532 541 molecular molecular JJ 542 547 level level NN 547 548 . . . 549 551 In in IN 552 555 the the DT 556 563 culture culture NN 564 570 system system NN 571 579 reported report VBN 580 584 here here RB 584 585 , , , 586 587 ( ( ( 587 588 1 1 LS 588 589 ) ) ) 590 593 the the DT 594 600 growth growth NN 601 607 factor factor NN 608 609 ( ( ( 609 611 GF GF NNP 611 612 ) ) ) 613 621 stimulus stimulus NN 622 629 induces induce VBZ 630 634 cord cord NN 635 640 blood blood NN 641 642 ( ( ( 642 644 CB CB NNP 644 645 ) ) ) 646 656 progenitor progenitor NN 657 662 cells cell NNS 663 665 to to TO 666 677 proliferate proliferate VB 678 681 and and CC 682 702 differentiate/mature differentiate/mature VB 703 714 exclusively exclusively RB 715 720 along along IN 721 724 the the DT 725 734 erythroid erythroid JJ 735 742 lineage lineage NN 742 743 ; ; : 744 745 ( ( ( 745 746 2 2 LS 746 747 ) ) ) 748 752 this this DT 753 767 erythropoietic erythropoietic JJ 768 772 wave wave NN 773 775 is be VBZ 776 789 characterized characterize VBN 790 792 by by IN 793 797 less less JJR 798 802 than than IN 803 804 4 4 CD 804 805 % % NN 806 815 apoptotic apoptotic JJ 816 821 cells cell NNS 821 822 ; ; : 823 824 ( ( ( 824 825 3 3 LS 825 826 ) ) ) 827 837 asymmetric asymmetric JJ 838 847 divisions division NNS 848 851 are be VBP 852 861 virtually virtually RB 862 868 absent absent JJ 868 869 , , , 870 872 ie ie FW 872 873 , , , 874 887 nonresponsive nonresponsive JJ 888 901 hematopoietic hematopoietic JJ 902 913 progenitors progenitor NNS 914 918 with with IN 919 921 no no DT 922 936 erythropoietic erythropoietic JJ 937 946 potential potential NN 947 950 are be VBP 951 957 forced force VBN 958 962 into into IN 963 972 apoptosis apoptosis NN 972 973 ; ; : 974 975 ( ( ( 975 976 4 4 LS 976 977 ) ) ) 978 981 the the DT 982 988 system system NN 989 991 is be VBZ 992 996 cell cell NN 997 1005 division division NN 1006 1016 controlled control VBN 1017 1018 ( ( ( 1018 1021 cdc cdc NN 1021 1022 ) ) ) 1022 1023 , , , 1024 1026 ie ie FW 1026 1027 , , , 1028 1031 the the DT 1032 1038 number number NN 1039 1041 of of IN 1042 1051 divisions division NNS 1052 1061 performed perform VBN 1062 1064 by by IN 1065 1069 each each DT 1070 1074 cell cell NN 1075 1077 is be VBZ 1078 1087 monitored monitor VBN 1087 1088 . . . 1089 1100 Single-cell single-cell JJ 1101 1108 reverse reverse JJ 1109 1133 transcriptase-polymerase transcriptase-polymerase NN 1134 1139 chain chain NN 1140 1148 reaction reaction NN 1149 1150 ( ( ( 1150 1156 RT-PCR RT-PCR NNP 1156 1157 ) ) ) 1158 1166 analysis analysis NN 1167 1170 was be VBD 1171 1178 applied apply VBN 1179 1181 to to TO 1182 1186 this this DT 1187 1194 culture culture NN 1195 1201 system system NN 1202 1204 to to TO 1205 1216 investigate investigate VB 1217 1221 gene gene NN 1222 1232 expression expression NN 1233 1235 of of IN 1236 1243 diverse diverse JJ 1244 1253 receptors receptor NNS 1253 1254 , , , 1255 1262 markers marker NNS 1263 1265 of of IN 1266 1281 differentiation differentiation NN 1281 1282 , , , 1283 1286 and and CC 1287 1300 transcription transcription NN 1301 1308 factors factor NNS 1309 1310 ( ( ( 1310 1314 EKLF EKLF NNP 1314 1315 , , , 1316 1322 GATA-1 GATA-1 NNP 1322 1323 , , , 1324 1330 GATA-2 gata-2 NN 1330 1331 , , , 1332 1335 p45 p45 NN 1336 1341 NF-E2 nf-e2 NN 1341 1342 , , , 1343 1347 PU.1 PU.1 NNP 1347 1348 , , , 1349 1352 and and CC 1353 1361 SCL/Tal1 SCL/Tal1 NNP 1361 1362 ) ) ) 1363 1365 at at IN 1366 1374 discrete discrete JJ 1375 1381 stages stage NNS 1382 1384 of of IN 1385 1399 erythropoietic erythropoietic JJ 1400 1411 development development NN 1411 1412 . . . 1413 1420 Freshly freshly RB 1421 1429 isolated isolate VBN 1430 1437 CD34(+) cd34(+) JJ 1438 1443 cells cell NNS 1444 1453 expressed express VBD 1454 1458 CD34 cd34 NN 1458 1459 , , , 1460 1465 c-kit c-kit NN 1465 1466 , , , 1467 1471 PU.1 PU.1 NNP 1471 1472 , , , 1473 1476 and and CC 1477 1483 GATA-2 gata-2 NN 1484 1487 but but CC 1488 1491 did do VBD 1492 1495 not not RB 1496 1503 express express VB 1504 1508 CD36 CD36 NNP 1508 1509 , , , 1510 1524 erythropoietin erythropoietin NN 1525 1533 receptor receptor NN 1534 1535 ( ( ( 1535 1539 EpoR EpoR NNP 1539 1540 ) ) ) 1540 1541 , , , 1542 1550 SCL/Tal1 SCL/Tal1 NNP 1550 1551 , , , 1552 1556 EKLF EKLF NNP 1556 1557 , , , 1558 1563 NF-E2 NF-E2 NNP 1563 1564 , , , 1565 1571 GATA-1 gata-1 NN 1571 1572 , , , 1573 1575 or or CC 1576 1588 glyocophorin glyocophorin NN 1589 1590 A a NN 1591 1592 ( ( ( 1592 1595 GPA GPA NNP 1595 1596 ) ) ) 1596 1597 . . . 1598 1600 In in IN 1601 1606 early early RB 1607 1609 to to TO 1610 1622 intermediate intermediate VB 1623 1629 stages stage NNS 1630 1632 of of IN 1633 1642 erythroid erythroid JJ 1643 1658 differentiation differentiation NN 1659 1661 we we PRP 1662 1671 monitored monitor VBD 1672 1675 the the DT 1676 1685 induction induction NN 1686 1688 of of IN 1689 1693 CD36 CD36 NNP 1693 1694 , , , 1695 1699 Tal1 Tal1 NNP 1699 1700 , , , 1701 1705 EKLF EKLF NNP 1705 1706 , , , 1707 1712 NF-E2 NF-E2 NNP 1712 1713 , , , 1714 1717 and and CC 1718 1724 GATA-1 gata-1 NN 1725 1729 that that WDT 1730 1739 preceeded preceede VBD 1740 1750 expression expression NN 1751 1753 of of IN 1754 1758 EpoR EpoR NNP 1758 1759 . . . 1760 1762 In in IN 1763 1767 late late JJ 1768 1774 stages stage NNS 1775 1777 of of IN 1778 1787 erythroid erythroid JJ 1788 1798 maturation maturation NN 1798 1799 , , , 1800 1803 GPA GPA NNP 1804 1807 was be VBD 1808 1819 upregulated upregulate VBN 1819 1820 , , , 1821 1828 whereas whereas IN 1829 1833 CD34 cd34 NN 1833 1834 , , , 1835 1840 c-kit c-kit NN 1840 1841 , , , 1842 1846 PU.1 PU.1 NNP 1846 1847 , , , 1848 1851 and and CC 1852 1858 GATA-2 gata-2 NN 1859 1863 were be VBD 1864 1870 barely barely RB 1871 1873 or or CC 1874 1877 not not RB 1878 1886 detected detect VBN 1886 1887 . . . 1888 1890 In in IN 1891 1899 addition addition NN 1899 1900 , , , 1901 1912 competitive competitive JJ 1913 1924 single-cell single-cell JJ 1925 1931 RT-PCR rt-pcr NN 1932 1935 was be VBD 1936 1940 used use VBN 1941 1943 to to TO 1944 1949 assay assay VB 1950 1954 CD34 cd34 NN 1955 1959 mRNA mrna NN 1960 1971 transcripts transcript NNS 1972 1974 in in IN 1975 1982 sibling sible VBG 1983 1990 CD34(+) cd34(+) JJ 1991 1998 CD38(-) cd38(-) JJ 1999 2004 cells cell NNS 2005 2020 differentiating differentiate VBG 2021 2023 in in IN 2024 2034 unilineage unilineage JJ 2035 2044 erythroid erythroid JJ 2045 2053 cultures culture NNS 2053 2054 : : : 2055 2059 this this DT 2060 2068 analysis analysis NN 2069 2076 allowed allow VBD 2077 2079 us us PRP 2080 2082 to to TO 2083 2097 semiquantitate semiquantitate VB 2098 2101 the the DT 2102 2109 gradual gradual JJ 2110 2124 downmodulation downmodulation NN 2125 2127 of of IN 2128 2132 CD34 CD34 NNP 2133 2137 mRNA mRNA NNP 2138 2142 from from IN 2143 2153 progenitor progenitor NN 2154 2159 cells cell NNS 2160 2167 through through IN 2168 2173 their their PRP$ 2174 2189 differentiating differentiate VBG 2190 2199 erythroid erythroid JJ 2200 2207 progeny progeny NN 2207 2208 . . . 2209 2211 It it PRP 2212 2214 is be VBZ 2215 2224 concluded conclude VBN 2225 2229 that that IN 2230 2234 this this DT 2235 2240 novel novel JJ 2241 2248 culture culture NN 2249 2255 system system NN 2255 2256 , , , 2257 2264 coupled couple VBN 2265 2269 with with IN 2270 2281 single-cell single-cell JJ 2282 2288 RT-PCR RT-PCR NNP 2289 2297 analysis analysis NN 2297 2298 , , , 2299 2302 may may MD 2303 2312 eliminate eliminate VB 2313 2316 the the DT 2317 2328 ambiguities ambiguity NNS 2329 2338 intrinsic intrinsic JJ 2339 2341 to to TO 2342 2351 molecular molecular JJ 2352 2359 studies study NNS 2360 2362 on on IN 2363 2376 heterogeneous heterogeneous JJ 2377 2388 populations population NNS 2389 2391 of of IN 2392 2405 hematopoietic hematopoietic JJ 2406 2428 progenitors/precursors progenitors/precursor NNS 2429 2436 growing grow VBG 2437 2439 in in IN 2440 2447 culture culture NN 2447 2448 , , , 2449 2461 particularly particularly RB 2462 2464 in in IN 2465 2468 the the DT 2469 2476 initial initial JJ 2477 2483 stages stage NNS 2484 2486 of of IN 2487 2498 development development NN 2498 2499 . . .